UNITED STATES OF
AMERICA
+ + + + +
FOOD AND DRUG
ADMINISTRATION
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TRANSMISSIBLE SPONGIFORM
ENCEPHALOPATHIES
ADVISORY COMMITTEE
(TSEAC)
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BETHESDA, MARYLAND
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WEDNESDAY,
JULY 17, 2003
This transcript has not been edited
or corrected, but appears as received
from the commercial transcribing
service. Accordingly, the Food
and
Drug Administration makes no
representation as to its accuracy.
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The
Advisory Committee met in the Versailles Room at the Holiday Inn Select, 8120
Wisconsin Avenue, Bethesda, Maryland 20814, at 8:00 a.m., with Suzette A.
Priola, Ph.D., Chair, presiding.
PRESENT:
SUZETTE A. PRIOLA, Ph.D., Chair
JOHN C. BAILAR, III, M.D., Ph.D., Member
ARTHUR W. BRACEY, M.D., Member
LISA A. FERGUSON, D.V.M., Member
PIERLUIGI GAMBETTI, M.D., Member
R. NICK HOGAN, M.D., Ph.D., Member
RICHARD T. JOHNSON, M.D., Member
RIMA F. KHABBAZ, M.D., Member
SIDNEY M. WOLFE, M.D., Member
CHARLES E. EDMISTON, JR., Ph.D., Temporary Voting
Member, Topics , 3 & 4
KENRAD E. NELSON, M.D., Temporary Voting Member, Topics 2, 3 & 4
TERRY V. RICE, Temporary Voting Member,
Topics
2, 3 & 4
DAVID F. STRONCEK, M.D., Temporary Voting Member,
Topics 2, 3 & 4
SHIRLEY J. WALKER, Consumer Representative
STEPHEN R. PETTEWAY, JR., Ph.D., Non-Voting
Industry Representative
SHEILA D. LANGFORD, Staff
ALSO PRESENT:
DR. DAVID M. ASHER, OBRR, CBER, FDA
DR. STANLEY BROWN, CDRH
DR. YUAN-YUAN CHIU, CDER, FDA
DR. MICHAEL DUNN, V. Pres., Chairman of the
Regulatory Committee, GMIA
DR. JAY EPSTEIN, Director, OBRR, CBER, FDA
DR. ROBERT HILLS, Health Canada, Ottawa
DR. GEORGE MASSON, President GMIA
DR. TERRY MORRIS, APHIS
DR. PEDRO PICCARDO, CJD
DR. MORRIE POTTER, CFSAN, FDA
CAPTAIN EDWARD RAU, Environmental Health Officer,
NIH
DR. RON ROGERS, Health Canada, Ottawa
DR. ROBERT ROHWER, Director Molecular
Neuro-Virology Unit, VA Medical
Center, Baltimore
DR. WILLIAM RUTALA, UNC
REINHARD SCHRIEBER, Chief Manufacturing Officer, Deutsche Gelatine
DR. ROBERT SOMERVILLE, IAH Edinburgh, UK
FABRIKEN STOESS, AG, Gelita Group
DR. DAVID TAYLOR, SEDECON 2000, UK
NELSON BROOKLANG, Ortech International
DANIEL R. DWYER, ESQ., Kleinfeld, Kaplan &
Becker, Counsel to GME
CHARLES FILLBURN, Nutranax Laboratories
PAUL HAFFENDEN, TerraCell
MERLYN SAYERS, M.B., B.Ch., Ph.D., Carter
BloodCare
WAYNE E. VAZ, Serologicals Corporation
AGENDA ITEM PAGE
OPENING/INTRODUCTION:
William
Freas............................. 6
CONFLICT OF INTEREST STUDY:
Katherine
McComas........................ 11
TOPIC 1 - SAFETY OF BOVINE BONE GELATIN:
Background
& Introduction:
Morrie
Potter...................... 13
Questions
to the Committee:
Yuan-Yuan
Chiu..................... 18
Market
Trend in the U.S.:
George
Masson...................... 21
Questions/Comments:...................... 27
Manufacturing
Process for Bone Gelatin in U.S.:
Michael
Dunn....................... 39
Questions/Comments:...................... 54
Manufacturing
Process Bone Gelatin in Europe:
Reinhard
Schrieber................. 60
Questions/Comments:...................... 74
GME
Validation Studies of Bone Gelatin:
Robert
Somerville.................. 75
Questions/Comments:...................... 94
Gelatin
Manufacturing Process:
Robert
Rohwer..................... 101
Questions/Comments:..................... 116
Risk
Analysis of Infectivity:
Ron
Rogers........................ 120
Questions/Comments:..................... 135
USDA
Gelatin Policy:
Terry
Morris...................... 136
AGENDA ITEM PAGE
Questions/Comments:..................... 142
Open
Public Hearing:
William
Freas..................... 144
Daniel
Dwyer...................... 145
Committee
Discussion & Voting:.......... 150
TOPIC 2 - BSE IN CANADA:
Potential
Exposure of Blood Donors in
North
America to the BSE Agent:
Jay
Epstein....................... 192
Questions/Comments:..................... 193
Review
of BSE in Canada:
Robert
Hills...................... 195
Questions/Comments:..................... 210
Open
Public Hearing:
Wayne
Vaz......................... 222
Merlyn
Sayers..................... 229
Questions/Comments:..................... 236
Committee
Discussion:................... 238
TOPICS 3 & 4 - TSEs AND DECONTAMINATION OF
MEDICAL EQUIPMENT AND FACILITIES:
TSEs,
Decontamination & FDA Regulated Products:
David
Asher....................... 241
Principles
of TSE Inactivation:
Robert
Rohwer..................... 251
Questions/Comments:..................... 269
Basis
for WHO Recommendations:
David
Taylor...................... 270
Questions/Comments:..................... 289
AGENDA ITEM PAGE
Reducing
Risk of CJD Transmission Through Surgical
Procedures: Experience in UK:
Philippa
Edwards (Pedro Piccardo). 299
Questions/Comments:..................... 311
TSE
Agents/Infection Control in USA Hospitals:
William
Rutala.................... 315
Questions/Comments:..................... 330
Infectivity
of Air Emissions & Residues from
Simulated
Incineration of Scrapie Tissues:
Edward
Rau........................ 338
Questions/Comments:..................... 350
TSE
Infectivity: Experience With Models for
Validating
Decontamination of Surfaces:
Stanley
Brown..................... 351
David
Asher....................... 361
Questions/Comments:..................... 375
ADJOURN:
Suzette
Priola.......................... 379
P-R-O-C-E-E-D-I-N-G-S
8:03
a.m.
SECRETARY
FREAS: Dr. Priola, members of the
public, invited guests and public participants, I would like to welcome all of
you to this our 14th meeting of the Transmissible Spongiform
Encephalopathies Advisory Committee. I
am Bill Freas. I am the executive
secretary for this Committee. At this
time, I would like to go around and introduce to you the members at the head
table, starting on the right hand side of the room.
The
first chair will soon be occupied very shortly by Dr. Pierluigi Gambetti. He is a professor and director Division of
Neuropathology Case, Western Reserve University. Okay. Then the second
chair will soon be occupied by Dr. Richard Johnson, professor of neurology at
Johns Hopkins University. And then
going around the table, the people who are here, Dr. Arthur Bracey, associate
chief Department of Pathology, Saint Lukes Episcopal Hospital. Next is Dr. Lisa Ferguson, a senior staff
veterinarian, U.S. Department of Agriculture.
Next
is Dr. Nick Hogan, associate professor of ophthalmology, University of Texas,
Southwestern Medical School. Next is
Dr. Rima Khabbaz, associate director for Epidemiologic Science, National Center
for Infectious Diseases, Atlanta, Georgia.
Around the corner of the table is a gentleman, whom I'm going to ask to
join us at lunch time, if that is okay, Dr. Nelson. Could I ask you to join us at lunch time instead of in the
morning?
DR.
NELSON: Certainly.
SECRETARY
FREAS: This is my mistake. I apologize. Dr. Nelson will be a temporary voting member, and he will join us
right at lunch time, and if you could just sit over in the FDA section up until
Topic 1 is over. And when I read the
Conflict of Interest statement, hopefully, that will be explained. My apologies for not checking before I started. Okay.
Next
is our Chair, Dr. Suzette Priola. She
is an investigator of Laboratory of Persistent and Viral Diseases of the Rocky
Mountain Laboratories. Next is our
consumer representative, Ms. Shirley Walker, vice president of the Health and
Human Services, Urban League of Greater Dallas in north central Texas. Next is Dr. Sidney Wolfe, director of Public
Citizen Health Research Group, Washington, D.C.
Next
is Dr. John Bailar, professor in America's University of Chicago. Next is our non-voting industry
representative, Dr. Stephen Petteway, director of Pathogen Safety and Research,
Bayer Corporation. Three Committee
members in addition to the two that are joining us shortly could not be with us
at all for this meeting. They are Mr.
Val Bias, consumer representative, Lynn Creekmore, staff veterinarian and Dr.
Stephen DeArmond from the University of California.
I
would now like to read into the public record the Conflict of Interest
statement for this meeting. "The
following announcement is made part of the public record to preclude even the
appearance of a Conflict of Interest at this meeting. Pursuant to the authority granted under the Committee Charter,
the Director Center for Biologics Evaluation and Research has appointed Mr.
Terry Rice and Drs. Kenrad Nelson, who I just asked to leave the table, and
David Stroncek as temporary voting members for Topics 2, 3 and 4 of this
meeting.
In
addition, the associate commissioner of External Relations of FDA has appointed
Dr. Charles Edmiston as a temporary voting member for Topics 2, 3 and 4 of this
meeting. Based on the agenda, it has
been determined that the Committee will not be providing advice on specific
firms or products at this meeting. The
topics deemed discussed by the Committee in open session are considered general
matters issues.
To
determine if Conflicts of Interest exist, the Agency reviewed the agenda and
all relevant reported financial interests from meeting participants. The Food and Drug Administration prepared
general matters waivers for special Government employees, who required a waiver
under 18 U.S. Code 208. Because general
matters topics impact on so many entities, it is not prudent to recite all
potential Conflicts of Interest as they apply to each member.
FDA
acknowledges that there may be potential Conflicts of Interest, but because of
the general nature of the discussion before the Committee, these potential
conflicts are mitigated. We would like
to note for the record that Dr. Stephen Petteway is serving as a non-voting
industry representative member for this Committee. He is employed by Bayer and thus has interests in his employer
and other similar firms.
Listed
on the agenda are speakers making industry presentations and/or updates. These speakers have financial interests
associated with their employer and with other regulated firms. These speakers were not screened for these
Conflicts of Interests. With regard to
FDA's invited guest speakers, that's all other speakers, except those from
industry, the Agency has determined that the services of these speakers are
essential.
The
following interests are being made public to allow the meeting participants to
objectively evaluate their presentations and comments that they may make. Dr. Robert Rohwer has disclosed he has
financial interest with various firms that could be affected by the Committee
discussions. Dr. William Rutala
receives consultant fees from several firms that could be affected by the
Committee discussions. Dr. Robert
Somerville has research supported by the Gelatin Manufacturers of Europe. His expenses to this meeting were also paid
by the Gelatin Manufacturers of Europe.
Dr. Charles Weissmann holds patents related to Prion Disease work.
Members
and consultants are aware of the need to exclude themselves from the
discussions involving specific products or firms which they have not been
screened for the Conflict of Interest.
Their exclusion will be noted in the public record. With respect to all other meeting
participants, we ask, in the interest of fairness, that they address any
current or previous financial involvement with any firm whose product they may
wish to comment upon. Waivers may be
available by written request to the Freedom of Information Office."
That's
the end of the Conflict of Interest statement.
I do ask that throughout this meeting before it starts if you would
check your cell phone or your pager and, please, put it in the silent mode, so
it won't disrupt those people sitting next to you.
Next,
the FDA is continually trying to improve its Advisory Committee Program and to
reduce any perceived Conflicts of Interest.
It has asked Dr. Katherine McComas from the University of Maryland to
conduct a survey of this program, and I would like to give her the opportunity
to tell us how we can help her with this survey and how the survey is being
conducted. Dr. McComas, either
place. Keep talking and they'll turn
the mike volume up.
DR.
MCCOMAS: Okay. Good morning and thank you. I'm Katherine McComas and I'm a faculty member
at the University of Maryland, and I'm here today to conduct a study of what
people know and understand about the Conflict of Interest procedures that the
FDA uses to monitor real or potential Conflicts of Interest of its Advisory
Committee members. This is a study that
is being conducted across multiple meetings.
This is the 11th meeting I have attended across the centers
at FDA, including CBER.
For
those of you in the audience, I've distributed a questionnaire on your chair
and I have also distributed a different questionnaire to the Advisory Committee
members. If you have an opportunity
today to complete this questionnaire or tomorrow, there is a box on the
registration desk where you can drop it.
Otherwise, there is a business reply envelope that you can just drop it
in the mail as soon as you can. Your
participation is voluntary, but it is important. The more responses we get, the better we are to provide feedback
to the FDA about what people know and understand about the Conflict of Interest
procedures, and what may be done to improve satisfaction, if necessary, with
the Advisory Committee process.
Again,
I appreciate your participation and if you have any questions, my contact
information is included in the letter, in the questionnaire and I would be
happy to provide any answers. Also,
when the study is done, the responses will be available in summary form to
everyone who is interested. So thank
you very much for your time and have a great day.
SECRETARY
FREAS: So if you got here early and did
not receive a questionnaire, the questionnaires are on the table outside and,
please, everybody on the FDA staff will be more than glad to help you if you
have any questions with this questionnaire.
Dr.
Priola, I turn the microphone over to you to start the meeting. Thank you.
DR.
PRIOLA: So since we have a very full
agenda today, we will just get started with the first speaker, who is Dr.
Potter, who will give us background on Topic 1.
DR.
POTTER: Good morning. FDA has been considering the safety of
gelatin with regard to BSE for a number of years, and has come to this
Committee on a number of occasions to get its recommendations on FDA's guidance
to gelatin manufacturers and users. The
safety of gelatin is determined as you've told us before by the safety, the
source materials in the degree to which the gelatin manufacturing process
destroys prions that enter the system.
Questions
to the Committee have dealt with these two factors and how well knowledge about
TSE's was reflected in FDA guidance for assuring the safety of gelatin for food
and cosmetic use. Before 1996, FDA did
not include gelatin within its recommended restrictions concerning bovine
ingredients in FDA regulated products.
In 1996, FDA began to review its position on gelatin in light of new
information that associated BSE exposure with Variant CJD in humans and new
data from a study on the effect on infectivity of gelatin processing that
suggested only partial effectiveness.
In
1997, this Committee met to consider the safety of gelatin and to provide an
assessment on the overall risk to humans associated with imported gelatin. This Committee made the following
observations: First, that the
scientific information available no longer justified excepting gelatin from
restrictions recommended by FDA for other bovine derived materials from BSE
countries. Second, that bovine gelatin
injected or implanted forms posed a higher risk of transmitting BSE to humans
than gelatin that is ingested. Third,
that brains and spinal cords from cattle from BSE countries should be excluded
from raw materials used to produce gelatin for human consumption. Fourth, alkaline or acid processing in
gelatin manufacturer may only reduce rather than eliminate BSE infectivity, and
the Committee called for better validation studies, particularly to investigate
the other steps of gelatin manufacture.
And finally, that porcine gelatins appear to pose no known risk of transmitting
TSEs to humans.
After
the 1997 TSEAC meeting, FDA issued its gelatin guidance document which remains
the current FDA position in policy on the production of gelatin. In this guidance, FDA proposed the following
recommendations concerning the acceptability of gelatin in FDA regulated
products intended for human use. First,
that importers, manufacturers and suppliers should determine the tissue,
species and country source of all materials used in processing gelatin for
human use.
Second,
that bone and hides from cattle from any source country that show signs of
neurologic disease should not be used as raw materials. Third, gelatin production from bones and
hides obtained from cattle that reside in BSE countries or countries that do
not meet the latest BSE related OIE standards should not be used in injectable,
ophthalmic or implanted FDA regulated products or in their manufacture, but may
be used in FDA regulated products for oral consumption and cosmetic use by
humans if the cattle come from BSE-free herds and if the slaughter house
removes heads, spines and spinal cords directly after slaughter.
Fourth,
gelatin produced from bovine hides from any source country may be used in FDA
regulated products for oral consumption and cosmetic use by humans if
processors insure that the hides have not been contaminated with brain, spinal
cord or ocular tissues of cattle residing in or originating from BSE
countries. Fifth, gelatin produced from
bovine hides and bones may be used in FDA regulated products for human use if
the gelatin is produced from raw materials from countries like the United
States that observe OIE standards and have not diagnosed BSE in their national
cattle herd, that is RBSE-free. And
finally, gelatin produced from porcine skins from any source country may be
used in FDA regulated products for human use.
In
1998, this Committee met again to discuss gelatin among other issues. FDA's guidance, based on the 1997 TSEAC
recommendations, was presented to the Committee to consider several new pieces
of relevant information. For example,
the infectivity of dorsal root ganglia and low level infectivity in bone marrow
and the growing number of BSE cases being discovered in Europe. The Committee considered this new
information and decided gelatin could be safely sourced from bones and hides of
cattle in BSE countries as long as the recommendations in the guidance were
met. That is that the cattle came from
BSE-free herds and the high-risk materials were removed after slaughter.
And
this is at present the status of the safe source factor for gelatin. Continuing on with the other key factor,
that of validated effectiveness in the manufacturing process, in June 2001, the
Committee was given an update from the Gelatin Manufacturers of Europe on the
interim validation study results on the inactivation of BSE through the gelatin
manufacturing process. This was an
information sharing meeting only and no questions were posed to the Committee.
The
Committee reviewed the study design and the preliminary data and requested a
presentation of the final results as soon as they were available. The Committee is now about to get its wish
as GME will present their completed studies, and we will hear other marketing
and manufacturing information on gelatin in North America and Europe. After you have heard this new information,
we would like you to comment on the studies and to consider the current gelatin
guidance in light of these completed studies and other relevant information.
And
I think, according to my schedule, Yuan-Yuan will now charge up the
Committee. Thank you.
CHAIR
PRIOLA: Thank you, Dr. Potter. Dr. Chiu will now present the questions for
Committee.
DR.
CHIU: Good morning. First, I would like to thank Dr. Priola and
the Committee members to take the time to come out and also we have sent you a
huge package, gelatin studies protocols and procedures and the results. We appreciate how much time you need to
really review those studies. In the
early days, in the 1998 year, when the Agency and the Committee together made a
decision for the Agency's recommendation on gelatin was based on previous study
which the Committee thought was somewhat flawed.
So
generally, this reason follows the advice of the Committee to then redesign the
studies and then today, you know, we have the new study results. You did not review the interim results, but
today we have the final results off of five studies. We're hoping, you know, with the presentation today and the
background information you have you will be able to help the Agency to answer
two questions.
Next
slide, the first question is "Do those results of these new studies
demonstrate a reduction in infectivity that is sufficient to protect human
health?" And we are only limited
to hear the question to bovine bone gelatin is consumed by humans through oral
or topical administration. The question
is not for gelatin of other administrations, such as the injection, you know,
implantable. We would like the
Committee to focus on oral and topical administration.
Next
slide, now, the first question, you know, the answer could be yes or no or in
between regardless, you know, the answer we also would like you to answer the
second question. There are two
parts. The first part is "Do the
scientific data and the information available support the current FDA
recommendations on bovine bone gelatin for oral and topical
administration?"
The
current recommendations, next slide, is on this slide. The general policy of FDA is for FDA
regulated products, the bovine derived material should come from cattle not
bone residing as slaughtered in BSE countries, but the Agency also provides
some exemptions. The exemption could be
a total exemption unconditional, such as milk, dairy products and the milk
derived product. But some of the
substances, you know, the Agency provide conditional exemptions, and the
gelatin for oral and topical use are giving conditional exemptions.
So
if the cattle actually is coming from BSE countries, then that condition is the
cattle must be from a BSE-free herd and also at the slaughter house the head,
the spine and spinal cord should be removed.
And this is from BSE countries.
Now, some countries may not have BSE cases, but there is consider of
high-risk of BSE. Then the
recommendation is the heads, the spine and the spinal cord should be removed as
the first step in the slaughter house.
So the first question is whether this current recommendation still is
valid, based on the scientific information we have today.
Next
slide, if the answer is yes, then that's the end of it. If the answer is no, then we would like to
know what changes the Committee would like to recommend to our current
policy. The changes can be in all
different directions. You may consider
we can actually grant a total exemption to the gelatin for oral and topical use
or you may consider to modify the current recommendation the FDA has, either by
strengthening or by relaxing the conditions.
So we are anxious and grateful you will give Agency your
deliberation. Thank you.
CHAIR
PRIOLA: Okay. Thank you, Dr. Chiu. Our
next speaker is Mr. Masson, who will discuss market trends in the U.S.
DR.
MASSON: Yes, good morning
everybody. Madam Chairman, I would just
like to thank the Committee and the FDA, in particular, about the opportunity
to address the Committee. As we have
heard from Drs. Potter and Chiu, it has been a long and winding road, the saga
of gelatin, and we hope today that we can reach a satisfactory conclusion and
see gelatin taken off the file, so to speak, having reassured you of its
safety.
My
first slide, please. Can I have the
first slide, please? Okay. Thank you.
Well, just an introduction of I'm currently the president of our
industry association, the GMIA, and also president and CEO of one of its
members at Russelot. The next
slide? A bit of history as to GMIA and
credentials, so to speak. Our
association was formed in 1956. We have
six members, all NAFTA based, four from the U.S., one in Mexico and one in
Canada. And we've listed here the
typical working committees by which we run the institute. There is no particular order of precedence,
but the technical and regulation committees as you can imagine, indeed, are the
primary focus of most of our work, I guess.
Next,
in terms of what we represent, we, as you see, represent roughly 22 percent of
the global gelatin production, and almost 100 percent of all the gelatin made
in North America. And, indeed, three of
our members are also affiliates of the Gelatin Manufacturers of Europe. And I should have added actually that one of
the other members is an affiliate of the Japanese Gelatin Manufacturing Group.
Next,
please. This lists our objectives. As you see, we try to monitor and inform our
members of any and all regulations which can impact gelatin. We are the liaison with FDA, USDA and other
regulatory authorities, and we gather and distribute technical information to
our members, endeavor to promote a broader knowledge of gelatin and encourage
its wider consumption. And we provided
the forum as you've seen from our committee information on all of the major
aspects concerning technical, environmental and safety issues.
And
as time has gone by and as other industry associations have been formed around
the world in Japan, South America and so on, a major function which has emerged
has been to liaise with them to ensure that technical information and
regulation information, etcetera, is shared with the other associations around
the world. And I just participated, for
instance, in the Japanese meeting or the Asia Pacific meeting, which was held
in Japan, just last month, as an example of the increasing international
corporation among the industry associations.
Next,
please. This slide lists the primary
uses of the concentrated, obviously for today's purposes, bovine bone gelatin
and, as you see here, this is a list of the major uses for bovine bone gelatin
in the United States. It is listed in
the standing order of use with photographic still being the largest consumer
going on down to food. There's less and
less bovine bone gelatin used in food products, by food we mean confectionery
and marshmallows or whatever else.
It
is being, I guess, more replaced there by pigskin porcine gelatins. But anyway, those are the primary uses and,
as you see, no matter what the end use, all the gelatins are produced through
the same manufacturing processes and my colleagues will be describing those in
some detail in a few minutes.
Next,
please. To give you an idea of the
scale of the gelatin business globally, and in particular the bovine part of
that production, we have listed here the various theaters, so to speak. Europe is still the biggest gelatin
producer. Significantly so with 117,000
tons out of a total of some 270,000 tons around the world. Above that, I would have to say, over 25
percent or so is actually bone gelatin.
The U.S. in total we make something like 60,000 tons. And again, these are all gelatins, whether
bovine or porcine or bovine hide or bovine bone, porcine skin.
And
as you see, of the 60,000, about 17,000 tons is actually bone gelatin. Other covers is Asia Pacific, the Asia
Pacific regions and South America, and you see about a third of their gelatin
is of bovine bone origin. So totally,
bovine bone represents almost 80,000 out of the total of 270,000 tons. And to give you an idea again of this
international value industry total globally is not that big. It's 1.5 billion dollars equivalent.
Next,
please. What we tried to do here is to
put to ourselves a few questions, the elimination of which, I think, will be
helpful to the committee in looking at bovine bone gelatin, in particular, in
the U.S. The first question, as you can
see is, "Can the U.S. gelatin industry supply total U.S. capsule
industry's needs?" The answer is
no. And this illustrates how that is
the case.
As
you saw earlier, the U.S. bovine bone gelatin production totals some 17,000
tons, but of that 11,500 are needed for photographic and other non-capsule
uses. So you see that that remains,
there remains only about 5,500 tons which can be used by the capsule industry,
but the total needs, in fact, are 10,000 tons, and this means that the
shortfall roughly 4,500 tons of bovine bone gelatin has to be imported and they
come primarily from Europe, also from Japan and India, and immediately derived
nevertheless from U.S. bones or from bones from other countries.
Next,
just continuing that theme, of those 4,500 tons of which we need to import,
"Can they be derived solely from U.S. bones, even if it's not actually
manufactured in the U.S.?" Again,
the answer is no. As you see here, the
total amount of U.S. bones which are made available to the gelatin business is
roughly 130,000 tons and because of its use in photographic production, whether
in Europe or in the U.S., and also for manufacturing bovine bone gelatin by
other companies outside of the U.S., the amount remaining available for
pharmaceutical gelatin production here is only 28,000 tons.
The
next line which is in bold print illustrates that we need roughly 6 tons of
bones to make a ton of gelatin. So that
the 10,000 tons of gelatin, which the capsule industry needs, is actually
equivalent to 60,000 tons of bovine bones and, consequently, you see the
shortfall here is roughly 32,000 tons, so to speak, to be able to make all of
the capsule industry requirements strictly from U.S. bones. So in other words, the deficit has to be
sourced from bone suppliers outside of the United States.
Next,
so then I apologize, this is a little bit of a busy slide, but the bottom line
is that there are, indeed, other sources outside of the United States, but even
though the quantity is maybe available for various reasons in terms of
surveillance, the inspection procedures and so on, it's not so obvious that the
quantities, the tonnages, which are listed in the second line or the second
section are, indeed, available and because of the various restrictions and so
on, you see, in fact, that the bone and that those numbers diminished to rather
smaller numbers.
And
this really drives to the heart of the matter.
This is the crux really of what we want to get at today and my
colleagues will be addressing this individually and then in the public comment
session later, the question of how we can determine the BSE status adherents
and also the question as Dr. Chiu referred to of just when they have to be
removed in the gelatin bone process.
Again, we'll be traveling to that in some much more detail in later
presentations.
Next,
please. I guess that concludes my
presentation, unless there are any questions.
CHAIR
PRIOLA: Yes, are there any questions
for Mr. Masson?
DR.
MASSON: Thank you.
CHAIR
PRIOLA: Oh, Dr. Hogan?
BOARD
MEMBER HOGAN: I have one question.
CHAIR
PRIOLA: Just a second, Mr. Masson,
there's a question.
BOARD
MEMBER HOGAN: Sorry. I had one question, perhaps it is contained
in this information you provided us, which is quite huge. In terms of the amount of gelatin that is
derived from Europe, could you tell us something about the country breakdown,
that is it's most from the UK, France, Switzerland, etcetera?
DR.
MASSON: Yes, I think you'll find in the
information packet there is a detailed breakdown of the various imports. The consumption in the U.S., actually the
total marketing, is closer to 80,000 tons.
And as you saw, we make 60,000.
There is a net import/export situation.
The U.S. does export gelatin, but basically to get to the 80,000 that we
need, we need effectively a net import of 20,000 tons.
Those
20,000 tons come from quite a variety of countries and, indeed, they are listed
in the information packet. We didn't go
into the detail of it here, because it's somewhat difficult to differentiate,
certainly differentiate country by country.
It's a little bit more difficult to differentiate within certain
countries whether it is bovine gelatin or porcine gelatin, which is actually
being imported. But basically the
primary countries who do export into the States would be France, Germany, not
so much any more from UK, for obvious reasons, Brazil, Argentina, Japan,
India. Those would represent the large
majority of the total import picture.
And
again, the variety of gelatins some of that is bovine bone, for sure, but also
a lot of bovine hide gelatin comes, for instance, from South America, and
bovine bone also from India. It's quite
a variety of types from those principle countries.
CHAIR
PRIOLA: Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yes, I didn't hear you
well and I apologize. When you said in
foods increasingly, there's less bovine gelatin and an increased used of
porcine gelatin. Was that porcine skin?
DR.
MASSON: Yes, one can make and, indeed,
one does use porcine bones, but the large majority of porcine gelatin made
around the world is from porcine skins.
And again just to elaborate on that point, the food industry, the
present manufacturing process of bone gelatin, which we'll hear much more about
in a few minutes, is a very long process.
It's a very costly process.
Whereas porcine gelatin and hide gelatin, certain portions, is a much
sorter process. And economically, therefore,
it's much more viable to utilize porcine gelatin, in particular, in the food
industry compared to bone, you know.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: The numbers went by
pretty rapidly, but it looked to me like the proportionate shortfall from U.S.
production is about the same as the proportionate shortfall when you add
production from U.S. bones processed elsewhere. Is that correct? I'm
looking at the second and third from the last slides.
DR.
MASSON: And again, could you just repeat
that?
BOARD
MEMBER BAILAR: Well, in the answer here
to question 1, the third from last slide.
DR.
MASSON: Yes.
BOARD
MEMBER BAILAR: There was a shortfall of
4,500 tons and a need of 10,000. And in
the next one, it was a shortfall of 32,000 tons and a total need of, was it,
60,000, maybe I have misread this. Yes,
60,000. It's about the same
proportions, but I understand why these includes other production and the other
does not.
DR.
MASSON: The shortfall with the U.S., as
you see, makes 17,000 tons, that is equivalent to over 100,000 tons of bones,
and as we said, basically the cattle industry needs 10,000 tons of gelatin and
only half of that effectively is made here in the States. The other half, because of lack of
availability of bones and lack of capacity in the States for bovine bone
production, has to come from outside of the States, and that's, as you mention,
roughly the same proportion. It's
almost 50/50. Does that help?
BOARD
MEMBER BAILAR: If I understand
correctly then, adding the U.S. bone processed elsewhere doesn't help much at
present?
DR.
MASSON: Excuse me, adding?
BOARD
MEMBER BAILAR: Adding gelatin from U.S.
bones processed elsewhere does not, at present, help very much.
DR.
MASSON: No, because again the total
demand for U.S. bones, because of the other applications, particularly for
photographic and other European and other countries utilization of U.S. bones,
they don't always end up as pharmaceutical gelatin. The end up more often as photographic gelatin, so there's just
not the amount of U.S. bones going overseas which can come back to the U.S. as
pharmaceutical gelatin for capsule production.
BOARD
MEMBER BAILAR: At what point is the
distinction made regarding the ultimate use of the gelatin?
DR.
MASSON: Regarding what, sir?
BOARD
MEMBER BAILAR: Regarding the ultimate
use of the gelatin. Is it all
processed? I thought it was all
processed in the same way.
DR.
MASSON: Well, my colleagues will
describe that in a great deal of detail, and it is more or less, yes.
CHAIR
PRIOLA: And Dr. Wolfe?
BOARD
MEMBER WOLFE: This is sort of a
follow-up on John's question. You
mentioned two factors that are rate limited, so to speak, in terms of the use
of U.S. bones. One was the capacity,
presumably, to convert U.S. bones into gelatin, and secondly, was the
unavailability or the shortage of U.S. bones.
I can't believe that the second one is really a problem. It is likely that only a small fraction of
U.S. bones are currently being exported to other countries for
reprocessing. I mean, is that correct
or not? I mean, it must be a limitation
on production, not a limitation on U.S. bones, and that gets to the issue of
why there couldn't be an increase. If
the capacity is the problem, why there couldn't be an increased export of U.S.
bones to European countries to use them, preferentially, in favor of bones from
BSE countries.
DR.
MASSON: Yeah, your point is well-
taken. The problem, however, is that
the largest consumer of bovine bone, as you see, is the photographic
industry. Out of the 130,000 tons,
which is produced in the states, over 100,000 or approximately 100,000 goes to
the photographic industry. And by
definition, therefore, the remainder simply isn't satisfactory, and we can't
drive -- the gelatin industry is at the bottom of the totem pole, so to speak,
in terms of creating greater availability of bones. The different industries sell so much bone that's made available
to us basically, and there is only so much.
BOARD
MEMBER WOLFE: Are you saying that the
bone either goes to photographic industry or elsewhere and that there isn't, at
the present time, bone from U.S. beef that is not being converted into
gelatin? I mean, what percentage of
the, theoretically, available bone from U.S. beef is, in fact, being converted
to some kind of gelatin? Because my
question is sort of getting to the issue of whether or not it is possible to
divert or not to divert, but just to increase the use of bone from U.S. beef,
even though you want to -- you said there's a tug between photographic gelatin
and other gelatins if the total amount of bone was available, you could satisfy
both of them. So just, specifically,
how much of U.S. bone is, in fact, getting converted into some kind of
gelatin? Half of it, two thirds of it,
all of it? What?
DR.
MASSON: Well, the bone that is
available is being converted. Again,
there are only a few bone producers of the major beef players, but only a few
of them actually make gelatin bone at some of their facilities. And again, it's supply and demand. We can't.
Basically, there's not enough demand from our side that would force them
or encourage them, let's say, to produce still more bone. It's that simple.
BOARD
MEMBER WOLFE: Okay.
CHAIR
PRIOLA: Dr. Johnson?
BOARD
MEMBER JOHNSON: Yeah, I may understand
the way this is processed. But it seems
to me that the ready solution would be that U.S. bone would be used for all
consumables, whether they be dietary supplements and then you could use the
foreign bone for photographic materials.
It's about even.
DR.
MASSON: On paper that's true, but
that's --
BOARD
MEMBER JOHNSON: That's what I'm looking
at.
DR.
MASSON: Yes. But it's rather simplistic, because again we can't make that
determination. It's those industries
who make that determination. The
photographic industry has determined that they will use bovine bone, and that's
their prerogative that we can't influence it.
BOARD
MEMBER JOHNSON: So a solution would be
if we deregulated photographic bovine bone, and that would be a
possibility. Does FDA regulate
photographic gelatin?
DR.
MASSON: No.
BOARD
MEMBER BAILAR: No, they can use
whatever they want.
DR.
MASSON: Yes.
BOARD
MEMBER JOHNSON: So you could split it
up.
DR.
MASSON: I'm sorry?
BOARD
MEMBER JOHNSON: If there's no
regulation on photographic gelatin, you're subtracting it out to produce all
this shortfall, why not make the photographic gelatin from British bones?
DR.
MASSON: I'm sure some is, but again, we
as an industry can't make that determination.
It's the photographic people who make that determination.
CHAIR
PRIOLA: Dr. Bracey?
BOARD
MEMBER BRACEY: Yes, in the information
that you present, the majority of the gelatin is used for photographic
purposes. It seems to me that there has
been a major move away from film based photography towards digital. Have you seen a reduction in the demand and,
in essence, your picture is a static picture, but what does it look like really
as far as the demand for photographic gelatin in the future?
DR.
MASSON: That's a very good
question. As you rightly observed,
digital photography is here in a big way and will continue to grow. But there is some complimentality between
silver halide, the traditional silver halide process, which does utilize
photographic gelatin and the digital business.
So that the two things, digital is growing certainly at a much more
rapid rate, but photographic traditional silver halide photography is still
very much en vogue and, indeed, you know, the last photographic companies,
Kodak, Fuji and so on still continue to invest quite significantly in the
traditional side of the business as well.
So the two things, I'm not sure of --
BOARD
MEMBER BRACEY: Well, I guess, what I'm
wondering is over the years the data in terms of total demand has been static
or has it been actually declining?
DR.
MASSON: I would say it is fairly
static. There has been a diminution for
sure in some sectors of the traditional silver halide, photographic side of the
graphic arts, for instance, probably uses any photographic gels any more. That has gone totally, more or less totally,
to the digital side. But the
traditional film that you or I shoot, the amateur film, medical x-ray and other
types of cinema, film photography for movies, those are still the traditional
situation, and that demand is still very much there.
CHAIR
PRIOLA: Yes, is there a question from
this side or answer?
MR.
SCHRIEBER: Thank you, Madam
Chairman. I would like to make --
Reinhard Schrieber.
CHAIR
PRIOLA: Could you identify yourself?
MR.
SCHRIEBER: From GME, and I would like
to make a remark about potential replacement for the photographic industry of
domestic bones and imported bones. The
following situation is the biggest manufacturer of photographic gelatin is
Eastman-Kodak sitting here in the United States. They are forced to use domestic bones, because as a ban on import
of bones from out of the U.S. into U.S., because the risk of bringing in bones
from maybe BSE risk countries is tremendously high to bring in just in case by
the bones BSE into the United States.
So
gelatin is safe to be imported, but importing bones from other countries, I
think, is of high-risk for this community here, so therefore it would really
replace and most probably negligible risk with gelatin by a big risk by
importing bones, degreased bones from other countries, who therefore is a
replacement in this way, I think, is not a good idea for the U.S. On the other hand, I think it is really
impossible to force Eastman-Kodak just out of using bones from the U.S. I don't know how their reply would be in
this case.
And
maybe one more question, answer to your question about from which European
countries is sourced in Europe has no bone at all coming for the last 20 years
from UK, so the European gelatin industry did not source bone from UK. We do not source bone from Ireland. We do not source bone from Switzerland,
Portugal, the so-called higher risk countries in Europe. All the bones, proven bones used by the
European industry are coming from either Germany, France, Belgium, Netherlands
or Austria. These are the source
countries. Thank you.
SECRETARY
FREAS: Because our meetings are being
transcribed, we're asking everybody who uses a microphone other than at the
table to identify themselves. That was
Mr. Schrieber, the chief manufacturing officer of the Gelatin Group.
CHAIR
PRIOLA: Okay. I think we'll move on to our next speaker. Thank you, Mr. Masson.
DR.
MASSON: Thank you.
CHAIR
PRIOLA: Our next speaker is Dr. Dunn,
who is going to explain some of the manufacturing processes for gelatin in the
U.S. and that might address some of the questions that have arisen.
DR.
DUNN: I also would like to thank the
FDA and the Committee for the opportunity to come in and speak with you today
about the practices of the U.S. gelatin manufacturers. My name is again Michael Dunn. I'm currently vice president of Gelita North
America, and I also serve as the chairman of the Regulatory Committee for GMIA. As you can tell on this slide, there are two
current manufacturers of bone gelatin here in the United States, Eastman
Gelatin, who provides to Kodak, they are primarily producing photographic
gelatin, and GELITA USA, who is primarily a pharmaceutical producer.
When
we put those together, though, the majority of this gelatin goes to the
photographic applications, although there is a substantial quantity that does
go to the pharmaceutical sector as well.
The limed share of the gelatin that we produce is limed bone
gelatin. We do, however, produce a
small amount of what we call Type A or acid bone gelatin, but this is a very
small quantity.
I
also wanted to note that the practices that I'm going to be talking about
today, as well as the processes, apply to both GELITA USA as well as Eastman
Gelatin. Could I have the next slide? So just to set the overall objectives, they
basically are two-fold today. I want to
adequately describe for you today what our current sourcing practices are, as
well as the processing conditions that we use to manufacture bone gelatin in
the United States.
I
also want to clearly confirm that the bone gelatin processing conditions that
we employ here are virtually the same that are currently used in Europe. And more importantly, they meet or exceed
the minimum processing requirements that were spelled out in the GME TSE
Inactivation Study Protocol. This I
want to make clear, because we want to make sure that any of the results, we
want to make sure that they are applicable to what we are producing here in the
United States, as well as what is being produced in Europe.
Could
I have the next slide? So when we get
to sourcing, in the U.S. degreased gelatin bone is sourced exclusively from
USDA inspected beef processing facilities in the United States, and this raw
material is derived solely from healthy cattle that have been deemed fit for
human consumption based upon both anti and postmortem inspections.
Could
I have the next slide, please? When it
comes to SRMs, the U.S. gelatin bone suppliers have been removing SRMs with the
exception of vertebrae since as early as 1998.
And right now, limited quantities of vertebrae-free gelatin bone have
been available from as early as fall of 2002.
Currently, there are no FDA or USDA requirements for the removal of SRMs
in the United States. We primarily do
the two above bullet points primarily because of EU regulations and we supply a
large number of customers that have business in Europe that must comply with
those kinds of regulations.
Could
I have the next slide, please? So let's
go on to the process. What I have
outlined here is an overview of what happens in a daily gelatin production. The major input, of course, to this is the
degreased gel bone. We're on the order
of about 100,000 pounds of gel bone per a production day. And we have an equivalent amount of
hydrochloric acid, so another 100,000 pounds of hydrochloric acid would go into
this next. We use at least a half a
million gallons of water in the production and, of course, there is a lot of
labor and energy that goes into this as well.
What
I'll be talking about primarily today is what goes on in this blue box here, in
terms of the DTL processing conditions.
The output we're looking for, of course, is gelatin. On a base of 100,000, you get out about
25,000 pounds of gelatin, and then about 50,000 pounds of dicalcium phosphate,
which is the primary byproduct of this process.
Could
I have the next slide? So overall, what
we're trying to achieve here, we're starting with the protein we call collagen,
which is an extremely fibrous insoluble protein and we're going to transform
that into a protein that is fragmented and soluble, but has a variety of very
interesting functionalities, which makes gelatin such an interesting
business. So there are three major
things we are trying to achieve here.
Initially,
we need to hydrolyze the collagen. We
do this by breaking, there is intra and inter molecular cross links between the
adjacent chains. We start to break up
peptide bonds, so that we're able to water extract this material from the
ossein that we're producing. Subsequent
to that, we spend a lot of time purifying and concentrating the gelatin. When we do that initial extraction, it's a
very dilute solution about 5 percent, so we have to take a lot of water back
out of that and then we purify the material from both a chemical, physical and
micrological point of view.
If
I could take the next slide, please? So
the incoming gel bone comes to us. It's
delivered by a truck or rail car and these are just simply typical
characteristics of that material, and we would use the same material to make
either the Type B or the Type A gelatin.
So the fat content ranges from 1 to 2.5 percent. The size of these chips is an 1/8th
inch to 5/8th inch. The
mineral protein ratio is about 2 to 1.
And the moisture content is about 6 to 9 percent. And in contrast, it's worth mentioning in
the EU all of the producers there have their own degreasing facilities, which
is different than the way things are done here in the United States. The big meat producers have their own gel
bone processing facilities, and they supply us with this finished bone chip.
Next
slide, please. The first pretreatment
step is what we call acidulation. But
what is happening here is the demineralization of the bone. This is where all that hydrochloric acid
comes into place. What we're trying to
achieve here is the production of what we call ossein, which is this
demineralized bone material. There's a
number of washings, hydrochloric acid washings during this process. We also remove a lot of non-collagen
impurities that come in with the raw bone.
The
concentrations that we're looking at here, maximum, 4 to 6 percent. The way this works is it's a counter-current
distribution process. We start out with
a dilute hydrochloric acid concentration, that's what the initial bone is
exposed to, and it's gradually raised up over this 4 to 5 days. It's a very exothermic reaction, and this is
why it takes to long to carry this out to dissolve out all of this
material. The typical ambient range as
far as temperature after this process is done, the residual acid, is washed out
for about a 24 hour period before we go on to the next step, which is on the
alkaloid side of things.
Could
I have the next slide? So if we choose
to lime, at this point, this is the breaking point we choose to make either
lime bone or acid bone, at this point.
In the case of liming, this is a lime pit that you're seeing up there in
the picture. Again, we being, this is
where we continue to hydrolyze the collagen molecules and there's a lot of
washing that goes on here with the refreshing of the lime solution, so we're
moving impurities.
There
is also something important that happens here chemically that is different than
porcine gelatin. You hydrolyze away the
asparagine and glutamine. You deanimate
those and form their respective acids which drops the iso-electric point of
that molecule from about 9 down to about 5.
So electrically, the porcine and the bovine gelatins are quite
different. We use a saturated lime
slurry to do this. The pH is
approximately 12.5. The liming time is
25 to 70 days that we're tying up this material in production for a long period
of time before we can make gelatin out of it.
Again, and the temperatures, these lime pits are agitated on a daily
basis. We're there to make sure we're
getting proper exposure to the alkaline material to the bone chips that are in
the pit. And these lime slurries are
completely refreshed on a weekly basis.
Next
slide, please. After that, there is a
washing and acidification step. We want
to neutralize the excess lime, again remove, wash out additional non-collagen
impurities, and we want to adjust the pH of the ossein slurry, so we can
prepare it for extraction. So this wash
out period under alkaline conditions is 24 to 48 hours under vigorous
agitation, temperatures from 45 to 70 degrees.
The neutralizing or souring of acids in this case are either
hydrochloric or sulfuric acid, and our target pH for this part of the process
range between 5 to 7.
Could
I have the next slide? In lieu of
liming or alkaline which is what we do most of the time, we're only talking a
few percent of the time we do this process.
We can do an acid treatment and produce Type A or acid bone
gelatin. So the purpose of this process
here is to condition and ready the ossein material for an extraction at a very
low pH. In the traditional process, we
use a sulfuric acid and we expose the ossein to a pH of about in the range of 1
to 2 for about 6 hours, and then we rinse that back to a pH ranging from 2.8 to
3.2. And this is where we will extract
the gelatin. This is, I mean, usually
pH to extract gelatin. Most gelatin is
extracted at much higher, more neutral pH.
We
also have an alkaline pretreatment option that we're looking at, that some of
our customers are looking at, because of all the discussion around sodium
hydroxide pretreatment. In this
situation, you would do this alkaline pretreatment prior to the ossein
treatment. And in this case, you are
able to maintain the pH at 13 or greater with sodium hydroxide for a period of
three hours.
Next
slide, please. Okay. Now, we've finished with the pretreatment,
whether it be for acid bone gelatin or lime bone gelatin and the rest of this
will be common to both of these types of gelatins. Now, we extract the gelatin.
This is where we've wetted the gelatin, we've hydrolyzed it, now we're
going to actually pull this, extract this out of that ossein particle to
produce the gelatin.
We
use demineralized water. What you're
seeing up there is a typical gelatin extractor. We do a series of extractions.
I said 4 to 6 depending on the plant and the company, the way they do
that. But the initial extracts are done
at a lower temperature, and what you will get out is a material that typically
has a higher molecular weight, a higher viscosity, a higher bloom strength.
As
you go to subsequent extracts, that material will become more degraded. It will have a longer profile of treatment
with time and temperature. And those
ending extracts will conversely have higher collagens, lower molecular rates,
lower viscosities and so on and so forth.
So the temperature range is from about 120 to 200 as you go through that
series of separate gelatin extracts that you are pulling out. The conditioning time for each extraction
ranges from 1 to 6 hours and it's 4 to 6 extracts.
Next
slide, please. When that extract comes
off, it's a typical, very dilute solution somewhere in the range of about 4 to
6 percent. So you're saying to get to a
dry product, we got to pull a lot of water out of here as well. So we have initial filtration, this is a
U.S. type filter, vertical leaf type filter.
It's precoated with diatomaceous earth and cellulose. And that basically is to give us initial and
improvement in the clarity. The
solution will also go on to ion exchange.
We want to protect these ion exchange collagens.
Could
I have the next slide, please? So
you're looking here at an ion exchange battery. You see three columns in the forefront and three in the
background. Those are batteries of
cation in that exchange columns. Of
course, the objective here is to deionize this material, depending on whether
it is pharmaceutical or photographic.
It gets more exposure to those columns depending on what is needed.
Primarily,
the cations we're removing are calcium magnesium and iron. On the anion side, it would depend on the
acid that we were souring the material with before we extracted it. And sometimes we use hydrochloric and
sometimes sulfuric. So those would be
the primary anions that would be removed under those conditions. And the finished product from an ash
standpoint would be somewhere between .1 and 1 percent, depending on the
product that we're making.
Could
I have the next slide, please? Now, we
begin to remove water, and we do this by using evaporative means
initially. So we have this 5 percent
solution that we're going to drive up to a 15 to 25 percent concentration. The evaporator you see there in the picture
is a triple effect plate and frame type evaporator. The output temperature is not too high. It usually runs about 125 to 130 degrees on the average. Basically, a temperature that will just make
sure the gelatin doesn't gel up in the production plant.
Could
I have the next slide? Then we have
another filtration. We heat it
again. We've concentrated that
material, so there is more particulate becoming apparent, in certain cases, and
then there is a chance that you may get some coagulated protein, so we have
another clarification step here. The
medium we use are exactly the same in the prior filtration cellulose and
diatomaceous earth, but we use a plate and frame pressure filter. The viscosity of this solution is increasing
now as we move along in the process, and this is what requires a completely
different configuration for filtration.
Could
I have the next slide, please? Then we
take the opportunity to adjust the pH, at this point. The final pH targets of the finished product are usually in the
range of 5 to 7. At this point, it's
usually just a fine adjustment and most typically it's done with sodium
hydroxide.
Could
I have the next slide? Then we do our
final concentration with evaporative means.
Again, this solution is becoming quite viscus, so we're concentrating
our thick, what we call at this point, our thick gelatin liquor. This is an example of a double effect plate
and frame type evaporator as well. And
the concentration here will be a fairly broad range here from 25 to 50 percent,
and this is because, I talked earlier about your initial extracts are much
higher viscosity, so you only will be able to drive those up to about a 25
percent. However, the latter extracts,
which have a much lower viscosity, you're able to drive those up to a much
higher concentration level, and that's what is done.
Next
slide, please. Then we go through a
sterilization step at the end of the liquid phase. After this, we're going to be going into a more solid mode with
the gelatin production, so this is our last opportunity to do something with
the liquid phase. So we use direct
steam injection. We use a temperature
that ranges anywhere from 138 to 149 C for 8 to 16 seconds, and this is
primarily to ensure the product, hygiene of the product.
Next
slide, please. Then we're taking
another tack here in terms of drying the gelatin. We're beyond evaporative means, so what we do is to increase the
surface area, then able to dry this material, we cool it down from about 120
down to about 70 degrees where the gelatin actually sets, starts to set, and
this is down with a glycol cooled heat exchanger. Then it is extruded out through these perforated heads to form
these noodles, which will range in size from under 2 feet long and about an 1/8th
inch thick, and they are deposited on the front end of a dryer, which is in the
next slide.
These
dryers are typically about 12 feet wide and about 150 feet long. The air quality we use is heated, dehumidified
and filtered air. The object is to
produce a stable product. It has very
low water activity. Typically, it has
10 to 12 discrete zones with different temperatures. There's a gradient that ranges from about 80 to 160 degrees
fahrenheit that goes across that entire dryer.
It takes about like 2 to 3 hours to get through this system, and the
final moisture content of the gelatin product is about 10 to 12 percent.
It's
a very touchy process. It's very easy
to melt the gelatin. If you try to dry
it too fast, you know, with too much water, the melting point is lower and it
is going to melt down or you can get case hardening. It's a very delicate process drying this gelatin effectively.
Next
slide, please. Then we do a milling
after the drying and our size is typically 8 mesh. That's our kind of working mesh size. We can do a variety of mesh sizes in the finished product, but
most of our intermediate products we're producing these intermediate extracts
that we use to do our final finished blending, and it's typically about 8 mesh.
Next
slide, please. So as these individual
extracts, whether it be 4 or 6, come off there, they are separated on the dryer
as discrete extracts. Those individual
extracts from daily production are individually blended to make sure that there
is no lack of homogeneity as that material is processed across that dryer. So we blend those with homogeneity. We sample those materials, as intermediate
product, and those that go in the dryers are weighed and go into storage as
intermediate product for future blending and mixing.
Next
slide, please. So there is our
inventory that we're building up with our daily production, and then based on
the specifications of our customers, we build mixes and we formulate mixes with
these individual extracts that we have been producing. These are much larger blends. Some of these are 10, 20, 40,000 pound
mixes, so now we have a high capacity blender that allows us to put those
together.
Many
times we'll make a much smaller small scale mix to make sure that we can blend
it properly, particularly if it's a new product. We can hit the specification before we go to the large scale
blend. So sometimes there is a series
of analysis that we've done it two or three times before we finish off the
finished product.
Next
slide, please. And then we provide that
product once we are ensured that it meets the specifications of customers. We'll package that up using drums, FIBCs or
small bags and then it is off to the customer.
I
hope that has given you a quick -- I had to go through that rapidly. There is a lot of information to cover
there, but you've got that in your handouts there. So I hope that was useful and I would be glad to entertain any
questions you have. And I also would
like to invite you to come out to see our facility in Sioux City, Iowa if you
would like to see first hand how we make gelatin.
CHAIR
PRIOLA: Yes, Dr. Bracey?
BOARD
MEMBER BRACEY: Yes, I have one
question. You said in the cation
exchange process that you treat the product in a different manner depending
upon the end use, i.e., photographic versus other. So, in essence, that suggests that there is the potential for
control.
DR.
DUNN: That's right. That's right. I mean, there are certain types of food products where you may
not go through the columns at all. I
mean, it depends on the ash content.
Typically, the ash if it was unprocessed, it could be as high as 2
percent, okay. In some cases, there
would be no need. And it would get very
sophisticated with the photographic realm whether you are interested in anions
and cations, you go through a cation and bypass an anion or you may go through
a secondary column.
You
know, we have a battery with three columns of each type. Usually, one is a lead column, lag column
and then there is a regenerate one under regeneration. So there is a variety of ways to go through
that ion exchange system, depending on what the specifications of the customer
are. You might have a food customer who
says well, ash is less than 2 or you might have a photo customer and it has got
to be between .1 and 2.5 or .1 and .25 or something like this. There is all kinds of variations on the
thing in terms of exposure to ion exchange.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: I understand from Dr.
Chiu that it is the processors who are responsible for the safety of
supplies. How is that monitored or
enforced here and abroad?
DR.
DUNN: You're talking about the supply
of our gel bone?
BOARD
MEMBER BAILAR: Right.
DR.
DUNN: Okay. We audit our suppliers.
One of the things that makes it a little bit easier here in the States
is we only have a few. We basically
have -- it depends on the company.
Between the two companies, I think, we have five or at most six
different suppliers. So it's not an
unmanageable deal to go in and audit these customers on a regular basis. We also know that USDA is in these
plants. They help us with this. As a partnership, they are in there auditing
all the time.
For
example, when we worked with the USDA because of these European regulations to
start taking our SRMs, back in 1998, they worked with us to do that, to go in
and validate those procedures and so on.
So we have an ongoing program in that respect and we work with the USDA
sometimes to do various things as well.
BOARD
MEMBER BAILAR: What about foreign
supplies?
DR.
DUNN: All of our suppliers here in the
United States, everything we source is here in the United States right now.
CHAIR
PRIOLA: Can you remind me, you said there
were Type B and Type A --
DR.
DUNN: That's right.
CHAIR
PRIOLA: -- process and the Type A is
acid?
DR.
DUNN: Type A is the acid. Type B means base.
CHAIR
PRIOLA: Right. And why do you choose one of those others?
DR.
DUNN: Like I said, we do very little
Type A. I mean, very little. We're talking probably less than a couple
percent, something like 2 to 3 percent, and that's all directed to the
pharmaceutical capsule industry, and there are reasons for that. Because of the way we process this material,
the ratio of viscosity bloom and the ratio of viscosity to concentration is
very different. We can acquire a very
low viscosity concentration ratio with this process for acid bone.
And
sometimes those customers who make the capsules require that they have a higher
concentration. And the limit usually is
viscosity. So if they can get a gelatin
that has a lower viscosity to concentration ratio, that allows them to bring
more gelatin into that capsule, and sometimes in the soft gel, it depends on
the drug fill and what is going on there, that can be very important. So it's very important for a number of
applications in the soft gel area.
CHAIR
PRIOLA: All right. So even though it's a small percent of the
time you do this process, most of it goes to the pharmaceutical industry?
DR.
DUNN: That's right.
CHAIR
PRIOLA: Then the sodium hydroxide
option, the base treatment, you said that's under review. Is that to see how that might effect --
DR.
DUNN: That's right. That's under review for acid bone. The most important thing it's under review
by our customers, and they are currently evaluating that to see if there is not
any other shortcomings of the fact that the sodium is there as opposed to the
calcium from the lime.
CHAIR
PRIOLA: Does it seem to change the end
product at all?
DR.
DUNN: From our prospective, it doesn't,
but that's why we're relying on the capsule manufacturers to do their full
evaluation and that's what we're looking for.
Okay. So we can do it. It's easy for us to do. It's not a problem for us to do that.
CHAIR
PRIOLA: Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yeah, I have a question
regarding the bovine bone sourcing practices.
You said since 1998, you have been removing the specified risk
materials, except for vertebrae.
DR.
DUNN: Yes.
BOARD
MEMBER KHABBAZ: Why that exception and
is it still practiced?
DR.
DUNN: That's a very difficult thing to
do, and there is really up until recent times there has been no
requirement. There are EU regulations
now developing and that's why there is concern there that that may be a
requirement coming into place as early as the end of this year. We're not sure how this is going to roll
out, so we're looking at this strategically.
Right now, there is not a requirement, but there is a big hurdle there
in terms of industry's ability to do this.
This
will cost us more money. It will reduce
the amount of bone available. Right
now, if you take the vertebrae and take it somewhere else, you reduce the
quantity right there by 25 to 50 percent.
And then there will be certain facilities that will just not be able to
do this with the equipment they have.
They won't be able to make this change without investing new
capital. But anyway, the prices we are
seeing now, you can get this material, small masses of this material now, but
it is going to cost you 50 to 100 percent more than the traditional. So, I mean, nobody wants to go there unless
we have to. It's going to be very
costly for us, our suppliers and our customers.
CHAIR
PRIOLA: Okay. Thank you very much, Dr. Dunn.
DR.
DUNN: Thank you.
CHAIR
PRIOLA: I think we'll move on to the
next speaker. It will be Mr. Schrieber,
who will describe the European manufacturing processes for gelatin.
MR.
SCHRIEBER: First slide, please. I would like to thank you, Dr. Priola, this
Committee and the FDA for the opportunity of presenting on behalf of the
Gelatin Manufacturers Association, GME.
Again, details about raw materials sourcing and the bone gelatin
manufacturing practices in Europe. My
name is Reinhard Schrieber. I'm the
chief manufacturing officer operating GELITA Gelatin Group. I'm 36 years in the gelatin business, and I
have served at European Gelatin Association for many years as president,
chairman of the regulatory committee and the chairman of our BSE Steering
Committee.
After
my American colleague, Mr. Dunn, has already substantially presented the
details of the bone gelatin manufacturing process, I would like to go only
relatively shortly into this issue. The
manufacturing processes in general and although the bone gelatin manufacturing
processes in particular are very similar to each other, not only in the U.S.
and Europe but all over the world. The
main differences which can be noticed between the continents are related to the
safety status of the raw material and the sourcing systems in place.
This
is why I like to focus more on these topics, whereas I would like to try as
well to connect the connections and the conditions of our study to those
existing in reality.
Next
slide, please. GME members have taken
several voluntary steps to ensure the safety of the raw materials. Long before the emergence of BSE, the
European gelatin industry has decided to use, and this applies for all types of
raw materials, only raw material coming from healthy slaughtered animals and
released for human consumption, regardless of whether this was mandatory or not
in different member states. So we don't
use any materials from fallen or sick animals.
So
traditionally, no material from fallen animals have been used by European
manufacturers. The three bones gelatin
manufactures in Europe have never used UK bones, but when BSE in the UK became
evident, they confirmed immediately in writing not to use UK bones. After the condition of BSE to humans was
detected, the GME members committed themselves to stop the use of skull bones,
the target which was reached in 1997.
This was further followed by the complete removal of spinal cord by
European meat packers only on request of the European gelatin industry.
In
parallel, our industry started to replace European bones to a certain extent by
imported bone chips, mainly from the United States, but also from other
countries outside Europe. In 1999, the
European gelatin industry was able to convince its suppliers to remove
vertebrae from bovine bones of all ages, which again was more than European law
required.
Next
slide, please. As I stated before, on
top of our European sourcing of our demand for bovine bones can only be covered
with additional imports from different countries. So we always force our suppliers in GBR II countries to
voluntarily take measures in order to increase the safety of our raw materials. GBR II country means that there are so far
no BSE case detected and the European has assessed that it is unlikely that
there will be a case, but it cannot be excluded.
The
U.S. is and Canada has been until recently GBR II countries. Together with our American colleagues, we
implemented the removal of spinal cord, also in the U.S., and one year before
we succeeded in doing so in Europe, we had forced our suppliers in India,
Pakistan, Nigeria to remove the vertebrae as a precautionary measure.
Next
slide, please. Most of the measures
which we had already implemented became mandatory by regulation in Europe some
years later. On top came the postmortem
rapid testing of all cattle older than 30 months. Furthermore, the removal of vertebrae as requested now by law
only for animals older than 12 months, but again in the bones we use in Europe,
there are no vertebrae in at all. So in
practice, the vertebrae is removed from all cattle in the European Union if the
bones are intended to be supplied to the gelatin industry.
I
assume that you are aware of all those regulations presented to you, I think,
by Dr. David Asher in February of this year.
With gelatin regulations, the EU fixed raw material sourcing conditions
and certain safety relevant procedures to all kinds of food grade gelatin. This has been presented to this Committee
two years ago by my colleague, Dr. Scheigas.
Those requirements are in line with the new study conditions, and our
regular intervals controlled by public veterinarians responsible for the supervision
of our plants, although the FDA has made audits to the gelatin bone
manufacturers in Europe two years ago, they went to all plants.
Next
slide, please. Because of the steps
taken by the industry, there was always only a very little chance that BSE
infectivity could be present in the raw materials used to produce
bovine-origin. To date, due to
additional more recently implemented controls like the postmortem BSE testing
and the careful removal of all SRM, it is almost impossible for highly infected
material to enter our supply chain.
Next
slide, please. As with any process and
systems, there is a certain possibility of error. What could happen, for example, animals with very low infectivity
might not be detected by the rapid BSE test.
But they are considered today as to posting no risk to human
health. The surveillance systems in
place might not be adequate in all countries.
The removal of SRM may not be done perfectly. The infectivity of bone marrow has not been finally
clarified. Based on our experience, we
believe that those risks are low, but they are not negligible. They will be quantified by the Scientific
Steering Committee of the European Union and then used in the coagulation of
the quantitative risk assessment, which is currently under development.
Next
slide, please. Last year, more than 9
million normal slaughtered animals were tested on BSE within the whole European
Union, including the UK. And 287
positive cases were found, which gives a ratio of 1 to 50,000. But our tests which had been done and our
study has assumed that all animals used were clinically infective. Supposing that the removal of SRM is not
effected perfectly and that those impurities may not be detected by the gelatin
industry when inspecting the incoming fresh bones, some might enter the process. Again, our tests and our study have assumed
that the bones from all animals contained the food quantity of infective spinal
cord and dorsal root ganglia. Well,
this gives a huge safety margin between the study conditions and reality.
Next
slide, please. Here again, the major
production steps applied during the commercial and the study manufacturing
processes, most of them have already been described by Dr. Dunn. All plants in Europe are ISO 9000 certified
for the quality management and they apply the HACCP principles. The combination of those is about equivalent
to GMP. FDA audits have been
successfully conducted in all European bone gelatin operations two years ago. And a further round of audits is scheduled
for the end of August and early September this year.
It
has to be noted that SGS and independent institute specialized in quality
certification carried out a validation audit.
And each of the bone gelatin plants of GME in Europe and there are no
known GME bone gelatin plants in Europe, and by these inspections all processed
parameters of our study design have been validated against minimum production
conditions in place in those plants.
Just
to clarify what this means, minimum conditions. In certain plants, for example, a higher concentration of the
hydrochloric acid or a longer liming time might be applied by one or the other
manufacturer compared to the conditions of this study. But we used in our study the minimum
conditions applied at least by all manufacturers.
Next
slide, please. One of the differences
in Europe compared with the rest of the world is the fact that in Europe bone
gelatin manufacturers have their own bone degreasing plants. In other countries, like the U.S.,
degreasing is part of the meat packers work.
In the Far East, for example, it is effected by independent specialized
companies. As mentioned before, only
bones from healthy slaughtered animals released for human consumption following
audit and postmortem inspection are collected from the meat processors, who do
then later the deboning of the carcasses.
In
the U.S., slaughtering and deboning is done normally at the same premises. In Europe, we have very often different
locations. So this means that the
carcasses of the animals are transported to a sausage manufacturer, to a meat
packer at a different place and during this transport, the bones are still with
the carcass. Only the SRM, the spinal
cord, the heads are gone, spinal cord is out, but the bones are still with the
meat.
The
incoming uncrushed bones are then inspected by the gelatin industry on sorting
belts for extraneous materials, including potential SRM contamination. Then the bones are crushed to small chips of
about 5/8ths of an inch, this fingernail size. Then the bones -- this means after crushing
that we have a big surface. And for
example, with the hollow long bones the inside would as well become an outside.
These
small bone particles are then degreased by hot water in a continuous flow
process at approximately 185 degree fahrenheit of an average period of about 20
minutes in equipment with high education.
This mix of water, temperature and movement separates fat and soft
tissue from the solid bone particles.
The little ones are then separated by sieves and cyclones, dried with
hot air, but the surface temperature of the bone particles will stay below 150
degrees fahrenheit to avoid degradation.
Then they are sieved to remove fine particles and stored in silos.
Next
slide, please. Demineralization to
remove the phosphates from the bones is carried out at the same conditions like
in the U.S. in a conduct current system.
The total treatment is about 4 days with hydrochloric acid of 4
percent. The remaining protein matrix
of the bones is called ossein.
Next
slide, please. To cut the cross veins
of the collagen acid or alkaline can be applied. This was addressed just before.
For a small portion of the total bovine bone gelatin production, it is
about 2 to 3 percent for special pharmaceutical soft gel capsules. The ossein is treated again for 24 hours
with sulfuric acid at the low pH and after some washes, the gelatin can be
extracted at a pH between 2 and 3.
So
standard bovine bone gelatin is normally extracted at a pH between 6 and
7. And the ossein is treated before the
saturated or over saturated lime solution for at least 20 days. As you have heard, the pH of this lime
solution, which is replaced several times during the process, is around pH
12.5.
Next
slide, please. To make sure that acid
bone will be as safe as lime bone, our industry looked into an alternative
process which would include an alkaline pretreatment, but without working the
special physical and chemical properties of this pharmaceutical as in bone
gelatin. Based on the knowledge that
after the bones are crushed potential infectivity would sit on the surface of
the bones and not inside the bone matrix, we assumed that a short time
treatment of about 2 hours with .3 molar sodium hydroxide solution should be
enough to inactivate infectivity if this pH is kept at 13 for this time.
Our
study results have shown that this treatment is very effective. But our study has also shown that gelatin
made by the traditional acid bone process did not show any detectable remaining
infectivity, which means there is a demand for this type of gelatin is still
very rare. You've heard that we are
depending, of course, on our suppliers to do it or not to do it.
Next
slide, please. During extraction of the
pretreated raw material, several single extracts are collected, each with
different physical properties due to an ongoing hydrolysis during the
extraction. It has to be stated that
due to the different requirements of the gelatin using industry, quite often
photographic, pharmaceutical and food grade gelatins manufactured from the same
raw material batch in sequence. Also,
Eastman-Kodak is manufacturing some pharmaceutical and some food grade gelatin.
This
means that all gelatin of one production day, including the photographic
gelatin, have to comply with the regulatory requirements for food and
pharma. When talking about food and
pharma, one has to keep in mind, as well, that the same capsules might be
filled today with nutritional products, being food, and tomorrow with Rx drugs.
Next
slide, please. For further
clarification, the diluted gelatin solution is filtered by different types of
equipment and filter media in the ossein and ion-exchange columns and
concentrated in the apparatus.
Next
slide, please. So final concentrated
gelatin solution is sterilized by direct steam injection. The temperature is at 4 bar. The pressure in the liquid phase, which is
very important, is a minimum of 280 degree fahrenheit and the temperature stays
for at least 4 seconds.
Next
slide, please. Finally, the sterilized
gelatin solution is chilled to set and then dried with purified and conditioned
air on belt dryers. Each production
batch, which is a single extract, is then tested on physical, chemical and
bacteriological properties. According
to customer specification, different production batches are then dry
blended. The final blends are again
tested under compliance with regulatory and customer requirements and then
released for shipment. These are the
common processes applied by the European industry.
Next
slide, please. There is one special
process which is done by only one company in Europe to manufacture gelatin with
low gelling strength for limited applications.
The degreasing is done of the bones in the common way, but then the bone
chips are autoclaved for at least 20 minutes under 3 bar pressure and 270
degree fahrenheit. After the
autoclaving, bone chips are rinsed with salt water. A certain quantity of gelatin goes into solution.
After
this gelatin solution is taken out, autoclaving at lower temperature and
shorter time is repeated several times.
Then these different extracts are collected, flocculated, ion-exchanged
and evaporated, drying, testing, blending, retesting and shipping is effected,
like with all other gelatins. Low
gelling strengths, the gelatin is used only for certain applications, and the
Committee members might remember that two years ago at this meeting, we already
explained that the main application is a confectionery licorice, although this
process has been successfully simulated during our study.
Next
slide, please. What are the conclusions
which we have drawn from this review presented here? So commercial mineral manufacturing conditions are reflected by
the GME study conditions. The GME
plants and process parameters have been validated for conformity against the
study design. The inactivation results
of the study, which will be presented next, are therefore fully applicable to
the practical gelatin manufacturing processes.
The study demonstrates the ability of the gelatin manufacturing process
to remove and inactivate infectivity even under conditions in which raw
material contain unrealistically high infectivity levels.
Last
slide, please. So safety of European
bone bovine gelatin is established on two principles. The safety of the raw material as required by GME practices and
EU law and the safety of our manufacturing processes as demonstrated by the GME
study. The Scientific Steering
Committee of the European Union has concluded based on all these principles, in
it's opinion, on the safety of gelatin that the risk is close to zero.
Madam
Chairman, Committee, that concludes my presentation. I would like to thank you and the Committee for your
attention. Thank you.
CHAIR
PRIOLA: Okay. Thank you, Mr. Schrieber.
Are there any questions?
Okay. Thank you very much. Our schedule says there is a break, I
believe, after Mr. Schrieber, so we can adjourn and return at 10:00, so that's
about 15 minutes from now. All
right. Thank you.
(Whereupon,
at 9:42 a.m. a recess until 10:03 a.m.)
SECRETARY
FREAS: We're going to go ahead and
resume the meeting.
CHAIR
PRIOLA: Okay. I would like to go ahead and get started. Dr. Hogan had a question for our last
speaker, Dr. Schrieber, that he would like the Committee to hear the answer
to. So, Dr. Hogan, do you want to?
BOARD
MEMBER HOGAN: Mr. Schrieber, I asked
just after our last talk about how the meat processors were audited, in terms
of providing safety of the raw materials to the gelatin manufacturers. Could you address that, Mr. Schrieber,
please?
MR.
SCHRIEBER: Surely. The standard procedure in Europe is that in
every slaughter house, every meat packing operation there is a public vet
present all time, every day as long as this operation works to supervise that regulation
is followed, removal of SRM is done and so on.
And besides this, the gelatin manufacturer are auditing their suppliers
on a regular basis, normally once a month or every two months, again inspecting
as well the commercial documents about where the animals have been sourced,
because commercial document which is required as well by law. So there's a double-fold. But the main thing is that the public vet is
present all day, all the time. Thank
you.
CHAIR
PRIOLA: Okay. Thank you, Mr. Schrieber.
We'll go on to our first speaker for this later morning session. That's Dr. Robert Somerville, who is going
to discuss the GME validation studies on bone gelatin.
DR.
SOMERVILLE: Okay. Thank you, Madam Chairman. It's a pleasure to be back in the USA where
I've spent quite a few happy years working a couple of decades ago. My task is to describe to you the actual
validation studies that were performed in three labs actually over several
years. There were several people
involved and I want to mention them.
First, Ad Grobben is perhaps the most important one of them all, because
he, as an employee of gelatin, which was a member of GME, actually performed or
was participating in all three studies in Edinburgh, in Holland and here in the
USA in Baltimore.
Phil
Steele, actually I should say that, I know a consultant to GME and is present
in the audience and I hope will assist in any different questions you might ask
me later on. Phil Steele is a
technician in my group and he assisted at not only in the work he did in
Edinburgh, but also in Holland to assist in the experiments there.
David
Taylor was my predecessor in running the inactivation group, and he initiated
the studies that we're about to describe and collaborated in setting up the
whole thing. He again is here in the
audience. I inherited the work from
David and responsibility for the work when David retired in 2000, so it's my
duty to report the results, but all the hard work was done before that.
The
work I'm specifically going to concentrate is on the Neuropathogenesis Unit,
which is part of the Institute for Animal Health in Edinburgh. It was funded by GME with further support
from the European Union. I should also
say that at the end of my presentation, I suggested that Bob Rohwer, who
performed the Baltimore studies, spend a few minutes describing the work that
was done in Baltimore.
The
next slide, please. I thought it would
be helpful to describe the basic mechanisms of TSE inactivation first, and
there are three ways in which inactivation or removal can take place. The first is through some form of
destruction through combustion, incineration, oxidation with hypochlorite,
hydrolysis of extreme pHs or with very high concentrations of highly effected
proteases and radiation can have an affect at very high doses.
Next
section, please. What possibly concerns
us mainly today is denaturation type of processes where materials hydrated, in
particular, will have a degree of inactivation effect and exposure to
chemicals, such as strong detergents or chaotropes, can also have an effect.
Next. And we have to look at treatment
variables. There are several biological
parameters that we must consider. The
strain of the TSE agent is particularly important and I will illustrate that in
a couple of slides time. The PrP
genotype may well be important, and we have to consider that. The tissue and the state in which the tissue
is presented in the experiment are also important. We have to consider physicochemical parameters such as heat temperature,
pH and the kind of chemicals that one uses.
And finally, the dynamics and kinetics of the reaction have to be
considered, the time, concentration of any chemicals involved and the
temperature are particularly critical.
Next. Okay.
This shows a slide of some data that was originally published in 1983 by
Kimberlin, et al, where TSE infectivity from two TSE strains was heated for
various lengths of time shown on the axis.
On the Y axis is the titre that was recovered after these
treatments. Two strains were used, as I
say the 22A strains and the 139A strain, and you can see that there is a lot of
rapid reduction in the amount of infectivity present, first. Then a plateau. So the reaction is biphasic with respect to time, and there is
little effect of time after initial exposure.
The
second point to notice is that was a strain difference, so the 22A strain on
this particular example is much more resistent to an activation than the 139A
strain.
Next. Now, in this slide, we're looking at what
happens when we heat at a constant time, 30 minutes, with a range of
temperatures, and what we can see here is that there is little reduction in
infectivity to start off with until we reach an inflection point, and then the
amount of infectivity drops rather rapidly, and that happens for both the TSE
strains that we're looking at here. But
you can see that the inflection point for these two strains differs, so that
for 22C, it's rather thermolabile, which might be a surprise to some. In fact, this temperature which starts to
inactivate is only about 70 to 75 degrees centigrade.
With
22A it's higher, about 97 degrees. But
we haven't specifically done experiments comparable to this BSE or BSE derived
strains, although I'm hoping to do them in the near future. But from the data that we have available, we
think that BSE derived strains are even more resistent to inactivation than the
22A strain here in red, which is the more thermostable of the ones we have
seen. So we can say the inactivation
process is biphasic with respect to temperature and dependent on
temperature. TSE strain, and I
mentioned the hydration state, and I'll come on to that in one moment.
Next
slide. Okay. This slide shows the effect of hitting again at 126 degrees
centigrade for 30 minutes autoclave, three strains of TSE, and you can see with
the 22C strain that all infectivity shown in red has been destroyed. The blue shows the starting titres. The two different blue bands are indicative
of two different PrP genotypes that the TSE strains were passaged in, and there
is no effective PrP genotype in this experiment available.
With
ME7, we cover both the types, a little infectivity, but with 301V, we cover a
lot more. Now, 301V is important to the
rest of this talk. 301V is the most
thermostable TSE strain that has been derived from the passage of BSE through a
particular strain of mice, the VM strain of mice. And it has certain advantages to these studies. Notably, it is very high thermostability,
and that makes it a greater challenge to the studies that we are performing.
On
the right hand side of the panel, you can see a different experiment where
material was heated in a dry oven to 200 degrees centigrade for either 20
minutes or 60 minutes. And I think the
contrast between what happened in the autoclave and the dry oven is really
quite remarkable. We get much less
reduction in infectivity and we've lost our strain differentiation. So there's no strain difference in the
results. And also material survives the
dry oven much better than it does in the autoclave. So that emphasized the point about hydration status. I think if we dry out infectivity, we make
it much more resistent to inactivation.
Can
I have the next one? Okay. This is an experiment where we have combined
temperature treatments with a range of pHs.
I don't suppose you can read this, but each line represents a different
pH from pH 7 up to pH 12 with three strains of TSE again, 301V, ME7 and
22C. And the point is to say that with
301V, in particular, we got very little reduction in the amount of infectivity
up to 100 degrees. Certainly up to pH
of 11.
We
didn't measure what happened at pHs greater than 11, whether we were getting
any reduction infectivity at pH 12 up to 100 degrees, but at 60 and below there
was very little reduction in the amount of infectivity recovered there. You do start to see, in effect, that pH 12
with the more thermolabile strains ME7 and 22C. So the suggestion is that high pH acts synergistically with
temperature when TSEs are inactivated.
Can
I have the next slide, please? So on
the left you can see a list of the things that I have been showing on all
previous, three or four slides, and results and conclusions. Thermostability is an intrinsic property of
TSE agents developed to kinetic mother which I'm not going into today, and so
forget about the rest. Thank you.
Next
slide. Okay. I want to move on now to reduction of risk of TSE infectivity in
gelatin. The challenges that we face,
there is very high resistent to inactivation, and the resistance increases on
drying up infectivity. There are
several available approaches. We can
remove by filtration, for example. We
can denature with heat and high pH or we can use at very high pH, we can get
hydrolysis of infectivity.
Looking
specifically at the risk reduction steps that are available in gelatin
manufacturing, the sourcing of bones, which Mr. Schrieber has just described,
is important as practices in precleaning the raw materials. I'm not going to discuss this. The standard gelatin extraction methods are
thought, were thought to be effective, and that is what the valid study of it
we've been involved in is designed to test.
And then the sterilization steps are steps which may specifically move
TSE infectivity and, of course, other contaminants.
Next
slide. Now, this slide shows the
results from the very first studies that were performed in the gelatin
manufacturing process. What was done by
Inveresk was to take any 7 grain homogenate and look at two components of the
process, either treating with hydrochloric acid, the liming step or the
combination of the two. The reduction
in titre after exposure to the hydrochloric acid was about 1 log, 1.2 was
measured. So that's 10-fold roughly.
Exposure
to lime for 20, 45 or 60 days resulted in a reduction in titre of about 2 to
2.3 logs. And you can see that even
after 60 days, these values are very similar.
So there was a small reduction of about 100-fold, but the time of
exposure had new extra effects. And the
combined treatment results in the reduction of nearly 3 logs, but you can see
that adding these two values together does not come to 2.8. So there isn't complimentary effect, but the
treatments are not completely out of it.
Can
you give me that slide, please? So
there is a reduction in infectivity titre measured by the acid and alkali in
combined treatment of any 7 homogenate.
The combined treatment is more effective than either single treatment,
but they are not titre, and time of exposure to costs in hydroxide does not
effect infectivity titre. And these
processes were not representative of the actual process involved in the plant.
There
is another study by Manske, et al, which showed that there was removal of
proteins under industrial degreasing conditions. These initial studies led to the desire for more systematic
studies to be performed.
Next
slide, please. As I've already
indicated, there was several experiments performed. In Edinburgh we performed four experiments, two alkaline
treatments using two TSE strains, the 301V strain, which I have described, also
the 263K strain, which is a hamster-facade strain, which we believe is
reasonably thermostable, but may not be quite as thermostable as 301V.
We
also looked at an acid process and we tested the addition of an NaOH treatment
in the acid process. Mr. Schrieber has
described the Dutch heat and pressure method and an experiment was performed in
the Netherlands to look at that process.
And as I say, Bob Rohwer will describe the sterilization filtration
experiments later.
Next
slide. Okay. The rational of the experimental design. The TSE source is a high titre BSE derived
model. It's thermostable. It's readily assayed in experimental
mice. We feel that the total titre is a
likely BSE contamination event during industrial processing the gelatin, as Mr.
Schrieber has already suggested. Short
incubation periods, but we have to be aware that we occasionally see very
extended incubation periods after heat treatments. And so we kept the mice under observation for up to 600 days.
The
limits of detection depend on concentration of the sample and the toxicity of
the sample. We cannot inject material
that is toxic, obviously, to the mice.
So sometimes dilution factors had to be included so that we could inject
the mice and not limit the clearance levels that we can measure. However, we feel that near optimum
demonstrate clearance levels are demonstrated from this model.
The
scaled down to simulate typical gelatin manufacturing conditions was performed
by Ad Grobben from earlier and that was reviewed by an international panel
prior to initiation, and as already indicated, Ad Grobben is here to answer
specific questions on that matter. And
the quality of gelatin was checked as the experiments proceeded and again I can
address those questions.
Next
slide. Okay. So this is what was done.
The raw materials were 1.5 kilograms of fresh crushed bones and 500
grams of intact calf backbone, spiked with approximately 10 grams of TSE
infected grain homogenate. Half this
back was injected into the spinal column and the remainder smeared onto bones
and dried onto the surface, and the backbone was then sawed into pieces. There was a degreasing process where the
bone chips were washing at 85 degrees centigrade to remove soft tissue and fat
after the spike had been added, and then dried in the hot air at 120 degrees
centigrade.
Then
the demineralization step was performed.
The bones were soaked in hydrochloric acids of increasing
concentrations. The ossein, of course,
remains as already described. Then the
liming process, the ossein was exposed to saturated calcium hydroxide of pH
12.5 for the minimum of three weeks, and then neutralized. On the acid treatment left over night, also
a pH 3 and then washed in water. The
NaOH treatment, which is included in the acid experiment, one acid experiment,
the ossein was exposed to .3 molars of sodium hydroxide pH 13 for two hours.
Then
the extraction process ossein was stirred gently with water at temperatures
from 60 to 90 degrees to a final gelatin concentration of 2.8 percent. And then purification steps were performed,
depth filtration, ion-exchange, heat sterilization and drying, and all steps
were designed to accurately represent the conditions of the industrial process.
And
it should be pointed out that in the larger process we used indirect heating,
but in the industrial skill process, of course, direct steam injection is used.
Could
I have the next slide? Okay. The spike, as I've already indicated, we use
the 301V strain in three of the four experiments, and we use it because it is
the most thermostable TSE strain tested so far, and it also is BSE
derived. We actually titrated the spike
on three separate occasions. We
actually had two spikes, Pool 1 and Pool 2, and you can see the values are very
similar in all three titrations that we performed with a value of about 7.7 in
each case.
And
as I've already mentioned, all clinically negative animals were observed for at
least 600 days, and then we examined the brains for any evidence of
pathological lesions of TSE infection afterwards. And all positive clinical cases were confirmed by pathological
examination.
Next
slide. Okay. Some results. So this is
the first experiment where the bone was spiked with 301V. The steps were performed degreasing,
demineralization and DCP, the dicalcium phosphate, which is a byproduct of the
gelatin manufacturing process, was also tested for residual infectivity and we
find little. The extract sample after
the liming initialization extraction had a little bit of infectivity here, and
you can see the individual numbers on the left, and that calculates, according
to the Carver Method, to titre of less than or equal to 101.8 ID50
per mil.
I
say less than or equal to, because if you use the Carver Method, you have to
make it -- and you've got incomplete groups at either end of your dilution
series, you have to make assumptions about what happened in that group. So we don't have a 10+1 group,
but we assume to get the number 1.8 that that value, all the mice would have
gone down the 10+1 dilution.
So the number over here is the total recovery calculated against the 10
gram spike that was used. So we got
from total infective load to 108.7 to total recovery of 10.5.
Then
the sample was taken through the filtration ion-exchange in concentration steps
and the sterilization steps, and a sample was then also measured for
infectivity, and no infectivity at all was recovered. And in this case, what these data say is that we couldn't detect
anything. We don't know what would have
happened if we had been able to inject a more concentrated sample again. So again, we can only say this is the limit
of the clearance that we have achieved.
So the total recovery is less than 103.8 starting with the 108.7.
Okay,
next slide, please. Okay. So this is the second experiment in the
alkaline process where we used the other strain, 263K, and I will go through
this a bit quicker. The total infective
load is 109. We recovered a
little infectivity again and the DCP, the dicalcium phosphate, and we also
recovered a little bit of infectivity in the extract sample totaling out to a
total recovery of less than or equal to 104.3.
Next
slide. The acid process here we again
had a spike which had a total infectivity of 108.8 and following the
steps of degreasing, demineralization, then the acid treatment and extraction
we had a recovery of infectivity of 106.2. In this case, we got a clear end point to the experiment, because
the neat fraction, all the animals came down to 10-2, none did, so a
nice, neat Carver calculation of 106.2.
After
the filtration, ionization, concentration, sterilization steps no infectivity
was recovered and we can say that is less than or equal to 104 logs
of infectivity were recovered.
Next
slide. Okay. Now, this is the variation on the acid treatment where the sodium
hydroxide step was included after the acid treatment had been performed. And when this was done, we find that no
infectivity at all was recovered in the titration and we can calculate total
recovery of less than or equal to the 103.3.
Next
slide. Okay. Looking at the data across the way, that's the first alkaline
process and you can see it went from 8.7 to less than 5 down to 3.8 with no
positives.
Next. Next?
Oh, there we go. Thank you. And this is the alkaline process with 263K,
and now you can compare the numbers directly with each other. So we start off with slightly higher spiked
titre and slightly lower recovery of infectivity at this point on the crude
gelatin extract.
Next. And next again. And with the two acid process experiments, we start off with the
spike of 8.8, 8.7. We recover a little
bit more infectivity than in the alkaline process at the crude gelatin extract,
but again when we look at the purified material, no infectivity is recovered
and we can, as I already said, indicate the clearance values from that part of
the process. And you can see now that
the acid process with the included NaOH treatment in here resulted in no
infectivity being recovered.
Next. So this summarizes the data and now I have
included on the right hand side the clearance factors that have been obtained
from the experiment. So we can say that
the alkaline process, the crude gelatin extracts have a clearance of greater
than 3.7, logs of infectivity and for the 263K experiment it was greater than
or equal to 4.7. The finishing, the
purification and sterilization steps have additional clearance factors that we
have demonstrated of greater than 1.2 and that totals over the two parts of the
process to greater than or equal to 4.9.
In
the acid process, we got a clearance from the -- in the crude gelatin extract
of 2.6. The sterilized gelatin after
finishing has got an additional demonstrated clearance of greater than 2.2 and
adding that together, we've demonstrated a clearance for greater than or equal
to 4.8. And then in the acid process
with the additional NaOH treatment, the overall clearance demonstrated is a
value of greater than or equal to 5.4 logs of infectivity.
Next. Okay.
So that summarizes what we have.
From the acid bone process, we have got substantial infectivity measured
before purification in the third experiment, but complete appearance after --
complete clearance after purification, including sterilization. Complete clearance, no infectivity detected
before purification if an additional sodium hydroxide step is included.
With
the alkaline process, there is greater removal of infectivity than after
equivalent acid hydrolysis procedure and there was complete clearance. No infectivity detected after purification,
including sterilization.
Next. So our conclusions are that the gelatin
manufacturing procedure was successfully scaled down and normal bone gelatin
was produced. Both degreasing and the
standard acid and alkaline treatments alone remove most, but not all, of the
implied infectivity before final purification of gelatin. The liming or alkaline procedure was more
effective and the additional sodium hydroxide step in the acid procedure
inactivates a residual detectable infectivity before purification. After purification, all samples do not show
any detectable infectivity. And again,
this was pointing out the removal of infectivity is cumulative, but not
necessarily additive.
Okay. I want to move on to report the data
obtained by the Dutch experiment where they applied pressure treatment to
produce their gelatin. They started off
with titre of their spike, total titre of 9.2.
They went through the standard procedures of degreasing and preheating,
then did the pressure treatment at 3 bar, 20 minutes, 130 degrees centigrade,
then extracted the gelatin. In the
crude gelatin extract, they showed that no infectivity could be recovered, and
the volume that comes to is less than or equal to 0.2.
So
they record a clearance factor of greater than or equal to 6.8 with this
process. They did nothing -- well, they
didn't follow this, the purification steps.
They didn't test that, but there was no infectivity in the gelatin
solution that would have come through this procedure anyway, so it would have
been a waste of time.
Next. So risk reduction and gelatin. We've had several descriptions of this
already in the earlier session about sourcing using only animals passed fit for
human consumption, omission of head bones and vertebrae from source material in
BSE infected countries, and we have shown the removal or inactivation steps or
removal of TSE infectivity during the gelatin extraction and purification process.
It's
also worth noting that the species barrier would reduce the effect of titre or
BSE being -- if humans were exposed to BSE from the source. It is worth also noting that the acids were
performed by injection intracerebrally, and this is by far the most efficient
route of infection, other routes of infection are less efficient.
Next. I'll skip that. That's it. Okay. Thank you very much.
CHAIR
PRIOLA: Are there any questions for Dr.
Somerville? Dr. Bailar?
BOARD
MEMBER BAILAR: I have a couple of
related questions. First, I find the
time deactivation curves somewhat troubling.
They suggest that some of the infected agent is being protected somehow. What is your take on that?
MR.
SOMERVILLE: Exactly that. That there is -- I didn't want to get too
much into the fundamental thoughts that I'm having at the moment about that,
but I think we're getting a dissociation reaction and a protective reaction
occurring when inactivation, heat inactivation is attempted. And the protected species that is formed or
the stabilized species is much more difficult to inactivate. It may be similar to the dried material that
I was showing in some of the earlier slides, too, and that we know is much more
difficult to inactivate.
BOARD
MEMBER BAILAR: Well, there are at least
a couple of other possible explanations.
MR.
SOMERVILLE: Sure.
BOARD
MEMBER BAILAR: One is that some of the
agents being protected inside little particles. There many be subtle differences in the chemical structure of the
ones that survive versus those that don't.
Which leads me to my second question.
In the intact animal, the infection occurs while the animal is alive. It gets circulated and I would presume gets
distributed throughout all the tissues and whatever titre is appropriate for
that. In the experiment, the infective
agent was added to the bone chips, that is at a considerably later stage of
things, where it might be more on the surface of any particles that remain or
it might stay on particles and so forth.
So I'm asking if you have looked into this, and if there is any reason
for concern about this difference in the sequence of when the infection is
added to the materials that you are processing.
MR.
SOMERVILLE: Well, let me answer this,
your question this way. I don't know if
it actually addresses what you are saying.
But the reason for doing the experiment the way we did it was to try and
maximize the exposure in the experiment.
So the thinking was that the greatest risk of BSE contaminating bones
was that during the slaughter process, and that spinal cord, for example, would
get spread down the vertebrae column included with it and dry onto it. So that was what was attempted to be
mimicked in the experiment.
I'm
not -- I suppose the other side of the question is how much infectivity in
living animals associated with bone and bone related tissue? Our primary concerns in that respect again
is to do with spinal cord in BSE infected cattle with spinal cord and ganglia
and related nerves and, of course, the brain in the skull. These should be removed, and again we're
asking the question what happens if they don't, and we've tried to include that
kind of thought in the design of the spiking of the experiment.
BOARD
MEMBER BAILAR: Have you tried to grow
out the infective agent that survives the steps for 20, 40, 60 minutes to see
if it remains highly resistent?
DR.
SOMERVILLE: No, not formally. It's an experiment I want to do, obviously,
but I haven't formed it. I don't think
-- David Taylor whether he has actually done that experiment, either. My prediction is that it would not be in the
protected form after passage through an animal, but we have to do the
experiment. Thank you.
CHAIR
PRIOLA: Dr. Petteway?
DR.
PETTEWAY: Thanks. I just have a couple of questions about the
process of doing the studies and setting them up. Just to make sure I understand, these were scaled down, coupled
steps, so that the spike was at the initial step and then removal was monitored
throughout the process without respiking it each additional step, correct?
DR.
SOMERVILLE: That's right. Yes, that's correct.
DR.
PETTEWAY: Okay.
DR.
SOMERVILLE: I think the experiments
that Dr. Rohwer will describe are looking at process of the final steps in the
process with spiking at the beginning of those individual steps.
DR.
PETTEWAY: Exactly. So that your final removal shows the
cumulative effect of the process to remove the input spike.
DR.
SOMERVILLE: Yes, yes.
DR.
PETTEWAY: I have one other question and
that's with the magnitude of the clearance numbers.
DR.
SOMERVILLE: Yes.
DR.
PETTEWAY: And the less than or equal to
or greater than or equal to. The
magnitude reflects the limit of detection of the assay.
DR.
SOMERVILLE: Precisely.
DR.
PETTEWAY: As opposed to what may
actually be the magnitude of removal.
The magnitude of removal is likely to be much greater than the numbers
reflect, because of the limit of detection of the assay, right?
DR.
SOMERVILLE: Basically, yes. We can only report what we observe.
DR.
PETTEWAY: Right.
DR.
SOMERVILLE: But we can also make some
predictions about what we know from other parts of the process.
DR.
PETTEWAY: Right.
DR.
SOMERVILLE: And that is why, as you've
said it before, it's important not only to look at the overall process, but to
look at individual steps and evaluate what they may be contributing to the
inactivation process or removal process.
However, as the study illustrated, for example, we also have to be aware
that individual steps will not be additive and that one part of the process may
remove the same thing as a later part, later stage might also remove, so you
have to be very careful when you're doing that.
DR.
PETTEWAY: But we can be confident in
the linking of these studies that based on the input spike that there was no
detectable infectivity based on the limited detection of the assay at the end
of the process?
DR.
SOMERVILLE: Yes, yes.
DR.
PETTEWAY: And then the last question I
have is the additional step with the sodium hydroxide. That was evaluated independently?
DR.
SOMERVILLE: What? It was a separate experiment, if that's what
you mean.
DR.
PETTEWAY: Yes, that was a separate
experiment?
DR.
SOMERVILLE: Yes.
DR.
PETTEWAY: Evaluated independently. Okay.
DR.
SOMERVILLE: Right.
DR.
PETTEWAY: Thanks.
CHAIR
PRIOLA: Dr. Hogan?
BOARD
MEMBER HOGAN: Very nice studies,
Bob. I had a question on when you are
calculating the clearance factor here, you've listed an equation that says
clearance factor is equal to gram spike times 10 the log titre spike divided by
milliliters of gelatin times correct factor times 10 to the log titre reduction
or gelatin. For somebody that can't
balance their checkbook, what do you mean by correction factor in the
denominator and why was that entered?
DR.
SOMERVILLE: Okay. The correction factors are to account for
the inherent losses in the process by taking a sample out for intermediate
titration or other evaluations. So
there is natural loss in the amounts going through the process. Does that deal with that?
BOARD
MEMBER HOGAN: Yes, that's great and it
makes good sense. The second question
is did you look at any place in the process where titre might have accumulated
or concentrated, such as inner vessels or on any of the columns or anything
like that?
DR.
SOMERVILLE: I think the short answer is
no. Unless Dr. Grobben would like to
comment on that. But as far as I'm
aware, there was no specific attempt to evaluate that.
MR.
GROBBEN: I do want to. I would like to comment to that, I
think. No attempt was done to try to
measure the infectivity which remains in the equipment, especially for
filtration and ion-exchange, because of the problem to extract that infectivity
from that equipment, so that was not done.
We just measured what was left in the gelatin.
CHAIR
PRIOLA: Go ahead.
BOARD
MEMBER HOGAN: That's what I
presumed. It's just very difficult to
get that stuff off to measure it regardless.
Now, am I to understand that in the gelatin processing process that
these filters would be reused batch after batch or are new filters introduced
in the manufacturing process either in Europe or the United States?
MR.
SCHRIEBER: May I answer this. There is no reuse. It's a one time use. It
may be the answer as well as with the ion-exchange columns. They are regenerated with either alkaline or
assay to purify for the next round of ion-exchange. So there is a constant chemical treatment after the gelatin has
passed those columns.
CHAIR
PRIOLA: Okay. If there are no further questions, we'll move on. Thank you very much, Dr. Somerville, and Dr.
Rohwer is going to present some data.
DR.
ROHWER: Can we go to the next slide,
the first slide here or do I control it?
Are you controlling it or am I?
Where is it here? Oh. Yes, please, go to the first slide. Thanks.
The gelatin manufacturing process is a diverse one. It has many generic features like the
contractionation for plasma, at least I see it that way having worked in both
areas. So they needed a protocol
representing as much of their collective production as possible. And the steps that we were asked to validate
in our laboratory were on bone gelatin.
Next. And we used the process parameters that were
selected by GME, their scale down and this took a lot of time setting this
up. Ad Grobben deserves a lot of credit
for this, as was mentioned. And from
our end, our major concern was about hazard control, and we spent quite a bit
of time on this as well. We did this
study at a scale that was much larger than we typically use in the
laboratory. We were using meters and
liters instead of 100 mls at a time and some of these steps did not fit easily
into the valid safety cabinets and that type of thing, so we had to figure out
other ways to do them. But in the end,
we were successful and it worked without a hitch when we finally got down to
doing it.
Next. The filtrations, the way this was done is we
tested several different types of filtrations that are used across the industry
and then pulled the filtrates, and that's what was actually titred. I'll show you that in a moment. And getting them all done though it took
quite a lot of time, because of the scale and the precautions we had to take to
do it safely.
Also,
all of the work that is done with gelatin has this complication that it is only
liquid above 50 degrees centigrade, so you have to keep things warm. You have to keep them warm on a large scale,
and so we developed a lot of technologies for doing that, which the tempering
beaker turned out to be one of our best tools, but circulating baths and hot
pads were also useful.
Next. Next, please. About hazard control, we used safety cabinets, bags to cover
everything up during the actual processes.
All joints between chromatography column unions and filtration things
were -- transfers were covered with plastic sleeves in case they leaked, put
things in large pots when we could. We
poured nothing. Everything was done by
pumping from one vessel to another in a safety cabinet.
Next. We are also concerned about
cross-contamination simply because of the scale that we were doing this on and
also because of the sensitivity of the results. And as a consequence, all new dedicated equipment was used for
these steps. Everything was disposed --
most things were disposable. The only
things that weren't were the stainless steel filtration vessels and a couple of
other things which could be autoclaved under sodium hydroxide for reuse.
Next. Next, please. We had a question about -- some discussion about the spike
earlier, and I think this was a very gratifying experiment for me. We've been trying to figure out whether our
spikes are relevant in our plasma studies and that type of thing. But in the case of bone gelatin, the most
likely source of infectivity is CNS tissue.
And as a consequence, in this particular case, at least we can say that
the brain derived spike is probably the most appropriate spike for testing
removal from this type of study.
And
personally, I think this is the relevant tissue and we can use it with
confidence. There are issues about
whether 263K or the less adapted BSE strain is more relevant. My feeling is that there are advantages to
both. Actually, clinically, hamster
263K looks a lot more like BSE than the less adaptive 301V strain. On the other hand, this is a strain that was
devised from BSE and so we use that as well.
The important thing is when you do two different strains, is what you're
looking for is the point of convergence between the two to give you some
confidence that the result you are getting are generalized more.
Next. The continuous process was done at the
Institute for Animal Health, and we are only working on the end stage process
right here. Robert has discussed the
rest of this. The continuity was
maintained by Ad Grobben, who took copious notes, and we also have a lot of
further documentation, which I'll share some of that with you in a moment.
Next. So here is the process we have been looking
at. This is the part that Robert has
been describing right here. Well,
actually, they carried it through this stage as well, but the only part of this
process that we're going to be dealing with is this part right here at the
bottom. The so-called purification
steps, the filtration, ion-exchange and UHT sterilization, and we're going to
look at step wise removals.
Actually,
we're going to gang these two together in one experiment. We're going to look at them independently as
well, and we're going to compare the cumulative versus the serial with the
individual testing of these two steps.
This was done just individually.
Next. Here is the basic layout of these
experiments. We have the infectivity
spike. It goes into the crude gelatin,
which is taken directly from production at the same stage of production. It's passed through the filtration
device. And in the filtration
experiments, on one arm, we took the filtrates and took it straight through the
ion-exchange columns and then titrated it.
On another arm, we took it over here and respiked it, figuring that we
may have -- hopefully, we had removed something in the filtration.
This
spike, at the most, would only double the titre that we started out with here
by respiking. If we got any kind of
removal here, we're just starting over at this point. And then testing the ion-exchange by itself. We wanted to use this so that we had
filtrated material to test the ion-exchange process with.
Next. The filtration steps, in fact, involved five
different protocols with various compositions of cellulose, sources of
cellulose and formulations depending on different manufacturing setups across
the industry. The filtrates from those
were all pooled and then they were titrated by themselves before being passed
to the ion-exchange column over here or respiked on this arm and passed through
the ion-exchange columns here. The
ion-exchange consisted of two columns, the cation exchange followed by an anion
exchange, and what we assayed was the eluate from both, the final eluate from
both.
Next. And then in terms of the UHT sterilization,
the ultra high temperature sterilization, we again started with gelatin from
production, infected that, spiked that with infectivity and then did the UHT
test and titred that. So it's a much
simpler pathway.
Next. Next, please. Here is a picture of Ad Grobben setting up the filtration
experiments. This is the filter
apparatus over here. We're
transferring, I believe at this step, we're preheating the filter with hot
water that has been heated over here, and I'm not sure that's what we're doing
there, but that's what this is doing.
This is the hot water. It had to
be preheated so that it was warm enough to keep the gelatin melted once we put
it in there. We've got another bath
heating up the gelatin to dry through the filter.
Next. Here's the filter being assembled. It's quite a large apparatus, compared to
what we're used to, but we were able to do all of this within the hood, though
the transfers had to be through this pump on the outside. There's the filter A being added.
Next. Here is the filter A being stirred in the
filter and then it's drained to form the filter cake in the bottom.
Next. Here is the filtration apparatus setup being
done. Here is the vent in case, because
you have to vent some air out of it in the early stages, and through HEPA
filter here. And here is the assembly
after the filtration is over.
Next. This was a keeper in that experiment, and
this was a failure. We always inspected
the filter cakes after the filtrations to make sure they were intact. There were no possibilities of leaks and
that kind of thing before we would keep the filtration as a successful
one. And so the only thing that goes
into this study were successful filtrations.
Next. Here is a picture of the column apparatus,
the ion-exchange apparatus. These were
gigantic by our standards. We set them
up on a mobile cart on a chromatography rack that we can roll, so that once we
got everything setup and ready to go, we could roll the cart over a very large
plastic bag and then cover the whole thing in this plastic bag and seal it up
during the actual experiment in case there were leaks. Thank God there were none.
Next. And here is the apparatus that we used for
the UHT Inactivation Experiment, and I can make that a little clearer in the
next slide, which is diagrammatic.
Next. The principles that we are trying to employ
in the UHT study that we were trying to mimic from the actual production
environment is, from my prospective in studies that I'll talk about later this
afternoon, infectivity is not intrinsically resistent. The problem is delivery of the inactivant
and the inactivant finding sanctuaries to hide from the steam, and drying is --
drying into a film is one of the biggest problems.
And
one of the nice features of the UHT process is the gelatin is being pumped
through a pipe in which live steam is being injected. There is no head space.
There are no sanctuaries. There
is no place for this stuff to dry.
There is no place for it to escape from the hydrolytic environment. We wanted to duplicate that as best as we
could.
Next. Next, please. So we did that with this apparatus where we filled this stainless
steel capillary and we used this capillary so that we could affect a very rapid
heating and cooling, because the whole process, the UHT process, is a 4 second
exposure to 140 degrees centigrade. So
how do you do that in 4 seconds? Well,
you have to get the heat to it in a hurry.
We didn't try to do it dynamically.
We did it statically. But we did
it in this way.
So
we have this chromatography capillary here.
We have a thermocouple, which is embedded in the tube. The probe is right about here. We have another thermocouple on the outside
to track what is happening in the bath.
And then to relieve any over pressure in the device, we have a water
column going here to a back pressure gauge, which ultimately if it were to
leak, it would go into this tube right here.
And this relieves the hydrostatic pressure that is developed by the fact
that we're heating this gelatin up in here.
But we've got within the gelatin column itself, we have no head space.
Next. We'll take that and the way we get our rapid
heating is through trial and error. We
set up a protocol where we can dip this thing, hooked to its various
thermocouples, up to this recording device into our 160 degree oil bath and
then as we see the temperature hit our target transfer temperature, which was
about 80 degrees, we quickly dump it into the 140 degree bath and it comes to
equilibrium in the successive period.
We then take it from there after 4 seconds has elapsed and dump it into
the other temperature. We're tracking
this whole thing on the computer. We're
watching it in real time as we're doing it.
Next. So we get curves like this. This is seconds down here. This is degrees over here. This is the outside of the capillary
tube. So we're seeing the capillary
dipped into the 160 degree bath, and here we're seeing the transfer into the
140 degree bath, and we're seeing it come to -- and this is the internal
thermocouple and it is coming to temperature very quickly thereafter, and then
at 4 seconds we plunge it into the water bath and that's the way there.
Now,
what I showed you first was the hamster experiment. We have now advanced to the mouse experiment. And there is one important difference. As we got more and more experienced with
this, we were able to get this ramp time down to shorter and shorter
periods. We had about 4 seconds on the
first one and about 1 second here. We
did four or five trials, actually, three in the end we focused on once we got
the method working. And then we picked
the best of those trials. And what I
have shown you is the temperature records for the two best trials for mouse and
hamster and that's what got titrated.
Next. Next, please. This is the results of all these experiments. The pooled filtrates gave a very
disappointing clearance. I was
expecting much higher than that. The
respiked column gave this, only about a half log removal, and remember for
these types of input titrations, we got about a .3 log error associated with
these numbers. The successive
filtration and ion-exchange gave about 1.8, which very interestingly, but probably
somewhat randomly, is exactly the same as the added values between these two.
But
I think it gives us some confidence that putting these things together, even
though the removal at each step is low, we are getting some significant removal
here of about 1.5 to 2 logs. The UHT
sterilization by comparison gave a much better result. Even that 4 second exposure is giving us
about 4 logs of removal. Attached to
this, we have about 6 logs cumulative.
And I think it is legitimate to attach these, because these are quite
different methods of removal versus inactivation.
Next. And this is just a comment on that, these
things were showing independent removal to the extent that we could detect it
with the lower levels that we saw there.
But in terms of looking at the total, at what was actually going on
there, the serial experiment is actually the best one to use, and that's the
one I think we should focus on. But
both of these steps were much less effective than I had expected. And I don't know whether it is because of
the matrix, the apparatus, the gelatin itself, but in the next slide I'll show
you some data.
Next,
please. In our experience, this is
about half these experiments were done by us.
The other half were done by other folks, but they were compiled for a
former presentation of the FDA and a TSE Advisory Committee meeting in October
7, 1997. And typically, and we've done
more of these since then, especially these depth filtrations, and they are
typically removing 3 logs or better.
So
there's something different about gelatin.
And it's either the way we did the experiment or it could be that the
gelatin is so overwhelming in terms of a competitive binder for the matrix that
we're not getting removal because of that.
But anyway, it doesn't fall into expectation. There is a warning in this though, which says that you have to
check these things. You can't
extrapolate from this cumulative experience and presume that it's going to work
in all cases.
Next. I've already dealt with this. Let's go on. Next, please. I just want
to point out that this UHT result is the worst case result. We did it under static conditions. It is heated from the outside instead of the
inside. 4 seconds is a minimum exposure
that is seen in the industry. And we're
using crude brain homogenate instead of material that has been already refined
by the process. And my guess is that
the stuff that has been through the process may be even more susceptible, but
that's a guess.
Next. I want to make one final point and that is
that the total exposure that these samples got really begins, at least for
sure, with the 263K case, with the 80 degree exposure. I mean, somewhere between 80 and 100
degrees. We have a series of
experiments which are actually on the next slide that I did in the '80s showing
that we start to see affect around 100 degrees, and we get total killing in a
few seconds at 121.
So
this ramp temperature is also contributing to the inactivation here. And if we take these ramp temperatures and
add it from 100, the ramp exposure plus the exposure temperature for the 263K
case and the 301V case and plot them on the same curve, which I'm going to do
next.
Next,
please. This is just showing you that
there is an effect at 100 degrees and above for the 263K case, at least.
Next. These are the ramp times for those former
experiments.
Next. Let's go on. Next. I just want to show
you this last slide. If you plot the
data from the 301V case and the 263K case on the same time axis down here,
including these earlier studies out here which were done at 121 versus 140, and
plot it back to the origin, you get a straight line through these things. Well, I first draw the line through
them. And what that is telling me is
two things.
One,
there isn't really any significant difference between the sensitivity of these
two agents to this process. And two, it
gives me some confidence in saying that if you are to extend this process to 10
or 12 minutes, you get another 4 logs or so removal. This is something that should be checked with actual kinetic
experiment and kinetic measurements, but it seems to me that this being a
minimum is a very -- this 4 seconds being a minimum exposure is a very encouraging
feature of this experiment.
Next. In conclusion, the purification steps are
removing 4 to 6 logs and the UHT step, in particular, provides a potentially
very secure inactivation step. Thank
you.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: The next to the last
slide you showed the susceptibility to heat over time, and what you had you
mentioned the straight line fit, but I didn't see any intermediate points there
that could really detect curvature in the line.
DR.
ROHWER: No, there isn't. What I'm saying is we're working with the
data that I have. And I think I also
said at the same time that it would be very nice to do a complete kinetic study
on this.
BOARD
MEMBER BAILAR: Yes, but I would not
conclude from that that it's a straight line.
DR.
ROHWER: Oh, I see what you're
saying. It may not be. You're right. From here to here, well, from here to here, extrapolation, I
don't know. I mean, it's hard. I guess, what would you say? You could have something like that, I guess.
BOARD
MEMBER BAILAR: I would say you do not
have the evidence on which to detect whether there is any curvature.
DR.
ROHWER: Okay. Well, I'll grant you that.
And all I'm saying is that this is -- let's put it this way, this data
is consistent with a first order process here, with these two samples behaving
very, very similarly.
BOARD
MEMBER BAILAR: Okay. That's all.
CHAIR
PRIOLA: Dr. Petteway?
DR.
PETTEWAY: That's a very impressive set
of experiments, Bob, especially dealing with the scale down, handling it
all. That's an extremely difficult
thing to do. But the 4.2 logs, was that
the magnitude with some residual infectivity found?
DR.
ROHWER: Oh, yes.
DR.
PETTEWAY: Okay.
DR.
ROHWER: Yes, I mean, we started with
7.5 logs.
DR.
PETTEWAY: Yes.
DR.
ROHWER: So there's still 3 or 4 logs
left.
DR.
PETTEWAY: And that was at 4 seconds
which is worst case?
DR.
ROHWER: Yes.
DR.
PETTEWAY: And what you're saying, I
mean, even given other points that would show a change in that curve, the
likelihood is 8, 10, 12 seconds, there would be nothing left is the point?
DR.
ROHWER: I was very interested this
morning when Michael Dunn pointed out in his presentation that in North America
anyway the typical time is 8 to 16 seconds, as opposed to 4 seconds, and
apparently gelatin can tolerate that quite well. If you would like to say something about that? Well, that's up to Sue. Sorry.
CHAIR
PRIOLA: Dr. Dunn, do you want to
comment on that?
DR.
DUNN: Could you say it again?
DR.
ROHWER: Yes, if I could repeat that,
what I just heard here is that there is apparently no problem extending that
time for 8 to 16 seconds.
CHAIR
PRIOLA: And Dr. Hogan?
BOARD
MEMBER HOGAN: Well, the question is why
does the European process use 4 seconds and is there a ramp up time to that or
is it just the batch is brought in, zap 4 seconds and then it is taken out?
DR.
ROHWER: I would like to defer to Mr.
Schrieber, if I could.
CHAIR
PRIOLA: Yes, Mr. Schrieber?
MR.
SCHRIEBER: What I explained in my
presentation already is that we used the softest condition we have found in one
of the gelatin plants in Europe. So
it's not uncommon to have like in the States a longer temperature or even a
somewhat higher, longer time or even somewhat higher temperature, but we had to
choose the minimum conditions founded in the three or four studies, and that's
what it is. You are right if the time
would be expanded to 6 seconds or the temperature would go up to 140 instead of
138, this would not really harm the quality of the gelatin.
CHAIR
PRIOLA: All right. If there are no other questions, thank you very
much, Bob.
(Applause)
CHAIR
PRIOLA: I would just like to say having
gone through the bulk of this, these infectivity studies in our rather thick
handout, that it is very impressive the work that Drs. Taylor, Somerville,
Rohwer, Ad Grobben and Schrieber have done studying inactivation of TSE
infectivity through the gelatin processes.
It's a lot of real nice work.
I
would like now to ask Dr. Morris to come up and explain to us the USDA's
gelatin policy.
DR.
MORRIS: Okay. Good morning and thank you for the opportunity to speak with your
Committee regarding APHIS's policies regarding the importation of gelatin.
COURT
REPORTER: Dr. Morris, hit the volume
button.
DR.
MORRIS: Thank you.
CHAIR
PRIOLA: I'm sorry. My apologies. It was supposed to be Dr. Rogers. I'm very sorry. That's my
error. Can we just go with that?
DR.
ROGERS: Yes.
CHAIR
PRIOLA: Okay. My apologies. I'm
sorry. You should have told me. I'm misaligned in the agenda. Okay.
So, in fact, we're not going to hear from Dr. Morris yet. It's Dr. Rogers who is going to give us a
risk analysis of infectivity.
DR.
ROGERS: Well, I guess the slide has
disappeared for a minute there, so don't start the thing until -- the timer
until it shows up. Is the mike on? Okay.
Thanks for inviting us down here today from Canada. I'm from Health Canada.
CHAIR
PRIOLA: We can't hear.
DR.
ROGERS: So is the mike on? Oh, it's on now? Oh, closer. Taller and
closer. How's that? Okay.
At Health Canada we have been doing a number of quantitative risk
assessments and part of our topic today that we will be covering is what's on
your agenda. But I did want to say that
what we're really looking at is the varying-CJD risk to consumers eating foods
containing small amounts of processed ruminant products. And I want to talk about some of our
modeling functions.
Next
slide, please. We have just completed a
quantitative risk assessment for basically products that contain beef extracts
and the beef extract industry certainly has a lot of similarities to the
gelatin industry, so some of the information I'm going to provide today will
certainly with some understanding of the overall picture. I do want to present today like the
quantitative model parameters for the evaluation on pairing CJD risks. I want to focus on the front end parameters
for risk analysis, and I want to provide some information on evaluating
uncertainty in the parameters and provide information on variability that we're
using in our models.
Next
slide, please. The purpose of our risk
assessments are really to provide information on two risk outcomes of the
probability of individuals acquiring varying-CJD through the consumption of a
product and the annual number of varying-CJD infections that could be predicted.
Next
slide, please. The approach that we're
using, basically, the first thing that we look at is the length from BSE agent
to varying-CJD. To date, there is no
direct evidence linking the acquiring of varying-CJD to particular
products. And I want to emphasize that
certainly the only information we had previously was the work of Simon Cozens
of the UK for food products which had some implicated meat pies, sausages,
these types of things in his work, but he has, in fact, recalculated his and
reevaluated some of that publications and in Edinburgh last year he has, in
fact, shown there has been no statistical significance to particular food
products and varying-CJD. So that's an
important picture.
The
presence of the BSE agent in the product of concern are not measurable by our
current techniques. The only thing that
we can actually still seem to have some type of laboratory analysis for is the
presence of CNS materials through IHCGFP and some neuro analyst techniques. The hazard identification basically has
established that there is a route from BSE to consumption exists. And so that's the reason for the
presentation of the modeling.
Next,
please. In Canada, we are using the
model that has basically been setup by the Kodak element. We have an issued statement. We do hazard identification, hazard
characterization, exposure assessment and risk characterization, but nothing
goes forward until you have hazard identification.
Next
slide, please. Our structure in our
risk characterization is depicted here.
Basically, we are looking for these probability statements in the
middle, which are outcomes, but we are looking at the infectivity consumed,
which really comes through our exposure assessments and the consumption
frequencies from the exposure assessment, and then the dose response models
that we have been developing, which are in the hazard characterization.
Next
slide, please. Our structure in hazard
characterization, basically, the main things, variables that we would be
looking at are the susceptibility in human population. We can say that certainly the we in our risk
assessment are looking at worst case assumptions. In fact, with the human population, we are not looking at
divergence, for instance, because of met type of codons, we say that all humans
are susceptible. We're not looking at
immuno-compromised or younger children.
There's no infant instances of that, so like our population
characteristics say that all populations are susceptible.
Infectivity
accumulation is one of the things we are looking at particularly with dose
response type of modeling. Our species
barrier from bovines to humans, I guess, what I would say there again is that
we are looking at the worst case. We're
saying that there is no species barriers.
It's a 1 to 1 ratio, but certainly when we're looking at the advice that
we get from the Scientific Steering Committee over in Europe that they say that
we should use the range of 1 to 10,000.
And,
in fact, the latest publications do say for oral transmission to food products,
you probably should be looking at 10,000.
And we are looking at risk assessments to be tried and say, for
instance, they do want things to be practical and realistic, but then we are
going with due caution. And so some of
these products that we're looking at to start with, we are emphasizing
generally worst case assumptions and seeing what those numbers generated look
like, and so that's what we have been doing.
Next
slide, please. In particular, it's the
dose response area for varying-CJD. And
in our models we are using a threshold dose response as well as an accumulation
dose response. I have a lot of slides
here, so I'll have to hurry along with this.
We'll talk about that a little further along.
Next
slide, please. The structure and
our exposure assessment just basically
analyze the model that we use.
Next
slide, please. And particularly, which
is of interest to this particular audience is the fact that we are looking for
the presence of BSE in cattle populations.
Our models are setup in such a way that we do know that the disease
status of a country changes and I think we have that from our own experience,
but we have them working along the lines, for instance, that food products and
gelatin products are produced over periods of time, so the BSE status and the
amount of BSE possibly infected cattle in the country change, and so we really
want to be able to adapt that to the different lots and processes.
The
tissue infectivity information that we use, basically, a lot of it from the
oral pathogenesis studies to start with from Dr. Wells group and that continues
on in the UK. Our source of infectivity
in the slaughtering areas, our sources of infectivity certainly depend on the
tissues that are used in the products, and I think you've been discussing a lot
of those today with specified risk materials, for instance.
And
this area here I put the word gelatin for the commercial product of what we're
looking at. In fact, this model was
developed particularly for beef extracts, but it is the front end that I wanted
to talk about today. Because what we've
been doing as well as we do look at consumption, the actual final products, the
amount of material that are in final products, and then the amount that go to
consumers and consumer individual servings.
Next
slide, please. So our quantitative
model prevalence of BSE into -- the BSE infected bovines in populations in the
screening procedures. The inference
from the countries of BSE surveillance, first of all, I want to say one thing
and that is that the products we were looking at were generally ones that were
coming from the European Union or could have been coming from other
places. But when we were looking at
prevalence data itself, because of the wide enhanced surveillance targets that
have been going on in the EU that we particular have some good observational
data there to work backwards from.
And
I must say that certainly the EU has also been doing a lot of missions out to
their member states to go for audit and compliance, and they have been doing a
number of good reports on that. That's
why we get some excellent data to sort of give some parameters around to put in
models. However, I'll tell you that
particularly our concerns are detected diagnosed cases are removed from -- are
diverted from food chains.
But
the incubating cattle are a question in the amount of infectivity in incubating
cattle, certainly one of the major things that we have been struggling
with. But in our particular
assessments, we're using 4 incubating cattle per adult cattle diagnosed and
we've done that, basically, from talking to experts in Europe. And the other thing that we're doing,
though, is that we're talking about the infectivity of tissues.
We
started off with giving them exactly the same infectivity as the clinical
animals to run through the worst case numbers, and then we scaled that
backwards. But I'll explain a little
bit later. But what we've done is
basically we try to group countries into low, medium and high prevalence rates
and so then I do have numbers on that, but again this presentation is going to
be a little small for that, but we'll get to that.
As
far as the abattoir screening itself it's concerned that certainly now there
are rapid tests involved, and we've seen that they were talking about 100
percent sensitivity, 100 percent specificity.
There's a number of rapid tests out there. We have done an evaluation of them and we've used the worst case
sensitivity for one particular rapid test, because we cannot tell because there
are varied tests that have been used in similar countries and so you don't
necessarily have all that information.
But
for modeling purposes, again, we're using the worst case. And for the ante-mortem, postmortem
inspections only for diverting BSE infected cattle from the food chain, we're
using a 2.5 percent removal from ante-mortem.
And I think most people know how very difficult it is to diagnose TSE
diseases and that they are very complicated.
However,
they have gone back and it is a requirement in the European Union to, in fact,
state where you are during your diagnoses, are they ante-mortem or are they
rapid test. And so that the range of
variability in ante-mortem depends on the country and in the awareness, the
education and in a lot of infrastructure elements. However, I can tell you that for what we've done, I've only seen
six reports so far from mission compliance audit states and the lowest amount
of divergence is 3 percent out of Belgium from ante-mortem and up to in the
high 30 percents in Germany. So there's
a lot of variability in the amount of BSE infected animals that are removed
postmortem. And this goes into the
models as well.
Next,
please. Next, please. Oh, sorry, I couldn't see it right. Okay.
So now, I'm looking at tissue infectivity. I just want to give a brief run down here that we are using .1
gram of raw unprocessed brain tissue from a clinically infected bovine as the
minimal or as the threshold dose in our models, at this time. I think that most of you are aware that that
is the amount of unprocessed tissue now that has been orally given to a cow
that has come down with BSE in the UK in the latest pathogenesis studies. That animal was 52 months. Again, like a very low dose, but it is our
starting point.
However,
we do put uncertainty around these things, again, up to, per program, 101
to 103 infectious doses. And
then the infectivity that we are assumingly using the same infectivity scaling
standardization to the trigeminal root ganglia, the dorsal root ganglia, the
spinal cord and emboli could possibly go into this slaughtering and stunning
procedures. And I have mentioned
already we are looking at the incubating bovine, and particularly the
sensitivity issue around that, and so we have a scale at different levels in
our final results.
Next,
please. Our sources of infectivity,
particularly, when we are looking at raw materials, it could be going into
things like gelatin. Our CNS emboli in
the blood, possibly spinal column cross-contaminations, blood itself, edible
fat contaminations, bone marrow, spinal column, and trigeminal ganglia.
Next,
please. And, in fact, this is the way
that we have started in our beef extract risk assessment that we sort of look
at in terms of tissue restrictions and no tissue restrictions, and
particularly, although I guess this was the top line here that would be very
much parallel to what could, in fact, be going on in the gelatin industry,
because in beef extract, we do have some productions that only use muscles only.
Next,
please. One of the things that we see
are really the number of bovines that are, in fact, going into batches and lot
production and silo storage in beef extract and this is very similar to some of
the things you are seeing in the gelatin production. So that we do have, in fact, calculated the probability of a
batch contamination, lot contaminations by the prevalence rates and by -- well,
it's a little complicated here for me just to go over that quickly, but it's
those calculations that we were looking for to say that there's a probability
that the consumer product is made from a contaminated product at that end.
And
so we are looking at the number of infected bovine tissues that go into the
batch or lots based on those country prevalence ratios as well. We also are looking at the infectivity
reductions, because in beef extract production, as well, you get a lot of
heating, wet heat, filtering, decanting and denaturation and, in fact, we have
tried to mix estimations on the log reductions there.
Next,
please. In terms of defining our
concerns and characteristics for these products, and I suppose this is one of
the difficulties that we do have with food products that contain small amounts
of a ruminant product ingredient, not always on the labels and not always
necessarily going to the ingredients.
In the beef extract production business, for instance, there is no GME
type organization.
We,
in fact, had to go to every country, major country, around the world that does
have beef extract production and do our individual investigations by companies
to find out the capacity of their equipment, the number of animals, for
instance, that -- first of all, one animal contributes so much tissue per lot,
and so there is a range of animals that go into lots or batches. And so the probabilities are all derived
from that type of information.
And
so like that's something I'm -- actually, I skipped over that a little bit, but
it's very important for this type of estimation of, for instance, that even if
you did have a BSE infected cow going to gelatin production, for instance, you
have to know the capacities of the equipment and the type of equipment and the
different processes and certainly that the gelatin industry have indicated that
there basically are very similar processes, a little different in the other
areas.
I
see I'm at stop time already, but I just need to go to the next slide,
please. Basically, these were the
variabilities and the components that we've been looking at, and I'm probably
way over time here. But can I just kind
of continue on just real quick?
Okay? Because certainly like our
particular interests is really in the production methods themselves, the
production practices, the sources of infectivity, all of these that we've quite
clearly documented in our written reports, which we would be glad to share with
this Committee at a later time.
And
the consumer product characteristics themselves, because there is differences
amongst the groups of products and within the groups of products. And so we have gone through actual analysis
of the amounts of materials that go in there, and then the consumption
characteristics themselves, because each product has a different consumption
characteristic and so we've tried to work that through with a point estimate of
the maximum values.
Next,
please. And the uncertainty issues that
we really weren't looking at in our reports and reporting them as sensitivity,
you really have to do a tissue infectivity incubating bovines or species
bearing the dose response.
Next,
please. And if you want to -- next,
please. Because this is basically the
charts that we prepare, and we are providing like product groups within our
report.
Next,
please. The BSE prevalence is basically
put into our charts.
Next. Our abattoir screening techniques.
Next. For divergence of BSE animals.
Next. Other production methods that we have been
going through with all information we collect and we can provide that as a
range.
Next. And then we've done production parameters
which are a range of ranges depending on the type of processing, etcetera, and
the types of tissues that are added.
Next,
please. And I think I'll skip this one
right now. Next, please. Next.
And this basically is just giving us some information on if you're using
rapid test and ante-mortem tests.
Next,
please. Because it was this type of
scatter diagram that we're trying to provide to sort of show or give the
information on the probability of the consumer batches themselves. If you've got large batches, small batches,
high prevalence, low prevalence, and so we tried to do some diagrammatic
information in our reports to give some idea of the dilution of tissues with no
infectivity or with infectivity.
Next,
please. And the difficulties I have
talked to, and next, please. Bottom
line is that these are the individual outcomes that we have been trying to
quantify in our risk assessments, and particularly, though that we -- you will
find that you can have a lot of problems with surveys, nutrition surveys,
etcetera, for the types of details that you would be looking for for trying to
get estimates on consumption values.
I'm
sorry to have gone over.
CHAIR
PRIOLA: Okay. Well, thank you. Are
there any questions for Dr. Rogers?
Yes?
BOARD
MEMBER WOLFE: You mentioned earlier on
that your assumption of the ratio of incubating cows to infectiveness is 4 to
1. What is the basis for that?
DR.
ROGERS: Excuse me, well, that's
basically expert opinion from Europe, because like we had talked to people that
had the experience with BSE for a number of years, and so that that is just
strictly an expert opinion. There's no
rationale for that except there is a range of incubating cattle that we do use,
but I can tell you the reason why we're using 4, at this time, is because we
have implied such harsh assumptions to the fact that there is, it has the same
amount of infectivity as the clinical animal.
BOARD
MEMBER WOLFE: Can you tell me what the
range is that you said that you use it for?
What is the range or ratio?
DR.
ROGERS: 4 to 10.
BOARD
MEMBER WOLFE: 4 to 10. Thank you.
CHAIR
PRIOLA: Okay. Thank you again, Dr. Rogers.
After carefully checking my agenda, now, we're going to hear from Dr.
Morris from the USDA. I apologize again
for putting you on the spot earlier.
DR.
MORRIS: Good morning. Thank you again for the opportunity to share
our Agency's policy on gelatin. Next
slide, please.
I
am Dr. Terry Morris with the National Center for Import/Export. I am representing the United States
Department of Agriculture, Animal Implant Health Inspection Service, Veterinary
Services.
Next
slide, please. We are headquartered out
of Riverdale, Maryland.
Next
slide. And we are under the direction
of Dr. Karen James-Preston.
Next
slide, please. Title 9, Code of Federal
Regulations, Part 94, 95, 121 and 122 gives APHIS the authority to regulate
animal products. Part 94 gives us the
authority to regulate gelatin.
Next
slide, please. We regulate gelatin
based on the presence or absence of BSE and the association with the BSE
affected region, BSE being Bovine Spongiform Encephalopathy. We have lumped gelatin into one of three
categories. One category would be
gelatin that is derived from non-ruminant species. A second category would include ruminant gelatin that is derived
from cattle that have no association with a BSE affected region. And the third category is ruminant gelatin
that has been derived and has an association with a BSE affected region. For the gelatin that has an association with
a BSE affected region, those regulations are found in Part 94, Section 18(c).
Next
slide, please. And pretty much to
summarize, 94.18(c), the gelatin that is derived from ruminants and the
ruminants are from a BSE affected region, that gelatin is prohibited entry,
unless the gelatin is imported for human food purposes, pharmaceutical
products, photography or any other similar uses that would not result in the
gelatin coming in contact with ruminants in the United States.
Next
slide, please. 94.19 addresses gelatin
derived from non-ruminant species. This
would include your pig, horse, poultry and fish gelatin. On May 27, 2003, an interim rule was signed
that modified the current verbiage in 94.19.
Currently,
next slide, please, the gelatin that is imported that is derived from pigs,
horses, birds and fish species must be accompanied by an original official
certificate endorsed by the full-time salaried veterinarian responsible for
animal health of the exporting country, and it must state the animal species of
origin.
Next
slide, please. 94.19 also deals with
gelatin derived from ruminants, provided those ruminants have not been in a BSE
affected region.
Next
slide, please. This part of the
regulation requires that each shipment should be accompanied by an official
original certificate endorsed by the full-time salaried veterinarian
responsible for animal health of the exporting government, and that certificate
must state four things. The first thing
it must state the animal species from which the material is derived. The second statement must include the region
in which the facility where the material was processed is located. The third statement would include a
statement that the material was derived only from ruminants that have never
been in a BSE affected region. And the
fourth statement must address dedicated facility conditions, meaning the
facility cannot receive, store or process any ruminant material from any BSE
affected region.
Next
slide, please. The last category deals
with ruminant gelatin that has been associated with a BSE affected region.
Next
slide, please. Ruminant gelatin that
has been associated with a BSE affected region must be accompanied by a
veterinarian import permit. A permit is
a legal document that authorizes the importation of controlled materials or
organisms or vectors into the United States.
For ruminant gelatin associated with a BSE affected region, the permit
would address the country of origin. It
would address the animal tissue species, meaning hide or bone. It would address the exporting and the
processing country of origin. Again,
we're looking at BSE-free versus BSE affected.
Next
slide, please. The next few slides
depict scenarios that address how APHIS would regulate the importation of
ruminant material under certain circumstances.
In this scenario, the ruminant material whether it be hide or bone is
derived of ruminants from a BSE-free country, but it is processed and exported
in a BSE affected country. In this
case, we would issue a permit for this material.
The
permit would require that the government certify the country of origin of the
raw animal materials and the government would also have to certify specific
conditions that exist within that facility and the BSE affected region. Again, that facility would have to be a
dedicated facility, meaning it cannot store, receive or process any ruminant
material from any BSE affected region, with the exception of milk and hides.
Next
slide, please. In the second scenario,
we address high derived gelatin only, sourced from ruminants. In this case, whether the hide is derived
from ruminants from a BSE-free or a BSE affected region, the fact that it is
processed in a BSE affected region requires the need for the permit. The permit, when issued, would require that
the government certify that the gelatin is hide derived only, and again because
the facility is in a BSE affected region, the facility would have to be
dedicated.
Next
slide, please. The last scenario
addresses bone derived gelatin. For
bone derived gelatin, and in this case, the ruminants are from a BSE affected
region. This material is allowed entry,
provided the individual obtains a permit.
And when we issue the permit, the permit would require that the individual
maintain affidavits that they obtained from individuals who they distribute
this gelatin to. The affidavits would
require that the individual certifies that the material will not be used as
livestock feed ingredient.
The
material cannot be incorporated into veterinary pharmaceutical uses or the
material cannot be incorporated into veterinary biologic products. And this goes back to 94.18(c), which says
that the material can be imported, provided it is imported for human food,
pharmaceutical products and other uses, photography, and other uses that does
not result in the material being introduced to U.S. ruminants.
Next
slide, please. To complete the process
for obtaining an import permit, you have to submit an application, which is VS
form 16-3. It takes about 2 to 3 weeks
between the time that we can process the application and turn around a permit
to you. The application can be
submitted electronically through our website.
It can be submitted by fax or my mailing it into our office. The permit is good for one year, and the
permit will only allow for the specific commodity requested from the specific
exporters, and it would have to go to the importer that requested the permit.
Next
slide, please. This is my contact
information, in the event that you need to contact our office.
Next
slide, please. And again, I wanted to
thank the Committee for the opportunity to share APHIS policies regarding the
importation of gelatin. I'm happy to
answer any questions you may have.
CHAIR
PRIOLA: Dr. Wolfe?
BOARD
MEMBER WOLFE: This is not meant to put
you on the spot, but as you know, the Department of Agriculture is seriously
considering, we have heard from others, on the verge of, lifting the ban on
importation of cattle, beef, from Canada to this country. You've outlined a thoughtful and, I think,
careful permit process that affects gelatin, for instance, which would come
from a BSE affected region, such as Canada.
Do
you not think that there is somewhat of a contradiction between being so tight
properly and restrictive about allowing gelatin to come in from there, but
seriously considering lifting the ban on meat from what would be the first time
the United States would ever have lifted a ban that previously existed from a
BSE affected country?
DR.
MORRIS: Yes, sir, the APHIS TSE working
group has devised a list of low-risk commodities and a list of mitigation
factors under which those low-risk commodities can be imported, the specific
criteria under which we would consider accepting these low-risk
commodities. That list has been
presented through channels to the White House and it is our understanding that
the White House has disseminated that list to the trading partners, so it is in
negotiation to make sure that all of our trading partners are aware of what the
potential actions would be.
BOARD
MEMBER WOLFE: So you're saying that
beef is presumably on a list as a low-risk commodity? Is that what I interpreter you're saying?
DR.
MORRIS: And I would have to look at the
list, but it's specific categories and it's specific ages. And, Lisa, if you want to help me out
here? Thank you.
BOARD
MEMBER FERGUSON: Yes, I'll try and help
you out. Speaking for the Department,
first of all, I would like to reiterate the point that you probably shouldn't
necessarily believe all the rumors that are in the press and everything that
you hear. There are lots of things
under consideration, not only at USDA, but through the entire Administration,
so all of the departments are contributing to these discussions.
And
the discussions are centered around, you know, is there a science-based way to
look at the situation? Are there things
that we can do, that are based on known science to address the situation with
Canada? And as Terry has described, at
least, we, at APHIS, have provided some recommendations for certain products
that perhaps could be considered low-risk and could initially be allowed for
import under certain conditions.
I'm
not at liberty necessarily to say what is on that list, what might not be on
that list, but we have tried to address it.
Okay. First of all, products
that are accepted internationally not to present a risk of transmission,
obviously, are not affected already.
But we are looking at a range of things to say these could be considered
lower risk than other things. And
really it's a wide ranging list that goes over a lot of issues.
BOARD
MEMBER WOLFE: Thank you.
DR.
MORRIS: Thank you.
CHAIR
PRIOLA: Okay. Thank you very much, Dr. Morris.
We'll now move on to the open public hearing portion of the
morning. So, Dr. Freas?
SECRETARY
FREAS: As part of the Advisory
Committee program, we hold open public hearings, so that members of the public
may wish to make comments to the Advisory Committee will have the opportunity
to do so. At this time, I have received
two requests. One is a written
request. This written request was run
off for the Committee members, posted in the viewing notebook out on the table
and some copies were provided for the public if you were here early.
The
second request is from Mr. David Bieging and he is at the microphone right
now. Welcome.
MR.
DWYER: Actually, I'm Dan Dwyer. Dave Bieging made the request and I'm going
to speak. I'm Dan Dwyer. I represent the Gelatin Manufacturers of
Europe, and I'm also speaking today on behalf of the Gelatin Manufacturers
Institute of America. You've already
heard from representative of these two associations this morning. These associations represent virtually all
of the gelatin produced in Europe and in the United States.
These
two associations have been working for many years, as you know, to ensure that
gelatin is safe and we've been pleased to be able to do so in cooperation with
the FDA. As we've discussed with FDA
previously, we would like, at this time, to comment on the questions that FDA
has asked this Committee to address today.
Specifically,
FDA's Question 1 currently reads "Do the results of these new studies
demonstrate a reduction in infectivity that is sufficient to protect human
health?" This question must be
interpreted in light of the normal circumstances surrounding gelatin
production. In particular, the question
focuses only on the manufacturing processes that were studied, but in practice,
as you've heard today, the safety of gelatin is based on two principles.
The
first principle is the use of raw materials.
As you know, in Europe this involves controls on raw materials imposed
by the European Union and by GME members.
The second principle is the use of manufacturing processes that can
eliminate any potential infectivity that might theoretically be present in the
raw materials. In Europe, this involves
the use of the processes that you've already heard discussed today and that
have been studied by GME.
These
two principles of gelatin safety apply as well to all bone gelatin regardless
of geographic origin. Therefore, we
request that when the Committee considers FDA's Question 1 it take these two
principles into consideration, that is we would recommend that the question be
revised to read "Based on the use of raw material sources and gelatin
manufacturing processes, as described in the information presented to the
Committee today, do the results of these new studies demonstrate a reduction in
infectivity that is sufficient to protect human health?"
FDA's
Question 2 addresses the Agency's guidance on gelatin. As you have heard already today from Dr.
Potter, in 1997, FDA issued a guidance document that established certain
parameters for the sourcing and processing of gelatin in order to avoid BSE
risk. At that time, the effect of the
gelatin manufacturing process on in infectivity had not been proven. The data discussed with the Committee today,
however, in our view, provides a basis for concluding that FDA's guidance is no
longer necessary.
Indeed,
as Dr. Chiu mentioned to you earlier, you may decide that gelatin should be
exempt that gelatin should be exempt from any FDA restrictions. At a minimum, we believe that the guidance
should be modified so as to improve the opportunity for European raw materials
to be brought into compliance with the guidance while at the same time
maintaining appropriate controls on the use of European raw materials and, as
Mr. Masson expressed before, ensuring a continued adequate supply of gelatin
for pharmaceutical use.
If
the Committee takes the approach of modifying the guidance in this way, we
request that the Committee consider two potential modifications to the
guidance. The current text of the
guidance has been provided already to you by FDA and, indeed, our recommended
changes to the text have also already been provided to you for your
consideration.
First,
FDA's guidance currently requires that "cattle come from BSE-free
herds." As a practical matter, the
term BSE-free herd refers to a herd in which there has not been a single animal
identified with BSE. In Europe, it is
mandatory, as you've heard, that animals over 30 months of age be tested for
BSE, whereas animals under that age are normally not tested, because they have
not been defined, at this time, to pose a risk to human health.
Thus,
in practice, a BSE-free herd is a herd in which BSE has not been detected in
tested animals. FDA's guidance in this
regard would be clearer if it were to include a brief explanation of the term
BSE-free herd by stating "BSE-free herd as determined by generally
accepted testing procedures."
The
second modification to the guidance that we would ask the Committee to consider
is one that this Committee has considered before. FDA's guidance currently requires that heads, spines and spinal
cords be removed from gelatin raw materials "directly after
slaughter." In 1998, this
Committee recommended that the removal of spines may be done at any time during
the deboning process. Indeed, the
removal of heads and spinal cords is not an issue as you heard, because they
are already removed before or at the time of slaughter.
Therefore,
it continues to be appropriate for FDA's guidance to be modified to permit the
removal of spines at any time during the deboning process. As the Committee considers FDA's Question 2B
then, we request that these proposed modifications to the guidance be taken
into consideration. A copy of our
recommended changes to the guidance has been distributed to you already for
your consideration, and it also has been made available to the public.
Thank
you very much. We appreciate the
opportunity to appear before you today.
SECRETARY
FREAS: Thank you for your
comments. Is there anyone else in the
audience who would like to address the Committee, at this time? If not, Dr. Priola, I would like to state
that we all have three more open public hearings throughout this meeting, and
we do encourage the public participation.
Thank you.
CHAIR
PRIOLA: Okay. So the questions put to us by the FDA are now open for discussion
and voting. Do we have the questions to
put up? So the first question, while
they're getting it up there, is simply, well, hopefully simply, do the results
of these new studies demonstrate a reduction in infectivity that is sufficient
to protect human health? Are there any
comments or any discussion from the Committee?
Yes, Dr. Hogan?
BOARD
MEMBER HOGAN: Since nobody else is
biting, let me take this opportunity to say that when I reviewed your article
and when I started reading this voluminous amount of material, I sort of looked
with the same sort of skeptical eye that I do when I accept papers for
publication, and I initially had, when I started reading, several questions
about processing and scale-down issues and residual infectivity, etcetera. But as I got deeper and deeper in this, we
concur that those had been addressed.
So
the initial questions that I have asked today, I am extremely personally
pleased with the results of these studies.
And while no study can be absolutely perfect, and I think that all the
questions that the original Committee in 1997 had regarding the data, in my
mind, have been answered.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: I agree that these are
very important experiments. They were
very well done. I read the reports also
as somebody who has done a lot of reviewing.
I do have one remaining question or set of questions. I'm not sure that we know enough about the
time course of deactivation and why some of the infective agents seem to be so
resistent.
CHAIR
PRIOLA: Well, maybe Dr. Rohwer would
like to address that more specifically, but, well, would you, Bob, would you
like to, since this is your day. I
don't want to speak for you.
DR.
ROHWER: You're asking a fundamental
question of TSE science, actually. And
it's something that is going to get a lot of discussion this afternoon. And so I don't know, I mean, I have another
talk that I'll be giving and it goes directly to that question, and Robert
Somerville has given his perspective on it, and we're going to hear from David
Taylor as well. And I think, is there
anybody else? I honestly can't
remember. Well, and David Asher has
some new data on this area as well.
And
you, yourself, put your finger, I think, on the central issues in your first
question to the panel, I mean, to the speakers earlier this morning about the
biphasic nature of these inactivations and what is behind them. And we don't know for sure. My own bias is quite different from
Robert's. I mean, I don't think there
is any intrinsic difference between these agents. What we're talking about is sanctuaries and an inability to
actually reach all of the agent. But
there are other interpretations.
You
pointed at one which is a genetic one and there are different ways that you can
look at these kinds of protections. We
don't have the answers to that. And I
think it is a residual question that haunts every single validation study,
inactivation study that is done, is to know just how far you can extrapolate
this data to zero.
I
would like to point out that this is not a new issue. It's something that has bedeviled the vaccine industry, water
purification, virtually any area in which you want to assure that something is
sterile, but you have no way of measuring the entire production lot to find out
whether it is or not. And we're kind of
in the same boat here.
BOARD
MEMBER BAILAR: Yes, but I'm not
concerned about extrapolating down past the last data point. You have data showing that the curve
flattens out, at least, to a considerable extent.
DR.
ROHWER: Yes, and the point that I'll be
making this afternoon is that the place where that flattening out occurs is
very context dependent. And you can
force it down or up depending on what kind of mixture you are inactivating,
what the conditions are and that type of thing. And so the one thing I can say about these studies is that the
knowledge that that occurs was part of the design of the study.
And
your other question about intrinsic versus extrinsic infectivity, the idea that
you have to introduce the spike into this spine preparation at the beginning,
and you can't know for sure whether you have really mimicked the invito
situation in which you would find the infectivity in a BSE infected cow is a
very appropriate one. However, in this
particular case, I feel more comfortable with it than practically any other
study like this that I have done, because it is the spinal cord and the ganglia
that we feel are the threat. They are
extrinsic to the bone.
And
what was done here is the stuff was actually injected into the spinal cord and
smeared on the bone, actually given an opportunity to dry on the bone, which is
something that probably actually happens, and is something that is very, in my
opinion, probably very dangerous to do with TSE infectivity. And so the original for the total process
experiment, which by the way I wasn't part of the experiment, but nevertheless,
my perspective on that is that that was probably just about as good a spike as
you could devise. And I mean, I can't
think of anything better.
You
could always argue with the downstream position of spiking homogenate into
these things, but even there I think we're talking about a worst case spike in
the sense that the homogenate is completely unrefined. And having taken this through the process,
you're liable to have stripped away some of the fats and things like that that
may be protective to these agents in pure brain type associations. But that's speculation on my part. I can't satisfy your basic underlying
concern there, because we don't have data on that point.
BOARD
MEMBER BAILAR: Well, I remain a little
bit concerned, because I recall reading basically in the Daily Press
that in the usual method of slaughter, bits of CNS material do get into the
peripheral tissues. Is that correct?
DR.
ROHWER: With penetrating concussive
slaughter, I think, it is without a question that that happens. And that's -- I don't want to comment on
that. There are people here from the
USDA who can probably tell us just whether that practice still occurs there or
not. I'm not sure.
BOARD
MEMBER FERGUSON: I'll answer that. Actually, the issue is with air injected
stunning, where you've got a captive bolt and then you've got holes drilled at
the end of it, and you inject a bolt, a blast of air, and that type of nomadic
air injected stunning is not used in the U.S. industry any more. Our colleagues at the Food Safety Inspection
Service are actually in the process of promulgating regulations that officially
prohibit that, but based on our understanding of slaughter practices, it is not
used in the U.S. Is it used elsewhere?
BOARD
MEMBER WOLFE: Well, like in the
countries where we're talking about, the European countries?
BOARD
MEMBER FERGUSON: In Europe it also
prohibited by regulation within Europe.
BOARD
MEMBER WOLFE: Within all of Europe?
BOARD
MEMBER FERGUSON: Yes, yes. Actually, well, within the EU. EU regulations prohibit it. So within the community, I think, you can
probably also then assume that any of those countries that are exceeding to the
community, the same thing applies.
CHAIR
PRIOLA: Dr. Somerville, I think, also
wanted to address part of your question, Dr. Bailar. Thanks, Bob.
DR.
SOMERVILLE: Can I just -- is this
on? Okay. Just to add to what Bob was saying and to emphasize what I said
at the beginning of my talk, was that in processes that were considering its
denaturation reaction which is, I suggest, leading to the stabilization of the
aging, past the drying processes that Barbara has just mentioned. There are other processes involved in the
gelatin extraction procedure which may assist in its destruction or removal.
I
suggest that possibly there is a degree of hydrolysis of infectivity which
would not necessarily depend on the stability of the agent in terms of its
denaturation properties, and also, of course, the filtration properties
described are of importance in removing, in the totality of the removal of
infectivity from the process.
CHAIR
PRIOLA: Yes, I think it is also worth
remembering that having sat through many of these Committee meetings and always
asking for data, I now have before me 2 inches of data, all of which point to
the same thing. That in the worst case
scenario you can still inactivate these huge doses of infectivity. And then in the real world we're talking
about starting material that doesn't even have, at least from the European
point of view, as we've heard, since they are now removing the vertebrae, it
doesn't even have that material there to start.
So
whatever contamination may be present is going to be significantly lower than
anything that has been discussed here today.
So at every step of the process, precautions are being taken that should
also be taken into consideration when you're thinking about these things about
total inactivation and sequestering evasion.
Are
we ready to vote, dare I ask? Does
anyone else have anything they would like to say now? Shall we call for a vote then?
SECRETARY
FREAS: There are currently nine voting
members at the table. I will go around
the table starting with Dr. Johnson over there. Dr. Johnson, how would you like to vote?
BOARD
MEMBER JOHNSON: I vote yes.
SECRETARY
FREAS: Dr. Bracey?
BOARD
MEMBER BRACEY: I vote yes.
SECRETARY
FREAS: Dr. Ferguson?
BOARD
MEMBER FERGUSON: Yes.
SECRETARY
FREAS: Dr. Hogan?
BOARD
MEMBER HOGAN: Yes.
SECRETARY
FREAS: Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yes.
SECRETARY
FREAS: Dr. Priola?
CHAIR
PRIOLA: Yes.
SECRETARY
FREAS: Ms. Walker?
MS.
WALKER: Yes.
SECRETARY
FREAS: Dr. Wolfe?
BOARD
MEMBER WOLFE: Abstain.
SECRETARY
FREAS: Dr. Bailar?
BOARD
MEMBER BAILAR: No.
SECRETARY
FREAS: The tally is 7 yes votes, 1
abstain vote, and 1 no vote.
CHAIR
PRIOLA: Okay. So we can move on to Part A of the second question, which is due
to scientific data and information available support the following current FDA
recommendation on bone gelatin. And we
can keep in mind that we can modify as the FDA has said we can modify this
question if we think it is necessary for this recommendation. So that's open for discussion. Dr. Bailar?
BOARD
MEMBER BAILAR: Before we vote on this,
could we have somebody from FDA say whether the modifications suggested are
acceptable?
CHAIR
PRIOLA: I'm sorry, the modifications
suggested by the gelatin manufacturers?
BOARD
MEMBER BAILAR: Yes.
CHAIR
PRIOLA: Yes. Would someone from FDA, yes, Dr. Chiu.
DR.
CHIU: I would put the question back to
the Committee. If the Committee think,
you know, the modification suggested by industry is acceptable, then we will
take that recommendation back to the Agency and then have internal discussion.
CHAIR
PRIOLA: Comments from the
Committee? I would like to read through
the gelatin manufacturers recommendations.
Is there any overwriting reason that anyone can see here to alter what
the FDA already has down, which seems to cover what it should in terms of
removing risk materials? Dr. Hogan?
BOARD
MEMBER HOGAN: No, I don't think it
should ever go under non-exempt. I
think that this is good. The question
is from the industry, why is it important to -- when you say BSE-free herds,
that covers it. I guess what you're not
allowed to use then are herds which contain animals that are younger than 30
months, and you would like to be able to do that. Is that the sense of why you want the modification? Since animals that are less than 30 months
are already assumed to be BSE-free.
CHAIR
PRIOLA: Dr. Schrieber, Mr. Schrieber?
MR.
SCHRIEBER: This request for
modification is based of an opinion expressed by the USDA. USDA has stated to FDA we do not consider
any herd in Europe being BSE-free. So
this means the current text, the way this is written, would exclude altogether
all European bones to be used for gelatin manufacturing and then exported into
the U.S. So therefore we need the
clarifications that under certain circumstances, and that's what we have
described, that the animals are tested according to current procedures in
Europe, that this would be, let's call it, equivalent to the BSE-free
herds. So that's one point.
And
the other request for the modification is what I said before. Due to the transport of the carcasses from a
slaughter house to a meat packer to sausage companies, with bone in, if the
request will stay, removal of spine, I'm not talking about spinal cord, this is
directly removed after slaughter. But
removal of spine has to be taking place directly after slaughter, this would as
well totally exclude the use of European bones, because this is not the
standard procedure.
So
we need some time frame during the further processing, because deboning is done
somewhere else and transport of carcasses without the bones is not
possible. This is the ratio behind our
request.
BOARD
MEMBER HOGAN: Well, then I would ask
Lisa, is that true the USDA considers no herds in Europe BSE-free, despite
testing?
BOARD
MEMBER FERGUSON: Well, I think what
we're dealing here with is the way our regulations are written. And our regs prohibit the entry of ruminant
from any country that is on the BSE restricted list. So, you know, since our regs are clearly prohibiting all these
animals, we can't necessarily make an exemption and say yes, okay, something is
free, something is not free.
CHAIR
PRIOLA: I'm sorry. Dick, go ahead.
BOARD
MEMBER JOHNSON: Yes. If this were modified by this Committee,
that would not affect the FDA regulations, and then you would have two
conflicting rules, right? Is that
right?
CHAIR
PRIOLA: Well, you're not necessarily
going to have two conflicting rules.
You know, the way our regs are written, we prohibit gelatin from
entering, as Dr. Morris has described, unless it can be demonstrated that it is
not going to go for animal use. Okay.
BOARD
MEMBER FERGUSON: So we don't make this
type of an exemption, you know, for stuff going for animal use, if that's
clear.
BOARD
MEMBER JOHNSON: I thought it was all
products derived from cattle that were from BSE positive countries that you
don't permit. But as long as we eat it,
it's all right? As long as humans eat
it.
BOARD
MEMBER FERGUSON: APHIS' authority is
related to animal health issues. APHIS'
authority is not related to public health issues, so our regs are written based
on that authority.
BOARD
MEMBER JOHNSON: But doesn't your animal
health issue say that products derived from, cattle products derived from BSE
positive countries cannot be brought into the country?
BOARD
MEMBER FERGUSON: Correct. Our regs in general prohibit not just bovine
products, but most ruminant products.
BOARD
MEMBER JOHNSON: Yes.
BOARD
MEMBER FERGUSON: From countries on our
BSE restricted list. However, I think,
as Dr. Morris explained in her presentation, there are certain things in the
regs that can be allowed entry and one of those things is gelatin under certain
conditions that is not going for animal use.
BOARD
MEMBER JOHNSON: That's in your
exemptions at FDA?
BOARD
MEMBER FERGUSON: Correct.
CHAIR
PRIOLA: This is USDA, Dick, so,
yes. They are USDA.
BOARD
MEMBER JOHNSON: USDA, that's okay.
CHAIR
PRIOLA: Yes, the FDA is strictly
concerned with oral or topical applications in humans of gelatin, so the USDA
regs aren't our concern. It's the FDA. It's this specific recommendation by the
FDA.
BOARD
MEMBER JOHNSON: Except it isn't --
wouldn't it be a regulation, for instance, that you couldn't bring in cattle
hides from Europe under the safety of animals?
BOARD
MEMBER FERGUSON: No, hides and skins
are exempted from our regs.
BOARD
MEMBER JOHNSON: They are exempted? Okay.
BOARD
MEMBER FERGUSON: They are considered,
yes, not to present a risk of transmission.
BOARD
MEMBER JOHNSON: Okay.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: How is herd defined? Is that all the animals on a single farm or
ranch?
BOARD
MEMBER FERGUSON: I don't know why you
guys are looking at me, because these aren't our regs. Actually, I have to admit, I mean, these are
the types of things that you always run into when you put that type of a thing
in a reg. It's very difficult to define
that. When we look at it from an animal
health point of view, it's a group of animals that is housed together. And if a premise has, you know, two separate
groups of animals that never come into contact with each other and are managed
completely differently, those could, technically, be considered two different
herds. But essentially, it's a group of
animals that are managed together and handled together.
BOARD
MEMBER BAILAR: Okay.
CHAIR
PRIOLA: Mr. Dwyer has been standing
there for a couple of minutes. Would
you like to make a comment?
MR.
DWYER: Yes, thank you. As you've explained, there is a complete
distinction between the FDA's guidance and the USDA's regulations. The USDA's regulations are intended only to
protect animals and not to deal with anything that the FDA has going on
here. If you go back and look at the
early meetings and transcripts of this Advisory Committee when FDA was
discussing with the Committee the formulation of this guidance, you'll see that
the requirement for BSE-free herd restriction was put in as one of a series of
restrictions in FDA's guidance that were intended to protect the safety of
gelatin.
There
wasn't much discussion, at that time, of what a BSE-free herd meant or how that
would be defined. Because it is
obviously possible to define it in many different ways, what the industry was
looking for is a way of defining it in a logical, rational way that is
consistent with current practice in Europe.
And that's basically it.
BOARD
MEMBER BRACEY: Although --
CHAIR
PRIOLA: Go ahead.
BOARD
MEMBER BRACEY: Although I think the
safety of gelatin has been certainly demonstrated to be rather robust today,
what bothers me is, in essence, a disconnect between two levels of
animals. One is the human where we are
considering saying that well, it's okay, based on the data that we see, to
allow humans to ingest these materials.
Whereas, on the other hand, another arm of the Government says that
another animal, which some of us think would be on a lower level perhaps than
the human, that it is not acceptable.
And,
you know, I really feel that we need to have some sort of harmonization,
because the message, I think, that -- if I were the public, I would be somewhat
concerned about the message that we would be issuing.
BOARD
MEMBER FERGUSON: Yeah, that's a valid
point. I would ask everybody, however,
to keep in mind that, you know, it's one thing to talk about an agent that is
coming from cattle and going directly back into cattle versus an agent that is
coming from cattle and is going into a different species. Granted, it has been demonstrated that there
is that transmission, but you do still have somewhat of a species barrier
there.
CHAIR
PRIOLA: Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yeah, and actually,
listening to the USDA presentation, I have that same reaction saying there is
an apparent contradiction here between conditions for animals and humans. But in thinking about it, I mean, you have a
potential amplification. I mean, these
are some different issues that go into place with animals and that's why I
didn't comment. But there is an
apparent contradiction. I agree.
CHAIR
PRIOLA: Yes, Lisa?
BOARD
MEMBER FERGUSON: Can I go back to the
term BSE-free herd? That's very
difficult to define and I don't want to necessarily sound too harsh, but in
some ways it is sort of meaningless. I
know we have struggled with those types of definitions as we tried to setup our
scrapie or CWD eradication programs.
And, you know, specifically, as we're doing our CWD program, we don't
necessarily define under the auspices of that program as herd as free until
they have gone through a 5 year period with extensive surveillance and a lot of
that. So it is a bit difficult to
define.
I
guess I'm not quite sure exactly what level of risk mitigation it's necessarily
adding in this guidance. Probably more
of the risk mitigation is coming from removing those tissues that are at
highest risk and also just through the inactivation of the process itself. So perhaps what we should consider is that
specific point even necessary in there or does it just cause more confusion
than it is really worth?
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: I feel like I just don't
know enough about all this. And I am
concerned about the definition of a herd.
Does this include animals that come from the same source, prior to the
time they are parceled out into different farms? Does it include any element of time? That is, you know, if all those animals there today are gone and
you bring in new ones, is that part of the same herd? What about overlap in time, which I understand is common in the
industry, that there is a continuing flow of young ones in and older ones
out. I just don't know enough about it.
CHAIR
PRIOLA: Dr. Chiu, do you want to
comment on what the FDA means by BSE-free herd or is there something more
specific you can tell us?
DR.
CHIU: Well, I will try. If I didn't get the picture across right,
then I will ask Dr. David Asher to add it.
I think in our original discussion we were thinking a herd is a group of
animals managed, you know, by the same people and also physically they are
together, so they are sort of separated from another group of animals. And also we think when we say BSE-free
means, you know, that group of animals in the past there was never, you know, a
BSE case among that group. In addition,
we were also thinking, you know, that group of animals were never fed with meat
and bone marrow, so therefore they don't have that kind of risk to contact BSE.
CHAIR
PRIOLA: Dr. Wolfe, did you want to say
something?
BOARD
MEMBER WOLFE: I just wanted to ask
Lisa, just from your perspective, what do you think the difference is between
this guidance or recommendation as it now exists and the way that the industry
would like to redefine it? I mean, the
reason I'm asking you is (a) you're from the USDA, but (b) you have just gotten
done saying you don't think the phrase herd has any meaning at all. So if it doesn't have any meaning, then what
is the difference between our current version and what they propose?
BOARD
MEMBER FERGUSON: That's a good point,
and actually I don't really see a whole lot of difference in true meaning
between what the industry has proposed and what currently exists. My sense of what industry has proposed is
trying to make it more realistic and to make it more meaningful in what fits
with industry practices, which is a very valid point, especially this one about
the removal of tissues and where.
BOARD
MEMBER WOLFE: I'm specifically just
talking about the herd definition.
BOARD
MEMBER FERGUSON: Actually, I mean,
after what Dr. Chiu has just said, you know, if those are the specific issues
that FDA is intending to address with that point, then I guess my suggestion
would be to put that in there as the guidance to say that these animals have
not been fed meat and bone meal, those types of things. That is a more accurate definition of the
risk mitigation measure and is more easily understandable and leads to less
confusion.
CHAIR
PRIOLA: Looking though this, I don't
have any trouble. I think that's an
excellent suggestion actually for the FDA to modify it according to what they
mean by BSE-free herd. The other
suggested modification by industry down there at the bottom, I'm somewhat
uncomfortable with, but you had mentioned that you weren't as
uncomfortable. Why exactly is that?
BOARD
MEMBER FERGUSON: Well, I think that's
probably just because of my understanding of slaughterhouse practices. And if this is saying, you know, as it
currently says, let me find it, "and if the slaughterhouse removes the
heads, spines and spinal cords directly after slaughter," that lends
itself to a lot of interpretation.
First of all, talking about spine directly
after slaughter, does that mean right after the animal is stunned and, you
know, hung up on the rail and bled out?
if so, that's not necessarily common practice. You need that vertebral column there to give some structure to
the carcass that's moving through the plant.
You know, and I think the point is that those tissues are removed at
some point in time during the process.
Although they are not going into the start of the gelatin manufacturing
process. It's not as much a point as
specifically when are they removed, it's that they are removed.
CHAIR
PRIOLA: Which the current guidance says
anyway. I mean, I don't see where the
industry modification makes that much of a difference if, in fact, they take
that out at the level of the slaughterhouse they take out that
requirement. The way I read it.
BOARD
MEMBER FERGUSON: Well, I guess from an
interesting point of view and actually let me rephrase that. From a Government point of view, as a
federally employed Government veterinarian that might be put in a position to
certify to this, I probably couldn't.
And it is just because of the way that that is worded, where this stuff
is removed directly after slaughter.
CHAIR
PRIOLA: Where does it say directly
after slaughter?
BOARD
MEMBER FERGUSON: Right in the text,
yes. If the slaughterhouse removes the
head, spines and spinal cords directly after slaughter.
CHAIR
PRIOLA: I just have after
slaughter. Do I have the wrong one?
BOARD
MEMBER FERGUSON: Can we put it up?
CHAIR
PRIOLA: Oh, I see. You're looking in the -- I see. It says directly after slaughter if it's
from a BSE herd. Later in the
recommendation it says if the slaughterhouse removes after slaughter. So there is two different ones.
BOARD
MEMBER FERGUSON: Yes, but even, I mean,
the later one remove head, spines and spinal cords as a first procedure
following slaughter, that just leaves open a lot of ambiguity and, you know,
there are some of our folks who are very literal, you know, when they would
read that and say oh, no, they didn't, you know, stun this animal, bleed her
out and then immediately remove things, therefore, I can't attest to that type
of certification.
CHAIR
PRIOLA: I guess again, could we ask
FDA, is there -- since that's a USDA interpretation of this recommendation,
does the FDA have the same sort of reservations or are they concerned about
those same sort of reservations as to when exactly the tissue is taken after
slaughter or is the discussion enough?
DR.
ASHER: No, I think the discussion is
very useful. My recollection of the
intent of the FDA with both those issues was that the reason why BSE-free herds
was specified but not defined was just to put the industry on notice that under
no circumstances did we consider material from a herd recognized to have BSE as
being an acceptable source for any kind of gelatin entering the United
States. No effort at the time was made
to define a BSE-free herd.
If
one were to try to define an acceptable BSE-free source, I would certainly
agree with Dr. Chiu that it would not simply be all tests of 30 month old
animals going to slaughter are negative.
The herd would have to have a certified history of never using food
supplements containing prohibited proteins.
There would have to be an adequate surveyance program, not just 30
months slaughter animals.
And
my personal opinion would have to include a sufficient number of older sentinel
animals and, of course, careful veterinary surveyance to make sure that all
sick animals were recognized. My
personal opinion also is that this Committee not entertain an assertion that an
animal that tests negative at 30 months poses no threat to the public
health. I say both those things without
attempting to influence the discussions of the Committee. Thank you.
CHAIR
PRIOLA: Would it be sufficient to say
something like a BSE-free herd is defined by the FDA, if that is in fact
defined somewhere, clearly?
DR.
CHIU: No, we have not put in
writing. And regarding the slaughter,
you know, the first procedure are directly after slaughter, I remember our
discussion in the past, was because if you remove spinal cord, it is not
possible, you know, to make sure entire cord, everything is removed. You might have residual, you know,
tissues. And if you carry that to
somewhere else and then remove the spine, then create contamination of other
tissues, in the bones of other tissues.
So we thought, you know, it would be better to remove, you know, the
spine, the vertebrae at the slaughterhouse.
That was the thought at that time.
CHAIR
PRIOLA: I guess the other thing to
consider is, again, given all the data we have seen showing inactivation of
infectivity following the gelatin extraction process, the issue of
contamination, cross-contamination by a spinal cord being removed at a
different part of the slaughter process may not be as major an issue given the
fact that now there are these five individual studies, all of which saying that
the gelatin process itself, as you get to the end, can remove extremely high
levels of infectivity under worst case conditions. So it's possible that this discussion as to when things are
removed and may not, given that data, be as critical as it might have been
before we had access to this data. Dr.
Bailar?
BOARD
MEMBER BAILAR: Dr. Priola, we have had
questions about some of the wording in this recommendation, this draft
recommendation. I have a question about
the last sentence that the processors are responsible for the safety of what
comes into them. Without offering any
guidance about that, would it be appropriate?
I don't want to vote against this.
On the other hand, I'm not very comfortable about voting for it.
Would
it be appropriate to defer action until the next meeting with a request that
FDA consider revising the wording? I
think the intent is fine. I have no
particular quarrel with the intent of the changes proposed by the industry, but
I think it needs some tightening up.
CHAIR
PRIOLA: Well, I think in a way that's
what the FDA is actually asking us to discuss.
Given what we have heard today and the current discussion, how can we
modify this or should we modify it in a way that addresses the concerns of the
Committee? So this, I would think would
be an opportunity to make that known, how you would want to do that.
BOARD
MEMBER BAILAR: I'm not sure we can
modify it on the fly this way. That's
why I would like to allow a little bit of time for people who know a lot more
about the process, the problems, than some of us on the Committee, and time for
some reflection about the implications of any changes.
BOARD
MEMBER WOLFE: I would agree with John,
because I think based on what Lisa has said, which, I think, amplifies the
understanding of the process somewhat and what other people are saying, that
the FDA has gotten some input from us, which is what Question 2B is, and it
would make most sense for us to get at the next meeting the new version of the
recommendation to vote on.
CHAIR
PRIOLA: Do you have suggestions for
changes that we can make? I mean, we
still have to actually vote on Question 2A, but would you have suggested for
recommendations?
BOARD
MEMBER WOLFE: I mean, defining, as Lisa
suggested, what a BSE-free herd is, sorting out the differences between
directly after, immediately after, first process or just after. I mean, there are three different ways of
describing in the current recommendation the timing between slaughter and
removal of spine, spinal cord and so forth, so I think, I mean, those are, I
think, two areas that need to be neatened or tightened up.
CHAIR
PRIOLA: Yes. Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yes, it's a question to
the FDA. Can we vote on this
recommendation and then leave to the FDA to wordsmith the BSE-free herd and the
timing of removal based on the discussion that they heard?
DR.
CHIU: I think we definitely can go back
to before BSE-free herd, you know. We
have some idea, you know, over the years, you know, we have in mind. We would like to get advised whether to
remove the spinal, the spine, the vertebras in the slaughterhouse is needed or
not because of the results you have seen, you know, from the validation
studies.
CHAIR
PRIOLA: Dr. Hogan?
BOARD
MEMBER HOGAN: Well, it seemed, the
validation studies suggested if you can start with really high titre material,
that you get rid of almost, I mean, virtually totally. So I think what you start with in the real
world is sort of irrelevant, because it's never going to be as high as what
they are starting with in these validation studies.
Now,
I am a little concerned that if you leave the spinal cord in and then you
drive, you know, 200 miles to have the spinal cord removed, it is going to dry
during that time period. Is that going
to sequester some agent that might be more difficult to remove later? But as I just said, I think the titres will
be much less than the validation studies.
So I personally wouldn't have a problem with that.
CHAIR
PRIOLA: Mr. Dwyer, do you have a brief
comment?
MR.
DWYER: Thank you. Just as a reminder, this Committee voted in
1998 to agree with or recommend to FDA the removal of the spines in the manner
that we have suggested in our draft modifications to the guidance. We have attempted to craft our draft
modifications to the guidance with respect to this issue, that is spine removal
in a way that is consistent with what this Committee recommended in 1998 as
reflected in the transcript of the meeting from April 1998. Thank you.
CHAIR
PRIOLA: Dr. Johnson and then Dr.
Ferguson.
BOARD
MEMBER JOHNSON: Well, I think it's very
impressive how much this does decrease the infectivity. On the other hand, we should go back to
remember that there is that inactivated tail or whatever you want to call it,
so there is inactivated particles, and think back to the Committee hearings
after the Cutter episode with Jonas Salk where they forgot, they neglected
looking at the unneutralized tail, which caused the whole Cutter episode.
Is
it there and I think we should consider that.
I don't think that's enough to change the rules, but I don't think it's
enough to say well, let's not worry about splashing a little spinal cord
around. I think we still ought to keep
that as tight as absolutely possible to keep the contamination membranes of the
spinal cord down.
So
I would agree that I don't think we need to change. I voted yes on 1, but on this I would not want to see it made
more permissive for the possibility of contamination despite the good
inactivation studies.
BOARD
MEMBER FERGUSON: I guess I would like
to briefly run through sort of the standard slaughter practice at least in the
U.S., and ask everybody to sort of think about the possibilities for
contamination. You know, an animal
comes in. It is stunned, rendered
unconscious, then, essentially, hooks are applied to the rear legs and it is
bled out. The animal is skinned. The head is removed and then, generally, the
carcass is split, at that point in time.
The
standard practice is to go ahead and remove the spinal cord, at that point in
time. That is the easiest time to do
it. But the issue is not necessarily
the removal of the cord. The issue is
the removal of the spine and that, you know, vertebral structure that allows
the carcass to sort of hold together and it's going through the rest of the
processing process.
So
if concerns are about cross-contamination from removal of the vertebral column
later in the process, I'm not quite sure where that cross-contamination is
going to come from, especially on bones and bone chips that are going into a
gelatin derived process, because even if you assume okay, you can get some
contamination when you split that carcass in half or if you have a missplit,
you're getting aerosolized cord that is going on the surface of that carcass,
and the bones aren't on the surface of that carcass. The meat on those bones is removed elsewhere in the meat cuts,
and the contamination isn't necessarily going to be on those bones, per se,
which is what is going into the gelatin process.
CHAIR
PRIOLA: I think we could -- at least,
the sense I'm getting is the recommendation as it stands needs some tightening
up in terms of clarifying definitions of BSE herd and when after slaughter
things need to be removed. The primary
question prior to this is do the scientific data and information available
support the following FDA recommendation?
If the answer is no, what changes?
Are there changes other than tightening up these definitions that anyone
would like to recommend?
For
myself, the removal of the vertebral column, I think, is a big issue for
European countries because of the European BSE, but given that here in the
United States there is as yet no BSE and they haven't yet moved, if I remember
correctly from this morning, to removal of the entire vertebral column, is that
right, that has happened. There is a
significantly different level of risk, if I understand correctly. So these rules seem to apply to really
primarily European BSE countries.
Should
we call for a vote on Part 2A and, if necessary, move on to Part B with
specifics? Are there any objections to
that? If the FDA has gotten what they
need from the discussion, which I think they have, we can move on to a vote for
2A.
SECRETARY
FREAS: I will go around and poll the
table exactly as last time. Dr.
Johnson?
BOARD
MEMBER JOHNSON: With the likely changes
made by FDA, do we vote yes or no?
CHAIR
PRIOLA: I think you vote no.
BOARD
MEMBER JOHNSON: You vote?
CHAIR
PRIOLA: And then we ask what changes
for 2B. Isn't that right? Well, actually, I think yes. Well, because I think that the
scientific --
BOARD
MEMBER JOHNSON: I would vote.
CHAIR
PRIOLA: Yes.
BOARD
MEMBER JOHNSON: If I looked at that
just as it says, which states there on the board, my answer would be yes.
CHAIR
PRIOLA: Yes. I think you can vote yes or no and we can still make
modifications in 2B, because this Committee has never hesitated to make
modifications.
BOARD
MEMBER JOHNSON: Then my vote stands,
Sue.
CHAIR
PRIOLA: So I'm sorry, so what is it
again, Dick, officially?
BOARD
MEMBER JOHNSON: It's a yes.
SECRETARY
FREAS: Dr. Bracey?
BOARD
MEMBER BRACEY: I would second that yes.
SECRETARY
FREAS: Dr. Ferguson?
BOARD
MEMBER FERGUSON: Yes.
SECRETARY
FREAS: Dr. Hogan?
BOARD
MEMBER HOGAN: Yes, but we need
modification.
SECRETARY
FREAS: Dr. Khabbaz?
BOARD
MEMBER KHABBAZ: Yes.
SECRETARY
FREAS: Dr. Priola?
CHAIR
PRIOLA: Yes.
SECRETARY
FREAS: Ms. Walker?
MS.
WALKER: Abstain.
SECRETARY
FREAS: Dr. Wolfe?
BOARD
MEMBER WOLFE: No.
SECRETARY
FREAS: Dr. Bailar?
BOARD
MEMBER BAILAR: No.
SECRETARY
FREAS: The industry, would you, please,
express your comments on this?
Okay. Out of the nine voting
members at the table, we have 2 nos, 1 abstention and 6 yeses.
CHAIR
PRIOLA: Okay. Under the part of 2B, even though we answered yes, if I tallied
right, there are three specific things that we would ask the FDA to clarify. And that would be the definition of a
BSE-free herd, to make the recommendations at slaughter, directly after slaughter,
you know, just after slaughter, if they could be more specific as to when the
vertebral column should be removed, and also, Dr. Chiu had asked specifically
about whether removal of the vertebral column is necessary. They wanted some clarification on that, too,
I believe. Are there any comments, Dr.
Bailar?
BOARD
MEMBER BAILAR: I would add a point also
about some clarification about the insurance by the processors that their
supplies are adequately protected.
CHAIR
PRIOLA: Well, I think that -- isn't
that in the last? That is in the last
sentence, right, gelatin processes should ensure?
BOARD
MEMBER BAILAR: It says the processors
should ensure, and I would like to know more about that.
BOARD
MEMBER HOGAN: Well, unfortunately, Dr.
Gambetti isn't here, but from my experience in removing spinal cords, there can
be left dorsal root ganglia and other nervous tissues depending on how you do
it. So I think the issue of vertebral
column if you're asking just for some comments, may be important if you want to
reduce that last little bit.
CHAIR
PRIOLA: Does anybody want to recommend
any specific language if we can, I don't know if we can, to give the FDA some
further guidance? You know, for
example, industry recommendation for BSE-free herd. Is that an appropriate way to qualify it, to introduce the
concept of testing according to standard procedures? Lisa?
BOARD
MEMBER FERGUSON: I guess I'm
uncomfortable with having testing in there as the only thing that's qualifying
the herd. You know, I don't think that
testing is necessarily the critical thing to hang your hat on. I think the point is lack of exposure, and
that should probably be where that definition heads.
CHAIR
PRIOLA: Are there any other comments as
to specificity as to modifications of the recommendation? Okay.
Sidney, do you have anything in terms of the slaughterhouse issue
directly after? I mean, how specific
should specific be, given again all the data we have heard this morning?
BOARD
MEMBER WOLFE: No, I understand that,
but, I mean, we heard that there is a vastly different slaughtering process in
Europe versus here, so what I thought I heard this morning from the Europeans
was that since it's done in a different place, it's not even within the
slaughterhouse. I mean, is that
correct? I mean, in Europe, you just
repeat what you said is the main difference between European slaughtering
techniques in terms of bone, in terms of getting the bone for gel, as opposed
to this country?
MR.
SCHRIEBER: The difference is only the
place of the removal of the bones.
BOARD
MEMBER WOLFE: The place, right.
MR.
SCHRIEBER: Just the place.
BOARD
MEMBER WOLFE: So it's not --
MR.
SCHRIEBER: Slaughtering practice is
exactly the same, I think.
BOARD
MEMBER WOLFE: But in one case, in this
country, the removal of the bones is in the slaughterhouse and there, somewhere
else? That's the difference. So it's the issue of transport and
whatever. So it's beyond just where in
the slaughterhouse. It's in the
slaughterhouse or not. The issue is
whether we think that it's okay for -- which is the issue the industry raised,
whether we think it's okay for the bone removal to be somewhere else with at
least risks to workers and others that are different than they would be if it
were all done within the slaughterhouse.
CHAIR
PRIOLA: Go ahead.
MR.
SCHRIEBER: In addition to this, the
places of the removal, which other meat processes are, are exactly under the
same supervision of the authorities of the public like the slaughterhouse
itself. They are meat processors, so
they have to follow the same rules.
It's just a question of distance.
It's not a question of how procedures are done, whether they are
inspected, whether they are controlled.
That's all the same whether it's here or there.
CHAIR
PRIOLA: Are there any other
comments? I guess I should ask the
FDA. Do you have sufficient information
in terms of what the Committee is asking for, for modification to the
recommendation based on the discussion and what was just said?
DR.
CHIU: Well, in my mind, I'm still not
quite clear, you know. We have read and
heard, you know, the study results and as many of you expressed, it's quite
impressive. So I am not quite clear,
you know, whether we get any advice. Is
it necessary to remove spine and if it's necessary, then when it should be
done? So if we can, you know, have a
little more discussion whether the spine, the vertebrae, actually needs to be
removed.
CHAIR
PRIOLA: Lisa?
BOARD
MEMBER FERGUSON: Yes, I guess I'll throw
my two cents worth in here and everybody else can have at it. I guess, I think it's important to
essentially limit the use of vertebral column in the production of gelatin or
say, you know, you are not using vertebral column in the production of
gelatin. I don't think it makes any
difference where or when that is removed, but to say yes, it's not included,
it's not going into the gel balm is important.
CHAIR
PRIOLA: Yes, I would actually agree
with that, that it's important that it is being removed given the data where
heard. Where exactly it's removed may
not be that big of an issue since you can activate, apparently, quite
effectively quite a bit of infectivity that might be residual on the bone
surface after removal of the spinal cord.
I
am actually comfortable if the FDA does tighten up the definition of BSE-free
herd. I am comfortable for myself with
the recommendation, how it sits, with just some tightening up of those
definitions, BSE-free herd, as well as being careful when you describe when the
vertebral column should be removed after slaughter. I think in Europe, all the vertebral columns are removed anyway,
so that is moot. It's just where the
removal is, and that is not a primary concern for myself.
Would
anybody else like to contribute? Is it
just too near to lunch? Are we running
out of steam? Well, if anyone has any
-- I mean, so I guess we have addressed the questions and if anyone else would
like to say anything after lunch, feel free to do that. When we restart the session at 1:30, 1:40?
SECRETARY
FREAS: Let's try 1:30.
CHAIR
PRIOLA: Okay. Let's go for 1:30.
(Whereupon,
the hearing was recessed at 12:45 p.m. to reconvene at 1:37 p.m. this same
day.)
A-F-T-E-R-N-O-O-N
S-E-S-S-I-O-N
1:37
p.m.
SECRETARY
FREAS: Okay. Thank you very much for rushing through lunch. In the afternoon, we are very fortunate. We will be joined by four new temporary
voting members. I would like to go
around and introduce them. I won't
introduce the whole table, just the four new temporary voting members. Well, I will introduce at least three of the
new temporary voting members.
On
the far side of the table is Mr. Terry Rice, Board of Directors, Committee of
10,000 from Windham, Maine. Would you
raise your hand, Mr. Rice? At the
corner of the table right in front of the screen is a new voting temporary
member for today, Dr. Charles Edmiston.
He is associate professor of surgery, Medical College of Wisconsin, and
he is also chair of the CDRH General Hospital in Personal Use Device Panel, and
he will be taking these issues from today to his center's panels.
And
we will be very shortly joined by Dr. Kenrad Nelson, who will be sitting next
to Dr. Priola, our Chair, and Dr. Nelson is a professor, Department of
Epidemiology, Johns Hopkins University School of Hygiene and Public Health, and
he is chair of the Center of Biologics Blood Products Advisory Committee. And
there is one more person, and that is also from the Blood Products Advisory
Committee. That is Dr. David Stroncek,
Chief Lab Service Section, Department of Transfusion Medicine, NIH. And to all four of you, I would like to
welcome you to the table and thank you.
CHAIR
PRIOLA: Okay. We'll go on to starting with Topic 2, which is BSE in Canada, and
I want to let the Committee know that this is an informational topic only. It's for our benefit. We're here to listen, and there are no
questions being posed to us. This is an
informational topic only. The first
speaker is Dr. Jay Epstein from FDA.
DR.
EPSTEIN: Thank you, Dr. Priola. Before Mr. Hills makes a presentation on
what is known about the reported case of BSE in Canada, I would just like to
take a moment to make a brief statement about FDA's current thinking in regard
to potential implications of that case report for blood safety policy.
FDA
is undertaking an assessment of the BSE exposure risk to blood donors in the
U.S. and Canada in light of the single BSE case that has recently been reported
in Canada. Although, it is premature
for the FDA to present any results of this assessment now, we believe that the
likelihood of exposure to the BSE agent for both Canada and the U.S. is and has
been very small.
The
exact magnitude of BSE risks for Canada and the U.S. will be difficult to
quantify because of methodological limitations. However, preliminary considerations suggest first that the risk
of exposure of blood donors in North America to the BSE agent has been
extremely low and is even lower now than it was several years ago. And then secondly, in particular,
implementation of the feed ban of 1997 both in the U.S. and in Canada
significantly reduced the likelihood of human exposure to the BSE agent for
both countries.
FDA
does not believe that there are sufficient data, at this time, to warrant
changing our blood donor deferral guidance.
However, we will continue to study this issue and will take further
action as appropriate. Thank you very
much.
BOARD
MEMBER WOLFE: I have a question.
CHAIR
PRIOLA: Dr. Wolfe?
BOARD
MEMBER WOLFE: Given that the spectrum
of countries for which there are limitations on blood donation go from UK, lots
of cases, to EU with some countries with very small numbers of cases in just
cattle, and that Canada is still a "moderate risk country" that has
had a case, the benefit risk equation is always important, and have you at
least tried to get some data, so that this question can be answered better
later as to what fraction of the blood supply in this country would be affected
if there was some sort of constriction on the ability of people who have spent
whatever amount of time in Canada?
Is
there at least some effort to collect that, because otherwise, as we learn more
about the possible risk, small though it may be, and the benefit, which is
having a blood supply that is not impaired some comes up to it? So just a simple question. Is someone trying to get a hold of those
kinds of data?
DR.
EPSTEIN: Yes, Dr. Wolfe. Thank you for that question. We are mindful of the need to try to assess
the impact on the blood supply of any potential change to our donor exclusion
policy and, indeed, we have already had dialogue with major blood organizations
on both the feasibility and scope of surveys that could establish the impact of
any candidate deferral policy related to Canadian exposure on the U.S. blood
supply.
And
more broadly speaking also, we are thinking about similar questions as they
might pertain to say exposure in Japan or other countries that have had case
reports of BSE in cattle. So that
enterprise is recognized and is ongoing.
BOARD
MEMBER WOLFE: So you are going to be
collecting data on what this impact would be?
DR.
EPSTEIN: Well, blood organizations have
been asked --
BOARD
MEMBER WOLFE: Right.
DR.
EPSTEIN: -- if they would collect such
data, and we have had preliminary statements of agreement.
BOARD
MEMBER WOLFE: Okay. Thank you.
CHAIR
PRIOLA: Any other questions for Dr.
Epstein? Okay. Thank you very much.
DR.
EPSTEIN: Thank you.
CHAIR
PRIOLA: Our next speaker will be Dr.
Robert Hills from Health Canada Ottawa who will discuss the review of Bovine
Spongiform Encephalopathies in Canada.
DR.
HILLS: All right. First, thanks very much for inviting me to
give you a little update of what the situation is in Canada right now with
respect to BSE. I will just wait for
the slides to come up. There is a fair
bit of information on the slides, so what I will try to do is go through it
relatively quickly. All right. Thanks a lot.
First
of all, I thought what I would do is give you a little bit of background to
what Canada has been doing with respect to BSE before we found a case in May of
this year. First of all, there was a
prohibition of the importation of products assessed to have a high-risk of
introducing BSE in Canada. There was
importation of meat and meat products only from countries that Canada
recognized as being BSE-free.
In
1990, there was a designation of BSE as being a reportable disease in Canada,
and any suspect case of BSE would be reported to a federal veterinarian. In 1992, there was the creation of the
National BSE Surveillance Program. In
1997, the same year as the U.S. did, as well, there was the implementation of
the feed ban of feeding rendered protein products from ruminant animals to
other ruminants. In the year 2001,
there was the creation of the Canadian Cattle Identification Program for Cattle
and Bison making it possible to trace individual animal movements from the herd
of origin to the slaughter.
Next
slide, please. This is just a quick
pictorial of sort of how Canada has approached it. Canada has adhered to the OIE guidelines on TSE risk
management. Up until our finding of the
case, we were considered to be provisionally free, and we have also done a risk
assessment that was completed in December of 2002 with respect to Bovine BSE
cattle in Canada, and in that risk assessment, we actually determined that the
likelihood of finding BSE in Canada would be remote. That has changed, but still remote.
Next
slide, please. Before we go on, what I
would like to do though is just bring you back a little bit in time, because we
did have a case previously. In 1993, we
did diagnose a case of BSE in a beef cow that was imported from the UK in 1987. The exposure of this animal to BSE occurred
prior to its arrival to Canada. The
index herd and all the UK animal imports were destroyed, at the time, and it
was subsequently determined that the UK herd, which was a source cow for this
particular animal, did have other infected animals, as well.
Next
slide, please. This graph here is,
again, a pictorial showing the importation of animals into Canada,
particularly, and we're it the North American Disposition of Imported UK Cohort
Members prior to the index case discovery in 1993. So there was importation of animals prior to our taking action in
1993 to eliminate those animals.
Next
slide, please. So getting on to our
first indigenous case. January 31,
2003, a 6 to 8 year-old downer beef cow from northern Alberta went to slaughter
to a provincially licensed meat facility.
Alberta Agricultural Food and Rural Development meat inspector condemns
the carcass as being unfit for human consumption.
At
that time, the head was collected and submitted as part of our National
Surveillance Program, the Federal Provincial Surveillance Program for BSE. And the carcass, at that time, because it
was condemned, was sent to inedible rendering.
Next
slide, please. On May 16th,
the testing was completed with a tentative diagnosis of BSE by the Alberta
Ministry of Agriculture. The sample was
then sent immediately to Canadian Food Inspection Agency's National Center for
Foreign Disease in Winnipeg, Manitoba where they also confirmed BSE, and then
the sample was subsequently sent to the Veterinary Laboratory Agency in
Weybridge, England, which is the OIE Reference Center for BSE. And on May 20th, they actually
confirmed our diagnosis of it being an actual case of BSE. Immediately upon notification, we notified
the OIE, in fact, that we did have a case of BSE.
Next slide, please. So what did we do from there? So we had a start in epidemiological
investigation. We basically broke it
down into three phases. The first
phase, we're calling that the case itself, which we'll call the animal trace
back, its immediate management, which we'll call the animal trace forward, and
the most probably origins, which is where did the animal get the exposure from?
Next
slide, please. This is a well used
graph. It is done in colors for a
particular reason. The red is the case
herd where the index animal was. The
blue line is considered the primary line of inquiry where we think the source
animal came from, which is in Saskatchewan.
The yellow line is considered the secondary line of inquiry. That was an Alberta line, and the green one
in the middle is, in fact, that we did discover that there was some commingling
with another herd. So those are the
areas, which we were tracing out as part of this investigation.
Next
slide, please. As I said earlier, the
index case was a 6 to 8 year-old Angus.
It was a member of a herd that was recently established within a two
year period between 2001 and 2002, and the animals that made up that herd were
from two farms. What we believe
initially from the age of the animal was that the expression of the clinical
BSE at this age offers the first epidemiological insight, which would probably
mean it was a low level BSE exposure given the age of the animal.
As
I mentioned earlier in a previous slide, the Saskatchewan blue line of inquiry
was the most probable avenue for which the positive animal moved to the Alberta
farm. That particular line of inquiry,
the animals were culled and depopulated and tested, and all tests were negative
by Prionics Western Blot and immunohistochemistry.
Next
slide, please. At the same time as we
were culling and depopulating and testing, we were also trying to confirm
through DNA testing the origin of the particular index animal. Unfortunately, the DNA testing did not come
back with a clear definitive result, and as a result, we needed to then move
down the second line of inquiry.
So
we proceeded with the depopulation and testing of animals in the Alberta line
of inquiry, which was that yellow line in the previous pictorial that I showed
you. We culled those animals and tested
those animals and all tests came back negative again from Western Blot and
immunohistochemistry. Even though we
didn't have a confirmed definitive answer for the DNA, it is still a probably
line of inquiry, and most probable one, is the Saskatchewan blue line for the
introduction of the animal to this farm.
Next
slide, please. The next phase, which
was the Animal Trace Forward Investigation, was to determine what would happen
with the animals that left the farm. So
there was movement. We looked for the
movement of the cattle from the index herd.
We looked at, as I mentioned earlier there was a green box, where there
was some commingling. We traced out
those animals. We culled them. We depopulated them. We tested them. We found that they were all negative by Western Blot and
immunohistochemistry.
Next
slide, please. So to summarize, the
Trace Forward and Trace Back Investigations, we had 15 premises that were
quarantined, an additional 25 herds were scrutinized and the tracing-out of
single animals or cohorts from the Saskatchewan line of inquiry.
The
trace out also included the identification and notification of the export of
five animals to the United States, which you all should be aware of, we did let
you know as soon as we found that out.
And in all, we had a culling of more than 2,700 animals, 2,000 of which
were 24 months of age and older, and all, as I mentioned earlier, have been
found negative by Prionics Western Blot and immunohistochemistry.
Next
slide, please. This is just a pictorial
or a graphic of the disposition of the carcass of the particular BSE index
case. It shows the yellow line, which
shows the line of investigation with respect to the use of the products for
laboratory testing. We have the lighter
blue line, which shows what happened with the processing of the hide. And then, again, we have the mauve or the
purple, which is moving the carcass to inedible rendering and what happened to
it from there. And as you can see, it
did get rendered into some meat and bone meal.
Next
slide, please. So we then looked at the
Feed Investigation. Since the index cow
was condemned unfit for human consumption, its carcass was sent to an inedible
rendering. And I would want to
reemphasize because it was sent to inedible rendering, there was no part of the
animal that actually went into the human food chain.
Next
slide, please. The carcass of the index
as I showed you in the previous pictorial, the carcass of the index case was
traced via Canadian Food Inspection Agency from the abattoir, the renderer, the
feed mill, the producer continuum to its direct allocation into pet food. And in the pet food case, we did find that
actually there was some pet food that actually was exported to the U.S. to
which we notified the U.S. when we found that out, and there was pet food in
Canada, as well. And there was the
production of meat and bone meal.
What
is important to take out of this though is the visit to the renderer and the
feed mills confirmed adherence to our feed ban or meat and bone meal feed ban
legislation on the product receipt, segregation, labeling and distribution. So there was no breach in compliance, at
that point, at the renderer. So it was
not labeled to be fed back to ruminants.
Next
slide, please. Further investigation
was the trace out of the feed to individual farms, and we did find that three
additional farms were quarantined when the investigation could not preclude
exposure of 63 head of cattle to the feed destined for poultry feed. In that case, it was evidenced that the farm
itself had allowed poultry feed to be fed to ruminants. The animals were culled and tested by
Prionics Western Blot and immunohistochemistry and, again, all those animals
came back negative.
Next
slide, please. This is just a
pictorial, again, of what I was describing before from the inedible material
from the index case going to the renderer.
It went up to pet food. It went
into poultry and some pet food, and there was the feed mills that we traced out
afterwards. So this is sort of managing
the risk of the disposition of the material from the index animal.
Next
slide, please. There were other
considerations that we wanted to take into account as we proceeded in with the
exposure investigation. We did look at
maternal transmission. We looked at
contaminated meat and bone meal used in feed products, particularly early risk
factors, any UK imports slaughtered prior to 1993 or other European imports. It was figured into our investigation.
We
looked at TSEs resident in other animals, CWD and scrapie as examples, and we
did look at the possibility of it being a spontaneous case. Our investigation right now is at the point
now, we are looking at feed products that are considered the most probable root
of exposure.
Next
slide, please. Again, this is just a
graphic again to illustrate the hypothetical foreign domestic exposures of the
index case, and it's just groups of, you know, what the possibilities might be
for the exposure of this particular case.
Next
slide, please. What we did find out in
the investigation is there was two potential meat and bone meal epidemiological
exposure roots that were identified.
The first was a feed concentrate and the second was a high energy feed
block. Both have incorporated meat and
bone meal, at some point in time. The
investigation did find though that the feed mill records and compounding
formulae confirmed that the meat and bone meal incorporated in both of these
products was curtailed in 1997 upon implementation of the meat and bone meal
feed ban.
Next
slide, please. So what can we
conclude? What we can conclude so far
is that discovery of BSE in Canada proves that the active surveillance and the
diagnostic programs were working, because we did find the case. Epidemiological evidence supports the
probability that BSE in this case animal was associated to exposure to infected
material through the feeding system, at some point, early in the animal's life.
Next
slide, please. What we felt that we
needed to do was to ensure that what we were doing was accepted and would be
recognized, so it was decided that what we would do is convene an expert panel
to actually go over our procedures and how we did it and what we were doing and
what actions we were going to be taking, and get their recommendations back to
us.
That
particular panel comprised of Ullie Kihm from Switzerland, Will Hueston from
the USA, Dagmar Heim from Switzerland, and we did have contact with Stuart
MacDiarmid from New Zealand, as well.
The first three met on June 7th to 9th and met
with the members of the Canadian Food Inspection Agency and Health Canada to
which the team was provided with an overview of the epidemiological
investigation. All actions taken to
date and the scope of the options and the measures being considered to adjust
domestic policies.
Subsequently
to that meeting, the team went back and did actually do a report, and what the
panel did find was that the -- they found that the risk management measures put
in place in Canada achieved the desired outcome. The surveillance did detect the case with BSE. The animal did not enter the food chain and
the measures in place have reduced the spread and amplification of BSE in
Canada.
Next
slide, please. They did come back with
some specific recommendations, however, for us to strengthen our current
situation. They did say that there
should be a prohibition on Specified Risk Materials in human food and animal
feed, including advanced recovery meat products, tighter controls on
non-ruminant feed, enhanced audit and compliance, strengthening the existing
cattle identification tracking and tracing systems that are existing in Canada,
enhanced disease testing and surveillance by increasing the coverage of fallen
and dead stock, downer and diseased animals, and to work at efforts to improve
the awareness among producers, veterinarians and the general public with
respect to BSE.
Next
slide, please. So what will Canada
do? Well, the government of Canada will
be responding to the recommendations of the International Team. We'll respond through our consultation
process with our provinces or territories, the Canadian industry, our U.S.
counterparts and other trading partners, and there will be a new policy measure
for Specified Risk Materials as being the first step.
Next
slide, please. I felt before we sort of
should go a little bit further, I would just give you a little bit of
background about -- because I'm going to be talking about Specific Risk
Materials Ban right now, which is the first step, that in Canada, 95 percent of
the slaughters is in federally registered establishments and the majority of
those animals that are slaughtered are between the age of 18 and 24 months.
5
percent of the slaughter is in provincial abattoirs and the majority of those
being over 30 months of age. Only
animals slaughtered in registered establishments can be exported. The provincial slaughtered animals can only
be traded within provinces and sold within provinces. If they are to leave provinces or to leave the country, they have
to be at a registered establishment.
Removing SRMs at the point of slaughter and disposing of them, we
estimate removes about 99 percent of the human exposure to potentially infected
material.
Next
slide, please. The immediate objective
of the SRM policy is to establish a requirement that the SRMs be removed at the
time of slaughter, and that they be removed from human use, human food and
human use. The new policy will define
Specified Risk Materials and require removal, as I mentioned earlier, at
slaughter.
The
list that I have right here are some of the things that we have been
considering as being the most probable.
They will likely include the brain, spinal cord, dorsal root ganglia,
eyes, tonsils, skull and distal ileum.
Next
slide, please. Following our first
step, as was recommended in the expert panel report, there will be other
measures that will be taken. There will
be areas that will be looked at with restrictions on animal feed and process
and protect human and animal health, that should be, expanded surveillance, as
was mentioned earlier, expanded food safety plans, comprehensive tracking and
tracing systems and national standards and approaches will be implemented in
Canada.
Next
slide, please. And that concludes a
very quick overview of what we did.
Hopefully, it did give you an idea of the scope in which we reacted and
what we looked at. I have listed here a
number of different sites that you can look at for updates. We're trying to be as open and as
transparent about our investigation or actions as we possibly can, and I would
encourage you to go to these sites and go to this to get the most up to date
information. That's it.
CHAIR
PRIOLA: All right. Thank you, Dr. Hills. Dr. Wolfe?
BOARD
MEMBER WOLFE: This morning, we sent a
letter to the Secretary of Agriculture, Veneman, strongly urging them not to
lift the ban on meat coming from Canada to this country, and one of the
questions we have was, and I will just read you three sentences, because it's
really the form of the questions.
Public
information regarding the enforcement of the Canadian Feed Ban, and we know it
went into effect in '97, but we also know the data from the United States show
very spotty and uneven enforcement, particularly, the first few years of the
ban, which is very similar to the U.S. ban and was enacted about the same
time. It's available, it's data
unenforcement. It's not on the website
of the Canadian Food Inspection Agency and a telephone call to the agency
requesting these data has not produced any information. Most tellingly, the report from the team of
international experts, which is, I assume, the one you just referred to.
DR.
HILLS: Yes.
BOARD
MEMBER WOLFE: That investigated the
Canadian government's response to the outbreak makes no mention of compliance
with the feed ban. It mentions the feed
ban, of course, but data incompliance.
DR.
HILLS: Yes.
BOARD
MEMBER WOLFE: It is simply impossible
to assess the wisdom of lifting the ban you wisely put in place, you, Secretary
Veneman, in this case, on an emergency basis without these data.
So
my question to you is what is your knowledge?
I thought your presentation was excellent. What is your knowledge of looking backward how effective the enforcement
of the feed ban has been from 1997 when it was imposed to now?
DR.
HILLS: Unfortunately, I don't have the
history to go back in time from '97 backwards or forwards. I know it has come up in our discussions
numerous times, being able to put some sort of quantifiable number to it and to
try to do that. I have not yet seen
that myself. We are trying to determine
that now, because we have had investigations.
We have looked at it. We have
found that our plants themselves have been in compliance.
As
I noted in our Feed Investigation, we did find though that there was a
possibility that if a farm was coproducing, that there was a possibility of
cross-contamination, if you want to call it that, and those are things that
we're trying to address now to improve.
BOARD
MEMBER WOLFE: Yes, I mean, needless to
say, it's the essential issue, because you have admitted that that's the most
likely place that the cow that got infected got infected from, and given --
DR.
HILLS: Well, no, that's not quite what
I said.
BOARD
MEMBER WOLFE: Well, I think it's all on
your slides, for the most part.
DR.
HILLS: No, I think what I said was we
found that there were three slides as part of the investigation from the index
case that actually found that there were three farms in B.C. that actually were
not in compliance, because they inadvertently had fed or that we couldn't
definitively tell whether or not the feed that was destined for poultry did not
end up having inadvertently been fed to ruminants. What I said was that our most likely possibility would be --
BOARD
MEMBER WOLFE: The feed.
DR.
HILLS: -- exposure before the feed ban.
BOARD
MEMBER WOLFE: For that particular cow.
DR.
HILLS: Yes.
BOARD
MEMBER WOLFE: But, you know, again, are
there going to be some data on enforcement?
I mean, I assume that once it went into effect, there was some kind of
government effort.
DR.
HILLS: Yes.
BOARD
MEMBER WOLFE: To check on enforcement.
DR.
HILLS: Yes.
BOARD
MEMBER WOLFE: When can we expect to see
those data?
DR.
HILLS: I can't give you a date on
that. I can certainly find out for you,
but I cannot give you that, but I am not aware of what date they are going to
be able to make that information available.
BOARD
MEMBER WOLFE: Okay. Thank you again, very good presentation.
CHAIR
PRIOLA: Dr. Johnson?
BOARD
MEMBER JOHNSON: Yes. Dr. Hills, a question that was brought up
early on was the possible U.S. origin of that cow, that it might be a North
Dakota or Montana cow, and it may have been our feed ban was the problem. Is there any further data on that original
origin of it, and has the U.S. origin been ruled out?
DR.
HILLS: The data we have right now
suggests that the line of inquiry was the source was the Saskatchewan farm
right now. We have no definitive
evidence that would say that it was an animal that was imported from the U.S.
CHAIR
PRIOLA: Dr. Gambetti?
BOARD
MEMBER GAMBETTI: Can you describe the
procedure that Health Canada uses to diagnose this particular animal and
suspected animal in general? You listed
Prionics Western Blot and the immunohistochemistry. Is that done in more than one area and in that particular animal
that turned out to be positive, were both positive? Can you, in other words, amplify a little bit on how the animal
or another animal are studied in Canada?
DR.
HILLS: If I understood your question
correctly, Canada has a national TSE laboratory network. The gold standard, the immunohistochemistry
test, is a test that is used, was used in all these laboratories, and so there
was, as I mentioned earlier, the National Surveillance Program.
The
Prionics test for Western Blot was brought in and we were evaluating it, at the
time, but it was brought in for use mainly because we had so many animals to
do, at the time, that we needed to find some mechanism by which we could up the
volumes, and at the same time, we feel that it was a mechanism by which we
could do validation testing on the Prionics test itself.
So
in doing that, the work was done in the Alberta and Winnipeg labs for the
Prionics test, but we're in the process now of looking at and getting that test
now distributed across the TSE laboratory network.
BOARD
MEMBER GAMBETTI: How many tissue
samples or brain area were examined, only one, the lower medulla or more than
one and were all of them, if there were more than one, were they all positive?
DR.
HILLS: I believe there was more than
one and yes, they were all positive, but I can't give you the exact number.
CHAIR
PRIOLA: Dr. Bracey?
BOARD
MEMBER BRACEY: Perhaps along that line,
I guess a question that comes up is the issue of with any test, there is always
the chance of having a false positive the more times you do an assay. How are you all, in essence, getting at that
and is there a plan to actually look at infectivity in some of these animals or
do you feel, in essence, confident enough with the assay system in terms of
eliminating that rare false positive?
DR.
HILLS: Well, that was one of our
concerns, was to go to tests other than the immunohistochemistry analysis, but
we felt that the information that we have generated now based on the culling
exercise that we have gone through with the animals that we have right now,
because each one of those animals was testing in parallel with the gold
standard test, that we felt that we're starting to now get the numbers that
would indicate whether or not the test is, in fact, what the manufacturer
suggests, which is 100 percent no false negatives. So we are generating the information now, and that is the only
way that we can do it.
BOARD
MEMBER BRACEY: What about the issue of
false positives though?
DR.
HILLS: Well, in doing the animals that
we're doing now, we have found no false positives, but that, as I said, was
only 2,700 animals. That is what we
have done. That is the claim of the
Prionics, I believe, is what they are suggesting is it is being 100 percent
accurate.
I
did mention that the animals that we were talking about were -- we were
targeting 24 months and older animals, because we still believe that there is
some possibility that the test will not work sufficiently well for animals
below that age, and so the testing system itself is probably a little
questionable for younger animals.
CHAIR
PRIOLA: Dr. Nelson?
DR.
NELSON: You mentioned that there were
2,700, I think, animals that were tested and found not to be -- not found any
positives, but how many were of the similar age to this animal? In other words, the infection in this animal
could have occurred six, seven years ago and not shown up if the animals
examined where younger.
DR.
HILLS: Yes, that's a very good
question. That's why when we were
looking at the culling and the depopulating, we were looking at the specific
herds, so we were getting an age distribution.
I can't give you the exact number of animals that were there, but
because we were targeting the animals older than 30 months, we would then take
into account some of those animals. And
some of those herds on the Saskatchewan side were breeding animals, so many of
them were older. I just can't give you
the exact number.
CHAIR
PRIOLA: Dr. Taylor, did you have a
comment?
DR.
TAYLOR: David Taylor from
Edinburgh. You mentioned how you
concluded that because clinical disease emerged somewhere between when the
animal was somewhere between 6 and 8 years-old, that this may be reasonably
construed as evidence of low level challenge.
Now, certainly, it is true in the UK that incubation period as we have
taken to broadly equate with age, because meat and bone meal was usually and
often only fed in calf food.
But
if the animal, in fact, received meat and bone meal A for the first time or
subsequently on several occasions after it was born, you can't really pin down
the incubation period. So it could
still be a high level dose if it got its meat and bone meal at a later stage.
DR.
HILLS: Yes, you are correct. I have no trouble with what you're saying,
but I will go back to our feed ban that we have in place right now. The fact that the feeding of animals back
and forth, the feeding of ruminant material back to ruminants is prohibited in
Canada, so the likelihood of that would be, in our estimation, remote, not nil,
but it would be remote. So yes, so that's
what we're saying is we do think it's preceding the feed ban, so that would
make it then '95, '96, somewhere around in there, and it is a possibility, yes,
at that time.
CHAIR
PRIOLA: Dr. Bailar? I'm sorry, Dr. Bailar, excuse me.
BOARD
MEMBER BAILAR: You mentioned the
Canadian system for identification, tracking and tracing of animals. How much help was that to you in your
investigation?
DR.
HILLS: It was significantly helpful for
us. Unfortunately, because we
instituted it in the year 2001, it really only was successful for animals that
were within that age group. Even though
all animals are tagged for movement now, what we can't do is really
definitively say, for example, the age of the animal, where the entire history
of the animal was, but for the younger animals, yes, we can do that. So after 2001, we can certainly trace it.
BOARD
MEMBER BAILAR: Can anyone tell us about
the present status of moves to have such a system in the U.S.? I'm sorry Dr. Ferguson isn't here now. I'm concerned, of course, about the
possibility that there might be a single case here sometime.
CHAIR
PRIOLA: Yes, and I'm not sure anyone
here can answer that. That is sort of a
USDA issue, not an FDA one, I think.
We're about to proceed to the open public hearing portion. Do you gentlemen have a couple of quick
questions? You have been standing very
patiently. Do you have a couple of
quick questions for the speaker?
MR.
HAFFENDEN: Yes, Paul Haffenden from
TerraCell. I would just like you to
comment. Several years ago, the
European Scientific Steering Committee assessed Canada and the U.S. as category
2 countries, equivalent risk, given the -- maybe you could comment on the
movement, high incidence of movement of animals between the two countries in
both directions, high incidence of movement of animal feed products between the
two countries in both directions, and then any comment on how you think that
might affect the adjustment and category risk now with this case in Canada?
DR.
HILLS: I don't think I can discuss
anything about how that is going to affect the categorization. I think that is something that somebody else
will determine, not me. But what I can
say is that given the trade between our two countries, there is movement of
both animals and feed across the border.
CHAIR
PRIOLA: If it's a very quick question.
MR.
BROOKLANG: Yes, Nelson Brooklang,
Ortech International, New York. You
made a distinction between federally registered and provincially registered
abattoirs and the age of cattle that are processed in those, and the fact that
provincial cattle don't get exported. I
wanted to ask whether blood byproducts used in the biotech industry, what I am
from, like Bovine Serum Albumin transferred, purified from bovine blood could
be collected from provincial abattoirs in Canada and sold in the U.S.?
DR.
HILLS: Good question. My recollection is that the material itself
is not. From the provincial licensed
establishments, the provincial government looks after that, and I do not
believe it then goes to the federally registered renderers, but I would have to
reverify that. I'm not 100 percent
sure.
CHAIR
PRIOLA: Okay. Thank you very much for your presentation. I will move on to the open public hearing
portion.
SECRETARY
FREAS: In response to our Federal
Register announcement, I have received two requests to speak at today's
open public hearing for this afternoon, and the first one is Mr. Wayne
Vaz. Would you, please, come to the
microphone and make your presentation?
MR.
VAZ: Good afternoon. My name is Wayne Vaz. I am representing Serologicals
Corporation. We are a leading supplier
of animal-based products of the global health care industry. We are based in Atlanta, Georgia with more
than 800 employees worldwide. We
greatly appreciate the opportunity to be here today to talk about the TSE
safety of our bovine products and their critical importance in global health
care.
Next,
please. Our goal is to raise the level
of awareness regarding the pervasive use of bovine products in the production
of life-saving drugs and other essential health care products. We want to present the facts according to
high safety and qualify of Serologicals' bovine-based products, and we would
like to work with the TSE Advisory Committee and regulators to further develop
industry guidelines to assure the continued availability of bovine products.
Next,
please. Serologicals is a global
provider of biological products and enabling technologies, which are essential
for the research, development and manufacturing of biologically-based life
science products. Some examples of our
products include antibodies, cell culture supplements, such as bovine albumin
and other products for diagnostic and research.
Next,
please. This is a listing of our
bovine-based products. We're focusing
on this, because these are the products that are believed to offer a
theoretical TSE risk. At present, we
have two manufacturing facilities, one in Toronto, Canada, the other in
Kankakee, Illinois. We have a third
facility under construction in Lawrence, Kansas.
Next,
please. Our bovine-based products are
used in the development and production of life-saving FDA-approved drugs, FDA
licensed diagnostics, medical devices and animal vaccines. Our bovine-based products are used in the
development and production of FDA approved biologics for treatment of cancer,
arthritis, Crohn's Disease, psoriasis, blood clotting disorders, spondylitis,
RSV and at least one genetically predisposed orphan disease.
In
the diagnostic area, our bovine products are used in the screening of U.S.
blood for key viruses, such as HIV and HCV, for screening of cancer markers and
in serological testing as a potentiator of blood typing prior to transfusions. In medical devices, our bovine products are
used in the production of a medical device that is used during surgery, and in
animal vaccine, our products are used for the cultivation of Leptospira, which
is used to produce animal vaccines for the treatment of Leptospirosis, which is
a worldwide problem in livestock. Also,
these bovine products are used pervasively in life science research as reagents
in the lab for protein assays and other lab assays like chromatography and
electrophoresis.
Next,
please. So in April of 2000, we
received an update, which was issued to manufacturers of biological products
from Kathryn Zoon, the former director of CBER, that essentially says avoid
using ruminant origin products derived from BSE-affected countries in the
production of FDA-regulated products that are intended for humans.
Make
sure you identify all the ruminant materials used in production of the
regulated products, and document the country of origin, and make sure you
maintain traceability records for each lot.
Of course, the purpose of this guidance is to minimize the TSE threat to
the public.
Next,
please. So this creates some regulatory
uncertainty regarding the products under development today that are made using
bovine ingredients. Also, there is a
risk of current production of approved drugs, which use bovine
ingredients. This may lead to a
possible interruption to the supply of these biotech drugs if BSE occurs in the
U.S. Bovine-based products provide
unparalleled performance. There has
been a few attempts to replace these products in cell culture, but they
typically result in lower productivity and higher costs.
Next,
please. I would like to switch gears
for a minute and talk about the prion infectivity clearance studies that
Serologicals has conducted. We scaled
down our manufacturing processes and we used a hamster-adapted strain of
scrapie agent as a model to emulate the BSE.
Like many presentations before, we used a 263K agent. We spiked known titres of infectivity prior
to key process steps, and using serum tenfold dilutions we titrated the
infectivity downstream to measure the ability of the intervening steps to
reduce infectivity, looking at that in-vivo using hamsters and looking at the
clinical signs, abnormal gait, tremors, ataxia or incoordination, also looking
at a histopathological examination of the brain tissue to confirm the clinical
diagnosis, and the characteristic protease resistance of the transformed
prions.
The
conclusion of these studies, if you look at our Bovine Serum Albumin product
line, our HS product line, looking at four process steps achieving a total
clearance of 16 log10, Bovine Aprotinin, a total of 17 logs and
EX-CYTE completing one manufacturing step to date, achieving 3.7 log10.
Next,
please. So we feel that these prion
clearance studies offer some objective evidence that these products are very
safe from a TSE risk. In summary, the
high safety and quality of our bovine-based products are summarized by the
following.
One,
many are manufactured from bovine blood, which is recognized as being low-risk
of TSE infectivity, according to the World Health Organization and the EC. Moreover, Serologicals use either plasma or
serum for added safety. One of the
theories is that it is believed that prions may reside in the cellular fraction
of blood, for example, leucocytes. We
only use bovines that are less than 30 months of age, and they are typically
less than 20 months of age.
According
to the DEFRA statistics in Great Britain, no BSE reported in cattle that is
less than 20 months. It is uncommon for
it to happen in less than 30 months. We
use only USDA-approved raw materials collected in USDA-licensed
establishments. All these products are
manufactured within an ISO 9002 registered GMP environment.
We
have completed and published prion and viral clearance studies, and this
compounded with the similar clearance studies that our customers have completed
that are producing these biologics, we also maintain EDQM certificates of
suitability, which again is an assessment of low TSE risk. We have a proven track record of safety in
global health care.
Next,
please. So this is an example of one of
the viral clearance studies that we have completed on our bovine product
line. Due to time constraints, I won't
get into this other than to say that we have demonstrated more that 6 log10
of bovine viruses.
Next,
please. So in summary, one,
bovine-based products are critical to the production of life-saving health care
products. Secondly, manufacturers of
FDA-regulated products cannot replace bovine ingredients quickly, easily or
economically. The high safety and
quality of Serologicals' products is supported by the low TSE risk raw
materials that we use, the controlled production and the research studies that
we have conducted that demonstrates robust virus and prion clearance ability of
the manufacturing process, and our track record of safety and success. We're pleased to work with the TSE Advisory
Committee to further develop the TSE risk guidelines covering these important
products to permit their continued use.
Next,
please. Some considerations. In addition to using low TSE risk raw
materials, why not recognize the value of prion clearance studies and let's
establish minimum acceptance criteria.
Let's have suppliers perform prion clearance studies to provide
objective evidence supporting the product safety. Also, why not consider prohibiting the sourcing from countries
with a high incidence of BSE, rather than just one or two cases?
And
finally, when it comes to setting policy, we would request that the FDA and the
USDA carefully weigh the impact to the end consumers, i.e., the patients,
producers of biomedical products, which are our customers and supply chain
producers like Serologicals.
Next,
please. Finally, we would like to leave
you with two contacts at Serologicals Corporation. If any of you wish to discuss this further, we would be happy to
do that. Our email addresses are
listed. Thank you very much.
SECRETARY
FREAS: Thank you for your
comments. Our next speaker for this
open public presentation is Dr. Merlyn Sayers.
DR.
SAYERS: Excuse me. Thanks for this opportunity to talk to
you. See if you can rustle up my first
illustration. It's a brave blood bank
of the talks in the immediate shadow of the regulators, so I am indebted to Dr.
Hills and to Mr. Vaz for giving me some narrative separation from Dr. Epstein.
No,
let's have the earlier slide, please.
I'm speaking to you as the CEO of Carter BloodCare, and that's the
community independent blood program that meets the transfusion needs of
something like 150 hospitals in the Dallas, Fort Worth area. I am also speaking to you as a former
chairman of the Blood Products Advisory Committee and as a consultant to this
group, and I only make those two comments to emphasize how keenly I appreciate
the challenges that the regulators confront and also advisory groups like yours
have to confront, as well.
By
way of a preface, let me have the first illustration. Something like 40,000 Americans donate every day and their health
history, their screening for markers of diseases that are potentially
transmissible by transfusion, their subsequent counseling if that counseling is
indicated, these elements constitute what is perhaps the largest public health
exercise in the country and might even be the largest public health exercise in
the world.
Now,
bear in mind that those 40,000 individuals that donate originally were some
50,000 individuals, close to 50,000. Of
course, many get deferred during the history and examination even before any
serological testing is done on those folk.
So what happens at a local level?
At
Carter BloodCare, at our blood program, last year we registered something like
270,000 individuals, 40,000 were deferred and they were deferred either
permanently or temporarily, as I say, even before testing. The majority of these deferrals certainly
are temporary deferrals, and they are attributable to medications that those
individuals might have been on. They
might have a low hematocrit. They might
have traveled to a malaria area.
But
those temporarily deferred individuals are of particular interest to us as
blood bankers, because potentially those individuals are individuals that we
could get back to continue their donation to the community's needs. We have been considering what has happened
to temporarily deferred blood donors for a long period of time.
May
I have the next illustration, please?
What is their subsequent conduct?
We looked to 500 temporarily deferred donors and followed them for two
years, and you can see from this illustration that 58.5 percent returned to
donate successfully. 8 percent returned
only to be deferred again, but 33.5 percent of that original starting
temporarily deferred group did not return.
So
for one third of temporarily deferred donors, that temporary deferral is so
discouraging an experience that those individuals resist all our entreaties for
them to come back and donate again.
Now, the likelihood of deferral is obviously proportional to the amount
of scrutiny that these individuals are subjected to.
So
let's have the next illustration. And
certainly, the amount of scrutiny that donors are being subjected to has
increased dramatically. You can see the
number of questions that donors were asked in 1988 and the number of questions
donors are asked in 2003. For those of
you that are donors, the donor history questionnaire does not list 160 separate
questions. It's probably closer to 40,
but each question has become so complex.
There are multiple questions.
There are questions within questions.
There are nested questions. So
what we now want are 160 pieces of information from donors.
So
where do these increasing scrutinies relate to our consideration for deferring
donors who have some geographic risk and our need to potentially exclude them
from the donor base? Let's show in the
next illustration. The number of donors
that we have deferred since the year 2000 now that we have introduced
additional scrutiny with regard to deferral for attempts to decrease the
likelihood of transfusion transmitted CJD.
You
can see that at our blood program in the Dallas, Fort Worth area, we have now
deferred something like 3,500 donors, and this sad tally is a significant
underestimate of the actual number of donors that have been deferred, and it is
an underestimate, because many of these donors are individuals who have paid
attention, taken heed of our broadcasts, our announcements, our publications
urging them about the new restrictions.
We have no idea of what that number is.
This number reflects only those individuals who fail to appreciate the
new restrictions that we are publishing and who came to donate anyway.
So
what sort of contributions might these individuals have made? Let's have the next illustration. The next illustration. Thanks.
How many donations have these 3,500 individuals made? Well, they have made something like 13,000
previous donations, and these are individuals.
These citizens are now people who are indefinitely deferred. They have obviously made important
contributions to the community blood program, and it is interesting that in
that conflict of interest questionnaire that was handed out this morning, you
were asked if you regarded it as important that citizens affected by decisions
are directly involved in the Advisory Committee process. And certainly, I believe that citizens, in
this case donors, are important and should be involved, but they frequently do
not get that opportunity.
If
you were to ask them how they respond to their temporary or permanent deferral,
let me show you some of the questions that have been posed to us during
counseling sessions when we have spoken to individuals deferred as a result of
some geographic exclusion.
Let's
have the next illustration. This is
what we get posed. What should I tell
my wife, my husband, children, my dentist?
What should I tell my family practitioner? Where can I get tested?
Where can I get treated? Will
this affect my medical insurance, my disability insurance, my life
insurance? Will this count as a
preexisting condition? Should I
reconsider having a family?
Let's
have the next illustration. Why do the
criteria change? Other donors have
asked questions along these lines. So
if I had been in the UK for one day short of three months, I would be
safe? If you're telling me that I can't
donate anymore, what are you telling patients who got my blood? Why didn't I hear about this from the
military? How many patients have got
this disease from my blood? How many
patients have got this disease from a blood transfusion anywhere?
Now,
these are tough questions and deferral criteria can be debated in the
relatively academic climate of these meetings, but justifications for deferral
that are acceptable here do not sit well when they are explained to the donor
community at a lay level. The donor
deferral process is essentially a contributor and an important contributor to
all those layers of transfusion safety that we recognize as being valuable, but
the process is also responsible for increasing numbers of former numbers whose
experience is marked by a sense of frustration and alienation.
And
this next illustration shows what has happened to the rate of permanent
deferrals amongst blood donors in our community. You can see that between 1999 and 2003, there has been a
threefold increase in the risk of permanent deferral of individuals in the
community. So just as we are concerned
about individuals who are temporarily lost and our difficulty with getting them
back, we are concerned, too, about the fact that the rate of permanent
deferrals significantly contributed to geographic exclusion is increasing so
dramatically.
So
in closing then, a few comments. For
many deferred donors, there is a credibility gap that our explanations just do
not bridge. And to avoid the risk of
transfusion safety being achieved at the expense of availability of blood for
transfusion, CBER's Blood Action Plan that was promulgated in '97, which
addresses increasing the blood supply and removing restrictions to safe
donation really needs our enthusiastic support and endorsement.
And
then lastly, and this sincerely is not meant as a plea for less safety or a
plea for less regulation, let me just say that it is easy to add eligibility
restrictions, and there are many recent examples, smallpox, SARS, West Nile
Virus, but the more difficult task of lifting restrictions that no longer serve
a purpose is a task that also needs to be addressed. Thanks.
SECRETARY
FREAS: Thank you, Dr. Sayers. Is there anyone else in the audience, at
this time, who would like to address the Committee on this topic before the
Committee?
MR.
FILLBURN: Charles Fillburn, Nutramax
Laboratories. I would like to ask Dr.
Hills, does he exclude the possibility that the lone BSE animal that was
observed is not due to a mutation? Have
you sequenced the gene?
SECRETARY
FREAS: Could you come to a microphone,
so our transcriber can record the comments, please?
DR.
HILLS: Bob Hills. Yes, we did look at the spontaneous
possibility, in other words the mutation of PrP gene. What I can say is that we did look at it. We have excluded it right now and for other
reasons, I really can't comment. There
are some publications coming out shortly with respect to that.
MR.
FILLBURN: Do you think it's possible
that it could arise again either here or in the United States simply as a
consequence of mutation?
DR.
HILLS: I --
MR.
FILLBURN: The reason I ask is we seem
to assume that the only way this can occur is through feed. If that's not the case, then we need to be
more aware that some of these restrictions that we're putting on may be
overkill.
DR.
HILLS: Well, I think there are ways
that we can look at the PrP gene to determine whether or not it is spontaneous
mutation based on that. Now, whether or
not you can determine whether or not you have sufficient testing power to
ensure that that one case you found is spontaneous or not, I don't know.
MR.
FILLBURN: Okay. I would just like to echo the comments of
Mr. Vaz that in how we react to the situation in Canada and our importation of
any products really has a dramatic -- can have a dramatic effect on health care
in the United States and it may be an extreme overkill. I would like to see the USDA and the FDA be
on the same page in how they try to treat this, that they be more realistic
about it, and demand more clearance work by processors who may be dealing with
these types of products.
SECRETARY
FREAS: Thank you for your
comments. Do you have a quick comment?
MR.
HAFFENDEN: I'll try to keep it really
quick. I would like to echo the same
comments, the same that was expressed by Mr. Vaz. We do collect, have up until May 24th collected
Canadian origin animal-derived blood products that are sold into the veterinary
and pharmaceutical industry. We have
collections in Australia and in the United States. These are critical supply raw materials, and I believe that we
do need harmony between USDA and FDA on guidance. We also do have an isolated herd, isolated BSE-free herd in
Canada and would like to volunteer to participate in a committee than can help
to set those guidelines and give some examples of what is really there.
I
understand why the USDA in particular and the FDA have to react quickly and
close doors, but I believe we need to put the resources shortly to analyzing
products on a product by product basis, not a global product entity, and reopen
those doors for products that are needed and critical.
SECRETARY
FREAS: Thank you for your
comments. There will be two more open
public hearings tomorrow, and at that time, we will be more than glad to
welcome your comments.
CHAIR
PRIOLA: This topic is open for
discussion by the Committee if anyone would like to make a comment or have any
additional questions. I know I have one
question that I actually forgot to ask Dr. Hills. You said you ruled out the possibility of this case having
originating in Saskatchewan or no, sorry, excuse me.
As
a consequence of exposure to CWD or scrapie, as well as as a consequence of a
spontaneous event, did you do that based on purely pathological assessment or
how did you come to the conclusion that this was not a case of a cow coming
into contact with a CWD infected deer or elk, a scrapie infected sheep,
especially since Saskatchewan is where the CWD is, right?
DR.
HILLS: Yes. That actually was a concern, which is why what we did when we
sent the sample over to Weybridge, we actually were asking them to look
specifically at the strain that they had in hand and to compare that with the
strains that they had and they saw no differences.
DR.
ROGERS: Ron Rogers, a little bit more.
CHAIR
PRIOLA: Okay.
DR.
ROGERS: I just wanted to say that some
samples have been sent over to a stacks group in the UK, and like they are
doing a differentiation on the glyco-civilization patterns, and so the CWD --
we had already previously been involved with them in some research activities
to look at CWD profiling, I guess you have to say at this stage. So this did have some positive material over
there already, and so this material also was brought over to sort of see if, in
fact, we can get those kinds of patterns.
CHAIR
PRIOLA: Okay.
DR.
ROGERS: So it's purely at the research
level that this was ruled out.
CHAIR
PRIOLA: So basically, right now,
pathological assessment, and you're doing the molecular assessment of the
PrP. Okay. Any other questions or comments?
Yes, Shirley?
MS.
WALKER: I find that it is interesting
that Canada is now moving to provide the restriction to add SRMs after
reviewing their new case, and we have just been asked to look at that language
in our present policy, so we might be cognitive of looking at and changing our
policy too quickly.
CHAIR
PRIOLA: Okay. If there are no other questions or comments from the Committee,
then I think we can move on to Topics 3 and 4, so the remainder of the
afternoon is going to be a general introduction to TSEs and decontamination of
medical equipment and facilities. The
first speaker will be Dr. David Asher from the FDA.
DR.
ASHER: Thank you. Well, it's a great pleasure to open this
session on decontamination of TSE agents, which has been developed jointly by
the FDA Centers for Biologics and Devices.
This topic is presented, next slide, please, in response to a request from
this Committee last year for more specific information before members felt
comfortable advising the Agency concerning appropriate decontamination of
tissue establishments where the TSE agents might be encountered.
Next
slide, please. FDA, of course, as part
of its mission, is responsible for helping industry to keep regulated products
safe and that includes keeping products free of pathogens, today's pathogen of
interest, of course, the TSE agent.
Properties of the TSE agents complicate those efforts. Thank you.
You have already heard some discussion about the context-dependency of
inactivation of TSE agents. I expect
you will hear more.
Scrapie,
of course, scrapie agent has not been completely inactivated after exposure or
after drying and then exposure to steam heat.
Fortunately, TSE agents are substantially inactivated in solutions of
sodium hydroxide, sodium hypochlorite and probably other chemicals. Hence, the World Health Organization
consultants have recommended decontamination in health care environments using
combined sodium hydroxide or sodium hypochlorite and moist heat. Some other authorities have doubted the need
for such harsh chemical treatments, and we would like the Committee today to consider
these different points of view.
The
situations for which we are soliciting advice today are for products regulated
by the Center for Devices, that is instruments and by the Center for Biologics
Instruments and Surfaces used in the production of tissue products and plasma
derivatives.
There
are, of course, somewhat similar though not identical situations that would
involve other centers, our Center for Drugs is particularly interested in
today's discussion, because, of course, some drugs have components of human or
animal origin. We are aware that some
of the problems involved in the production of food or feeds are similar, but
the contexts are really quite different, for example, of course, sodium
hydroxide poses certain problems in proximity to food products.
We
would certainly logically expect that some issues of the U.S. Department of
Agriculture, which regulates animal slaughter and meat production in interstate
commerce and the Environmental Protection Agency, which regulates water
affluence, including affluence from autopsy rooms, that they would have
relative issues, but this Committee is not advisory to those agencies, and we
do not solicit advice for those problems.
Next
slide, please. There is no question
that contamination of classes of products regulated by the Food and Drug
Administration have transmitted Creutzfeldt-Jakob Disease to human beings,
fortunately few, relatively few such cases have been recognized in the United
States.
Next. One such class of contaminated products is
reusable surgical instruments of which a contaminated cortical electrode shown
here in pieces is the best known example.
Next
slide, please. I am aware of only six
cases summarized here in which transmission of CJD has been plausibly
attributable to contaminated surgical instruments, and I would note that in
none of those was modern cleaning or steam water used to decontaminate the
instruments involved.
In
addition, at least two epidemiological studies of which I am aware have claimed
to demonstrate some association with previous surgery, though most studies have
not found that and the association is not particularly impressive.
Next
slide, please. You have heard that
inactivation by heating of scrapie agent is very much context-dependent. This is a slide of data from Bob Rohwer some
20 years ago demonstrating that scrapie infectivity in aqueous suspension was
reduced to the level of detection in less than five minutes at 121
celsius. That was carefully suspended
in aqueous solution. Although, at
boiling temperatures, although, there was also prompt reduction in infectivity,
a reduction, a resistant fraction, that's the term of art for Dr. Johnson, a
resistant fraction remained.
Next
slide, please. But when dried onto
surfaces, infectivity was readily detected even after autoclaving for an hour
at 134 degrees celsius.
Next
slide, please. Dr. Rohwer, who will be
our second speaker in this session, has confirmed that apparent-- or rather,
Dr. Taylor who will follow Dr. Rohwer has confirmed that apparent paradox, and
found that different strains of TSE agent in similar preparations, at least,
appeared to have differences in thermal stability. The conclusion of those studies were that in worst case
scenarios, autoclaving has not been validated to decontaminate all TSE agents
completely.
Next
slide. A number of factors listed here
must be considered in deciding whether there is a significant risk that a
contaminated instrument might transmit Creutzfeldt-Jakob Disease, including
infectivity of the contaminating material, the reduction in activity achieved
by cleaning and decontamination, and the root by which a susceptible individual
is exposed and Martha O'Lone will talk more about those things tomorrow. Our speakers later this afternoon and
tomorrow will address those and other issues.
Next
slide, please. Central nervous system
tissues for humans as for animals have been consistently demonstrated to be
infectious when assayed in susceptible animals.
Next
slide. But there is also -- next slide,
please. There is also a significant
though smaller likelihood that tissues of patients with Creutzfeldt-Jakob
Disease outside the central nervous system, and that's not just Variant
Creutzfeldt-Jakob Disease, that is typical sporadic Creutzfeldt-Jakob Disease,
will have some infectivity found and here are positive tissues listed.
Next
slide, please. Of course, fortunately,
most human tissues, fluids, excreta have never been found to be infectious.
Next
slide. Although, confidence in the
negative results is somewhat tempered because of the very small number of
samples studied, and the fact that the assays used were animal transmissions,
and there does appear to be some species barrier even between human beings and
primates, which might raise the level of the limit of detection.
Next
slide. Just a couple of examples of
really how small the number of tissues successfully studied has been.
Next
slide, and the next slide. In human CNS
tissues, the mean content of infectivity measured in the NIH series was
estimated to be about 105 primate intracerebral lethal doses per
gram, but note that one brain was found to be infectious at a dilution of 10-8,
and considering both that and the species barrier, it might be prudent to
assume such high levels of infectivity for high-risk human tissues, at least in
an occasional patient.
Next
slide. Because of the substantial
uncertainties attendant to the biology of the TSEs and the properties of the
agent effects of cleaning and decontamination, advice offered to public health
authorities in the United Kingdom and the USA concerning surgical instruments
has not been consistent, and I won't read these. But the UK CJD Incidence Panel has advised incinerating
instruments exposed to brains of patients with known CJD where our respected
authority, Bill Rutala, whose is going to speak later, has felt that cleanable
critical or even semi critical devices in contact with high-risk tissues of CJD
patients can be cleaned and sterilized by autoclaving either at 134 celsius or
121 celsius, etcetera. Our proponents
of both points of view are present here today, and we really would encourage a
discussion of these conflicting points or, at least, apparently conflicting
points of view.
Next
slide, please. The two FDA centers who
developed today's program have generally relied on advice concerning
decontamination that came from a consultation convened at the World Health
Organization in 1999 published the following year. That consultation was chaired by Paul Brown, who was then the
chair of this Committee, and our next two speakers, who are among the most
influential of the participants, are two of this Committee's former CDC
members, both of whom are in attendance today, were also in attendance at that
meeting. The consultation identified
recipients of potentially contaminated products as being the group of persons
at the greatest risk of iatrogenic CJD.
Next
slide, please. And they offered the
following general advice. They
acknowledged that decontamination is context-dependent and that one method may
not be completely effective in all circumstances. Cleaning facilitates decontamination using the best validated
methods available, essentially meaning based on actual pilot studies. And then they advised using what we call an
orthogonal strategy, that is using two different methods, methods based on
different physical chemical principles whenever that is possible.
FDA
staff might add that in choosing those orthogonal methods, that a method that
inactivates the agent is generally considered more reliable than one that
simply removes it, because when an agent is removed, there is always the danger
that it can be reintroduced back into the product of interest.
Next
slide, please. They recommended
single-use to instruments, destroying reusable instruments wherever possible,
but they acknowledged that there are obvious situations in which instruments
simply cannot be discarded, and that less effective methods than destruction
may have to be used.
Next
slide, please. The consultation
recommended a series of decontamination methods in order of decreasing probable
effectiveness.
Next
slide. I expect that they recommended
six such methods. I won't go through
them all here. I expect that they will
be discussed by several of the speakers later today and tomorrow, but note that
the first four all include the use of either sodium hydroxide or sodium
hypochlorite either with or followed by steam autoclaving.
Next
slide. The last of the six was to
autoclave 134 celsius 18 minutes with the caveat that in worst case scenarios,
that is where brain tissue has been baked, dried under surfaces, the
infectivity will be largely, but not completely removed.
Next
slide, please. And for surfaces or heat
sensitive instruments, they recommended sodium hydroxide or sodium hypochlorite
at room temperature.
Next
slide. A number of other treatments
listed here were dismissed as being inadequate.
Next
slide. We tend to think of the
decisions regarding effective decontamination in these four general categories,
and I suggest that it might be useful if the Committee addressed them in this
way, as well. The surface of these
situations are surfaces or instruments potentially contaminated with either
high-risk tissue or lower-risk tissue from a subject with definite or probable
TSE, and then the same categories for someone where TSE is not suspected.
Next
slide. We're fortunate to have with us
today Bob Rohwer, who has already spoken, David Taylor to review the general
principles of TSE decontamination and the basis for the WHO recommendations. Unfortunately, Dr. Philippa Edwards of the
UK, CJD Incidence Panel, is ill, but she kindly emailed a talk for us, and that
will be delivered by Pedro Piccardo, who is an alumnus of this Committee and a
most welcome recent addition to our CBER staff, dealing with TSE issues.
Bill
Rutala, who is consultant to the CDC on hospital infection control, will share
his extensive experience here in the USA.
Ed Rau will then report on interesting studies of incineration that he
and Paul Brown have been conducting.
And finally, today we'll end with Stan Brown of our Center for Devices,
and I, who will report some early results of models that we have been studying
based in part on the work of Charles Weissmann, who will speak to us tomorrow.
Next
slide. Then tomorrow the issue for
discussion will be presented for CDRH by Lillian Gill, Martha O'Lone and
Charles Durfor, and for CBER by Ruth Solomon and Dorothy Scott. Ellen Heck will review the needs of Eye
Bank. Christoph Kempf and Andrew Bailey
will represent the Plasma Proteins Therapeutics Association in discussing the
needs of plasma processes, including a study that they will propose.
Please,
note that, again, although the discussions will doubtlessly be of great
interest to other agencies of the U.S. Government, and especially to our Center
for Drugs, we do not solicit advice for these other agencies, only for
FDA-regulated devices, tissue products and blood products.
Last
slide, please. With that, I hope that
you find the program that we have assembled informative, and we anticipate
having useful comments during our open public hearing, and discussions and
votes by the Committee. Thank you very
much.
CHAIR
PRIOLA: Okay. Thank you, Dr. Asher. Our
next speaker will be Dr. Bob Rohwer.
DR.
ROHWER: Thank you, and let me begin
with an apology to those people who have heard this talk before. It is one that I have given to this
Committee actually in an earlier form of it in the past, and have given it
fairly frequently over the last few years, but I was asked to do it again just
because David felt that the place needed revisiting. And so we'll begin with the first slide. The main points I'm going to make in this
talk are that the susceptibility to inactivation of TSEs infectivity is within
the normal range for viruses and spores, but that the TSE infectivity is
resistant to disinfection or sterilization.
That may seem like incompatible statements, but I will try to show you
what I mean in just a minute.
The
susceptibility to inactivation is an intrinsic property of the agent, and this
susceptibility is normal. That's what
I'm saying, but the resistance is context-dependent and a property of the
environment of the infectivity.
Next
slide. The best sources for this at the
current time are these old papers of mine in Science and Nature, which
have the complete experiments behind the kinetics that I'm going to show you
here and this review, which puts it into a larger context.
Next. And then the WHO report, which Dr. Asher
just reviewed is also a very excellent source.
Next. I am going to begin by just talking about
the inactivation process itself. And
this is actual data taken from an activation process with the scrapie agent. This is was a hypochlorite experiment, I
believe. I have forgotten now. I put this together so long ago, but the
main points are the following.
If
I can have the next. We're going to
build this slide as we go through it.
The inactivation process, one way to think about this is if we think of
this is surviving fraction on this axis over here where we start with 100
percent survival, no killing. At 10-1
survival, we have destroyed 90 percent of the population.
Next
slide. So for example here, I mean,
here we're starting. If we convert this
to a 100 individuals, what we have got by the time we're here on this
inactivation curve is only 10 percent of the population left. We have killed 90 percent of the population
in these very first moments of exposure, and by the time we get to the second
log of inactivation, we're down to one out of 100 survivors. This is just by way of review of things that
you probably had in your elementary chemistry class, but we sometimes forget
this with time.
Next. And then if we look down here, we notice
that 90 percent of the kill occurs during this first tiny interval. Only 10 percent of the kill occurs during
this next interval. Only 1 percent
during this, a 10th of a percent, a 100th of a percent
during this interval right here. We're
getting less and less effect as we go along.
The vast majority of what is happening to this population is occurring
right here in the very first moments of exposure.
Next. And this is reflected by this component of
the inactivation, which is reflected by this line right here, and the
inactivation rate constant for this line is the inactivation rate constant,
which is defining the behavior of the vast majority of the individuals in this
population.
Next. Next.
This line describes a second component, and it is describing, next, a
much smaller proportion of the population.
About one in 10,000 of the individuals behaves like this.
Next. So what is going on here? We have susceptibility to inactivation as
defined by this initial rate of inactivation, is intrinsic to the agent. It is actually less complex and there are
fewer controlling parameters. Whereas,
over here, this population that is being inactivated at a different rate could
be gaining those properties in a number of context-dependent ways, and these
are different for each environment and they are much more complex.
It
could be due to the container. It could
be a factor. I mean, it could be any
combination of these factors, as well, but among the things that we have to
consider are the container, rough or smooth surfaces, reactive surfaces, porous
surfaces, cofactors like fats, proteins, oxidants, reductants, water, air, in
the case of autoclaving, buffers, pH, temperature can all affect the
inactivation suspension, whether what the source of the tissue is and its
composition, the procedure for making it, how it was homogenized, the dose
rate, various transfers.
And
then we have these kind of procedural problems with making these measurements
themselves, which are the accuracy limitations of the assay and its
reproduce-ability. This is a real issue
in the case of end point dilution titration of TSE infectivity, which is only
good to about .3 logs. And
cross-contamination is an issue, and it becomes an especially important issue
when we're talking about very low levels of survival at the very end of these inactivation
curves.
How
do we know that this survival isn't due to something that got transferred from
here, which has almost 100,000 times more infectivity in it? The way we know is we're very careful when
we do these experiments, but it's something that you have to be very careful
about.
Next. Next, please, and next. Let's go to the next slide. Click through to the next slide. Thanks.
In comparing agent properties, the properties that are intrinsic to the
agent are reflected in the initial rate of inactivation. That is when the vast majority is being
inactivated, and the interpretation is less complex. The size of the residual fraction is a complex function of environmental
parameters, and cannot be used to compare the intrinsic sensitivities of agent
strains.
This
is where I differ with Robert's perspective that he gave this morning. In other words, I consider these plateaus to
be a very important public health and agricultural problem in terms of
infection control, but they are not telling us that much about the intrinsic
properties of the agent. They are
telling us about the context of the agent.
They are telling us something about the context of the agent, and it has
been very difficult for us to figure out what that is.
Next. So let's consider an example from chemical
inactivation.
Next. This is a hypochlorite inactivation in which
we can see that. In the scrapie curve
right here, we have -- this is surviving fraction, the same kind of curve I
just showed you minutes ago down here, that on contact with hypochlorite, .5
percent, this is a normal concentration, which bleaches use, and we get an
initial very rapid killing down to about 3 logs, but then we hit a plateau and
there is at the level of 1 part per 1,000 or a 10th of a percent, we
have got something in this population that is protected from hypochlorite. It's not seeing it.
By
the way, we checked. The available
chlorine did not change significantly over the course of this infection. We did this same experiment with some controlled
viruses. These are bacteria
phages. They are non-involute
viruses. They are very easy to assay
and they are reasonably robust in some ways and not others. But here is PhiX 174 showing the exact same
phenomenon. It plateaus at a lower
level. FD and M13 like phages doing the
same thing over here.
Here
are these two. These two viruses were
spiked into the same kind of normal brain homogenate that the scrapie brain was
in, and they exhibited this behavior in a purified form in PDS. They were inactivated to the limit of
detection almost instantly on contact with bleach. Another example of context.
Next. Next, please. Well, two of the things that we are going to discuss here are the
things that work best for TSE agents, and bleach is one of them, and I hope
that David will be sharing. He has a
lot more data on bleach than I do, and I hope he will be sharing that with
us. But sodium hydroxide is something
that I have been pursuing for a long time, and this was an experiment a long
time ago with Paul Brown, one of our initial experiments, comparing CJD and 263
scrapie.
Again,
in a very highly dispersed 10 percent brain homogenate of these two infectious
agents, and adding sodium hydroxide at these concentrations, and I would just
ask you to concentrate on these first three lines here. At 60 minutes with one normal, we had
limited detection killing here and here for both CJD and scrapie. By 15 minutes, we had almost as much
inactivation. A 10th normal
did almost as well as one normal. It's
a very effective method.
On
the other hand, next slide, please, this is a table of -- it's now out of
date. There are more experiments would
could be put on here, but at the time I made this, these were all the sodium
hydroxide experiments that were in the literature, and we got very, very good
inactivation by sodium hydroxide, but there are examples here. These are the experiments that we had done,
at that time, but there are examples here where there is some activity left
after considerable amount of exposure, and that always was very puzzling to me,
but my guess is that it has to do with how the stuff is presented and
homogenized.
Next. I went back and revisited. We have revisited this subject with a
kinetic experiment on sodium hydroxide, which is presented here. And in this case, the black circles indicate
infectivity, and this is time of exposure, and we're seeing something that is
very similar to the sodium hypochlorite effect, except much more dramatic
even. We're getting a huge reduction on
contact, essentially, with sodium hydroxide.
This is the point that was taken in the shortest amount of time,
interval, that we could effect between adding the sodium hydroxide and then
adding the acid to neutralize it, and then taking the points. So it's right around 30 seconds. This is two minutes, etcetera.
But
on this same curve, I have got two other plots. One is a plot, which I am labeling denaturation in one
hydrolysis. And what do I mean by that? Well, we went back later and used a Western
Blot on these samples to see whether we could recover Western Blot signal or
not from these various fractions. And
the Western Blot, especially at the time that we did this, was not as sensitive
as the infectivity assay, so we couldn't detect it over as long a range, but it
was very clear that upon contact with sodium hydroxide, we destroyed the pk
resistance of PrP. It was gone. It was showing the same inactivation
kinetics, essentially, as the infectivity.
Whereas,
if we didn't pk digest and we just put the stuff on the gel to see whether
there was anything left, it also disappeared, and this is a disappearance by
hydrolysis. The protein is being
hydrolyzed. It no longer shows up on
the gel, and it is showing quite different kinetics. So one of the points that we can take home from this is that to
the extent that infectivity and PrP are related and the prion protein are
related, and I am not entirely convinced of that, but nevertheless, to the
extent that they are, it's denaturation that is the correlate with inactivation
of infectivity not hydrolysis. This is
basically good news, because it's much easier to denature something than
hydrolyze it.
Next. Heat inactivation will be the next topic.
Next. This is a -- I have just taken the 121
degree autoclave experiment out of that family of curves that Dave just showed
you and that I showed you earlier in the day, because it makes the points best
in my opinion. Here is a case where, at
the time we did this experiment, the story was that you couldn't kill this
stuff with autoclaving, you know, that 121 degrees was not sufficient to
destroy the infectivity from 263K hamster scrapie.
This
is a kinetic experiment showing that, and this was done not in an
autoclave. It was done in an oil
bath. The samples were sealed in
ampoules. They were plunged into the
oil bath, so that we could control their -- and I was using thermistors, at
that time. The temperature was being
recorded, so I knew when they got to 121 degrees. I knew what the ramp time was.
I had that on my recorder, and we could control the actual time of
exposure within very narrow limits.
So
this first point on this curve was taken after the 58 second ramp time to 121
degrees, so it had just got there. By
the time it got to 121 degrees, we had already destroyed 99.9999 percent of the
infectivity in that sample. On the
other hand, it took another 10 or 15 minutes to get to the limit of detection
of the infectivity. There was a
residual population that took longer.
And
this is a concern, and this was a very highly dispersed sample again of
infectivity, and you get quite a different story, next, please, if you do this
type of experiment. Now, I think David
is going to be showing a lot more of this in a few minutes, but later on, David
Taylor started doing these experiments using brain macerates. Now, this is not a homogenate. This is a mush of brain. It is not dispersed in fluid. It's a paste, basically, and it is being
exposed at these various temperatures in the autoclave, and this is the
untreated sample, and these samples are getting incomplete killing even after
these very extreme treatments. I mean,
this is quite extreme for steam inactivation.
Well,
you definitely have to say that this poses -- you wouldn't want this on your
scissors when they go back into the next patient, for example. This is an extremely important public health
result. On the other hand, what does it
tell us about the agent in what we're dealing with? Well, you can get a titre out of this, because you're at limiting
dilution here, and we can do something we call a parson, we get a parson titre
out of this type of sample. This is how
we make our measurements in low titre blood samples. And I have done this
on the next slide and just put these figures on next. This is where these samples would fall on this curve that I just
showed you. There has still been an
extremely high level of inactivation associated with these, but you have got
survival going all the way out to 134 degrees here, at 134 all the way out to
60 minutes for some of these samples that were done in these macerates.
Personally,
I think we're talking about the same story here. It is just a matter of what we're talking about, and the context
has been ramped up in the case of macerates versus homogenates, and the
survival lingers for longer periods of time.
Next. Next, please. So what are we dealing with here? These could be intrinsic differences, and that was a question
that came earlier in the day from Dr. Bailar and it's a legitimate one. I think it needs more study. A lot of us have this on our books. Robert said he has been planning to do this. I have been doing it. I have got these samples in the
freezer. I just have not gone back and
redone this experiment, redone the kinetics on these, but it is on the
books. Someday, it will get done.
But
when you talk about these heritable differences, the point that I want to
emphasize is that I feel that they have to be discussed on the basis of
inactivation rate, not residual infectivity.
And my own prejudice is that the rates will be exactly the same, because
what we're dealing with here is context, not intrinsic differences.
Aggregation
is another issue. This is something
that I was very interested in early on in my career, but I think we have this
under control at the moment with the way we are homogenizing and dispersing
things, and aggregation would give you a recognizable difference in the
inactivation kinetics. It would not
look like first order. It would be
first order with a delay. There would
be a delay in something like that contributing to that.
The
most likely reason for this, in my opinion, is compartmentalization. The inactivant is not actually reaching the
infectivity, and our challenge before us is to find ways to open and destroy
this compartment to get at the infectivity.
Next. I just have a couple more here. So if we compare these two moduses of
investigation, what we are using is 10 percent homogenate sonicated highly
dispersed versus whole brain macerate.
This is sealed in a serum bottle.
I can't remember, David. Do I
have this wrong? I think David will
correct me if I have got this not exactly correct here on how he has got these
set up. We are using an oil bath versus
an autoclave. Our samples were being constantly
stirred while we were inactivating them versus static. And, in fact, this is kind of an idealized
type of inactivation to get at the properties of the phenomenon. Whereas, this is a worst case scenario,
which gets at the worst case problems that might be confronted in the public
health or agricultural context.
Next. Okay.
Now, we had some dry heat data earlier in the day from Robert
Somerville, so I'm not going to go over this, except to say that if you dry
this material onto a surface, the inactivation properties become completely
different. It becomes much, much more
resistant to inactivation. However,
this isn't a completely unfamiliar phenomenon.
It happens with spores and it happens with other microbes, as well.
Next. And so, in fact, my own interpretation of
this in a nutshell is that what is happening in these experiments and where the
source of residual infectivity may be coming from in our ampoule type of
experiments is that as we stick our ampoule into the oil bath, it boils and
flashes off immediately, and we throw things up on the walls and they dry. We get little specks drying on the
walls. I was very religious about
trying to recover everything when I went back to reanalyze this material, so I
scraped the walls and got everything back into the test tube.
And
what if what is happening is we have the infectivity in a form in which it is
basically anhydrous. We have little
drips and drops here that end up in little droplets of fat. Fat when it is oxidized becomes a varnish,
which is, essentially, a plastic. And
so, basically, what we're subjecting this to is a dry heat sterilization at the
rate of parts per million in our case.
It's not something that's happening very often, but we create a dry heat
environment for a very small part of this infectivity, and that is what is
escaping. If the reagent can't kill it,
if you can't reach it, you can't kill it.
The
other example I like to give is that if you put brain homogenate in a Zip Lock
bag and throw it into one normal sodium hydroxide, nothing will happen to that
either. And so it has to be available.
Next. Next, please. Next. Not that one. That's not supposed to be there. So the point I want to make here is that 132
degrees uses a significantly higher temperature than 121 for steam
sterilization where the inactivation takes place in minutes or even seconds,
but 132 degrees is only incrementally more effective than a 121 degree
centigrade environment for dry heat sterilization where the inactivation takes
hours to days at those temperatures depending on what you're talking about.
So
this does form, I think, a rationalization for what we're seeing in this
situation, and it also tells us -- and this was the rationalization for trying
to remove all headspace from those devices in which we did the gelatin
inactivations that I showed you this morning.
We didn't want any opportunity, any place for drying to occur.
Next. I think there is just two more. Steam sterilization, the agent is not
intrinsically resistant to steam sterilization. There are problems with delivery.
Next. And for effective delivery, we recommend
surfactants, homogenization, high levels of dispersion, eliminate sanctuaries,
agitation is helpful. My guess is that
a refinant will also reduce the potential for protective associations and will
improve the ability to inactivate.
Next. Prevent drying, immerse in water prior to
enduring steam sterilization and combine two or more methods. And the processing details can be critical. Adhere closely to validated approaches, and
this is referring to this stainless steel result we'll hear more about
tomorrow.
And
where we should go with this. We need
to know more about the underlying principles of resistance, and we definitely
need more robust methods for sterilization, which will actually get at these
last little bits of infectivity.
Why
don't you end right there, and let me just end by saying that the way we
inactivate in the laboratory, our own instruments, is for stainless steel and
things that can take it and things that are recycled and go back into animals,
immediately after use they go into one normal sodium hydroxide. They are immersed in one normal sodium
hydroxide for at least an hour, and then if they can take it, they are put
through the autoclave under one normal sodium hydroxide. They are cleaned after decontamination under
those conditions, and then they are reprocessed in sterile packs back into the
facility for further use.
CHAIR
PRIOLA: Okay. Thank you very much, Dr. Rohwer.
Are there any questions before we move on to Dr. Taylor? Okay.
If there are none, we'll go on.
Oh, I'm sorry. David, go ahead.
DR.
ASHER: Can you comment on aluminum
vessels, please?
DR.
ROHWER: I didn't hear that.
DR.
ASHER: Aluminum vessels.
DR.
ROHWER: I still didn't hear it.
DR.
ASHER: Can you comment on the use of
aluminum vessels?
DR.
ROHWER: Oh, yes, right. We use a lot of sodium hydroxide in our
environment, and we learned early on that you don't mix sodium hydroxide with
aluminum. And, in fact, aluminum and
sodium hydroxide in an autoclave can explode and can be quite dangerous, so you
have to be very careful about that. So
we, essentially, have no aluminum in our BL3.
CHAIR
PRIOLA: I think I'll check our
BL3. I'm not sure if we have
aluminum. Dr. Taylor, if you
would? Our next speaker is Dr. Taylor,
and he is going to talk about decontamination of TSE agents and the WHO
recommendations.
DR.
TAYLOR: Thank you very much. Well, thank you for the invitation to speak
this afternoon. As you can see, coming
from the UK, I'm using thumb roll technologies, slides and overheads. I was warned there could be problems with
the electronic system, so I didn't bother with the front-line. I just brought the backup.
As
has already been discussed and as this group will appreciate, there has been
accumulating evidence over decades that TSE type agents are remarkably
resistant to a wide variety of decontamination methods, which are quite
effective with conventional microorganisms.
This does not mean to say that these methods have no effect, but rather
that they are impractical for usage in medical settings, etcetera. These include things like strong oxidizing
agents, phenolic disinfectants and even ionizing radiation.
Because
of this general resistance, there have been some known examples of iatrogenic
transmission where instruments or devices that were in contact with the brains
of CJD infected individuals went on to cause accidental transmission in
subsequent patients despite having been processed in some fashion or another.
Now,
I use the phrase in some fashion or another advisably, because the methods that
were used would not, in fact, be used nowadays, but David Asher showed you the
x-ray of implantation electrodes, which would be put into a marmoset to look
for infectivity after this was suspected of causing this disease in humans
through accidental transmission. In
this case, the electrodes were washed in benzine and in a well meaning exercise
to try and sterilize them, they were then exposed to alcohol and formaldehyde,
which we now know is not terribly good as far as TSE agents are concerned.
The
second example, which David Asher also listed, was instruments used on a
suspect case of CJD, neurosurgical instruments, I should say, were exposed to
hot air, 180 degrees centigrade, for two hours before reuse, and there was
transmission from patient to patient.
Now,
as I said, and as David Asher referred to, there is actually no convincing
evidence that we have seen accidental transmissions through neurosurgical
instruments, but some data suggests epidemiologically that there is perhaps
some evidence of this, but there is no hard and fast evidence.
Nevertheless,
with such dreadful diseases that are incurable, untreatable, there has been a
constant nagging doubt about transmission of CJD. To some extent, this was aggravated when Bob Will reported in the
UK the emergence of Variant CJD. As you
know, the number of cases has risen into the hundreds now and is still mainly
confined to the UK. The worrying aspect
of that, of course, was that the work of Moira Bruce clearly demonstrated that
the agent causing Variant CJD was identical to the BSE agent in cattle and
quite dissimilar to any other TSE agent that had ever been discovered.
Concerns
regarding accidental transmission of Variant CJD between patients was elevated
by the finding that New Variant CJD lymphoreticular system tissues in the
patient examined, infectivity or at least positive PrP was detected with 100
percent of these samples compared with nil percent of the iatrogenic sporadic
cases that were examined or in other controls.
That was a study here.
We
also know that in a limited number of studies, if you have archival tissue from
patients who end up with Variant CJD, in this case, appendix. You can find PrP in the appendix at the time
when the patient had no clinical signs of disease. So the potential for accidental transmission through surgery is,
at least, in theory enhanced by the fact that surgeons compared with
neurosurgical would much more commonly be invading lymphoreticular tissues
either deliberately or incidentally.
Now,
I would like to just show a few overheads if I may. Both David Asher and Bob Rohwer referred to the WHO meeting in
1999, which resulted in guidelines being issued. It was related to not only clinical aspects of CJD-like diseases,
but also to concerns for the practical issues, such as protection of laboratory
staff, pathologists, surgeons, etcetera.
Now,
within the guidelines, there is this table here, which you may not all be able
to see, which is almost a short form of what I started with, talking about
ineffective methods. And the only thing
I would say here is that I will go in to talk a little about this procedure
here, which is regarded as variably or partially effectively boiling in 3
percent sodium dodecyl sulfate, SDS, because this has been commonly banded
around as a probably relative effective procedure.
In
terms of the actual processes recommended, and David Asher did show you a
summary of this, incineration, I will say nothing about, because there will be
something said about that coming up shortly.
These procedures, they were based on what was known from the literature
on TSE inactivation at the time of the meeting. To my knowledge, not much has happened since then to alter the
views and recommendations in these guidelines, and they are listed in their
perceived order of effectiveness.
So
we start with emersion in sodium hydroxide and heating in an autoclave, as
opposed to going on here to immersing in hydroxide then transferring into water
and going on to autoclaving. Also,
here, the alternative is to immerse in sodium hypochlorite, and then going on
to autoclave.
Here,
we have emersion in hydroxide or hypochlorite, and then going into an open pan
and then autoclaving. This is because
one of the options here is the 134 degree centigrade porous load cycle in which
you cannot put fluids. So if you're
putting instruments through these after the fluid treatment, you must remove
them from the fluid.
We
then go on to suggestions for boiling.
These are listed in order of decreasing perceived effectiveness, bearing
in mind that WHO recommendations are, essentially, for the health community
worldwide, and that facilities and equipment availability will vary
tremendously, especially in some more deprived areas of the world.
Finally,
we go on to talk about autoclaving at 134 for 18 minutes. And then when it comes to things like
surfaces, we revisit procedures like sodium hydroxide and sodium hypochlorite. Then you can just do thorough cleaning if
you can't do anything else. And then
there are some questions about dry goods and autoclaving.
So
I have been asked to address or discuss with you the data, in a sense, that we
use to back up these recommendations, which I will do and finish with one or
two bits of additional, perhaps anecdotal information.
Will
you go back to the slides now, please?
I did mention that I would talk briefly about SDS, because simply
boiling in sodium dodecyl sulfate or concentrations as low as 3 percent has
been widely, well, fairly widely recommended as a very effective
procedure. However, in our own
experiments where we used 5 percent of this compound and even went on to
autoclave at 121 degrees centigrade, we certainly did not completely
inactivate.
Now,
we did get down to almost a limiting dilution.
In other words, we have reduced infectivity probably in the region of
10,000 fold or something like that, but within a medical care context to have
surviving infectivity at this level would be a concern. And so I present this simply to discuss an
idea that hot SDS is a universal panacea.
Now,
Bob Rohwer discussed with you his hydroxide data that he co-published with Paul
Brown in 1986, I think, and he also showed a list of, if you like, some
contradictory data. These are
publications, which are all saying much of the same thing, and that is that
sodium hydroxide looks to be pretty effective, but not completely so. A suggestion is that you are knocking down
infectivity, because this is at room temperature, by the way.
And
the one comment I would make to Bob about certainly our experiments compared to
his, I can't talk for many of the others, but clearly, you found complete
inactivation, but we didn't know it.
It's acknowledged in your paper that the sensitivity of your assays were
slightly reduced because of the toxicity of the hydroxide to the examples. In other words, you diluted these to make
them so that they could be tolerated by the hamsters.
In
our own experiments, what we found is that if we fiddled around considerably,
we could actually neutralize, get the pH down to neutral in the end products,
and provided they were injected very quickly into the brains of mice, we didn't
need to dilute. So there is a slight
difference in sensitivity between the tests.
I'm not saying that is necessarily the explanation, but it is possibly
so, because we do have a solid bank of data saying cooled hydroxide is not
completely effective.
Right. In our own studies, what we found was that
after exposure and, again, room temperature, this is a hamster agent, one molar
hydroxide, two molars for two hours. We
brought the infectivity level something down, certainly, but we were left with
about 4 logs of infectivity.
Now,
if we combine the hydroxide treatment with heat as has been recommended, then,
in fact, we find complete inactivation either when you add hydroxide to the
samples and immediately autoclave or when you hold in hydroxide for an hour and
then go and autoclave. And in other
studies, such as those from the Rocky Mountain Lab, they found that if you held
in hydroxide, then neutralized the pH and went on to autoclave, you still got
inactivation.
And
these are the various publications, which all come to the same viewpoint,
somewhat unusual in TSE studies to have so many publications saying the same
thing, that hot hydroxide is effective, whether this is a sequential process or
whether the hydroxide treatment is at the same time as your autoclaving.
Now,
in terms of sodium hypochlorite, which is one of the recommended procedures, we
did some studies quite some time ago with two strains of mouse agent exposed to
sodium hypochlorite containing various concentrations of available chlorine,
and we found that once you got up to about 8,250 parts per million of available
chlorine, you had a complete effect.
Now,
the data here, and these were studies that were done on behalf of the
Department of Health in the UK some time ago, and being extremely conservative,
the Department of Health accepted the data, but said well, to play it safe,
we'll make the recommendation that you should use sodium hypochlorite
containing 20,000 parts per million of available chlorine, which considerably
exceeds the lowest levels of efficiency here, but that's where the
recommendation came from to use 20,000 parts per million.
Somewhat
later, using two sources of BSE infected cow brain, we tested sodium
hypochlorite once again. Alongside it,
we also tested sodium dichloroisocyanurate, which is another chlorine releasing
compound, which is generally considered to have a comparable efficiency
compared with hypochlorite when compared at the same levels of available
chlorine.
In
these studies of these various concentrations of available chlorine, there was
no infectivity detected in any of the BSE cow brain samples treated with
hypochlorite. But when you looked at
the samples treated with the dichloroisocyanurate at comparable levels of
available chlorine, there were, in fact, a significant number of positives.
This
came as rather a surprise, but we found then by doing assays on the chlorine
content left after the exposure periods, that the sodium hypochlorite compared
with the dichloroisocyanurate, if you look at the starting and finishing
concentrations of chlorine, the hypochlorite much more readily gave up its
available chlorine during these decontamination procedures compared with this
compound. It may be that longer
exposures might be effective, but we are already up to two hours, which it's
getting a bit impractical to extend things beyond that.
Mention
was made of boiling. Well, we certainly
do have data, which have only actually ever appeared in an abstract sort of
meeting. They have never been formally
published, but we did find with 301V, that if you boiled for one minute, that
we have no detectable infectivity left compared with material exposed to
hydroxide at room temperature or microwaved for one minute.
Bob
mentioned the data produced based on 134 to 138 degrees centigrade porous load
autoclaving. This was in either BSE
infected cow brain, scrapie infected sheep brain or scrapie infected hamster
brain, and we had survival rates as shown here, which, as Bob suggested from
this graph, fall pretty far down on his survival curve. And, indeed, when we titrated, the starting
titre here again was 9 and a half logs.
It came down to about 2 logs or less.
So substantial inactivation, but, in fact, in terms of health care,
still a worrying amount of infectivity left.
In
terms of more recent studies using 301V, we had really surprising data for this
experiment where we autoclaved either at 134 or 138 degrees centigrade for
these periods of time with these weights of tissue. Now, the norm is, of course, as you increase autoclaving time
and/or temperature, you expect the efficiency of decontamination to increase.
In
these studies, the reverse was true. In
fact, we had more cases of TSE in the case injected with the samples from the
138 compared with the 134 process, which was statistically significant. If done on a one off basis, I would have had
severe doubts about the technical quality of our experiments here, but, in
fact, we had other experiments running at the same time, which showed the same
trends, perhaps not so impressively as here, but definitely showed the same
trends.
Also,
there were studies being carried out on behalf of the Department of Health who
insisted quite correctly that all of the equipment and the processes should be
independently monitored. And so we had
a third party monitoring the progress of these experiments, thermocoupling of
blanks for every single stage of the process.
And there are some of you that know there is still, I think, a T-chest
full of trace-outs for all these experiments.
So I have no doubt that we're seeing a genuine trend here.
I
mentioned that we're using 301V and we do know, as Robert Somerville mentioned
this morning, that 301V within the spectrum of the agents that we have tested
is certainly far more thermostable than others. These strains here are all most precise scrapie agents. 301V is our most precise BSE agent. There is a survival after autoclaving and
the blue bars are the untreated samples.
And you can compare the titre losses with the different agents after the
autoclaving process.
My
take on what was happening in these experiments was that in the past where we
found much more efficient inactivation or in some cases, complete inactivation,
we often used intact pieces of brain tissue.
In the more recent experiments, as Bob Rohwer said, we were using brain
macerate. This is undiluted brain,
which is just mixed up, so it's a homogenous sample to give you a blancmange
like material for autoclaving.
Now,
in putting these samples into what Ron described as long neck tubes, not
terribly long neck, but there is almost inevitably some smearing and drying of
the infectivity onto the tubes before you get to the autoclaving stage.
My
concept, my take of what is happening and what explains the results is that
during the porous load autoclaving process, which I must tell you, unlike the
gravity displacement system where there is usually a slow buildup of steam, the
porous load system involves a huge and rapid admission of steam into the
chambered autoclave, which, in my simple hypothetical structure here, is able
to fix any proteinaceous material in these fringes, and that paradoxically, if
that protein is PrP protein, the actual fixation process, which occurs early
and rapidly at the beginning of the steam process actually protects that
infectivity from the subsequent sterilization of the steam effect.
If
that was so, that would explain why the 138 degree samples were more positive
than the 134 since you would expect the rapidity and efficiency of that heat
fixation to be greater at 138 compared to 134.
I hope to show you in the next few slides that this is not all quite
cuckoo land.
We
do know that if you fix infectivity or fix infected tissues with formaldehyde,
you make that infectivity colossally more resistant to inactivation by
autoclaving. Here, we have 50 milligram
fragments or whole mouse brains that are infected with the strain called 22A,
fixes in formalin and then autoclaved.
And, in fact, 100 percent of the recipient animals have gone down in
disease. Whereas, in these experiments,
we were completely able to inactivate infectivity in these samples if they were
simply emerged in saline.
Similarly,
if you immerse infected mouse brains in ethanol, another protein fixative, and
then autoclave, you get remarkable survival of infectivity even though ethanol
fixed in autoclave, 100 percent recipient animals going down. So there is clear evidence that if you fix
the PrP protein by whatever means, you, in fact, stabilize it to the extent
that it is not normally taken out by the standard autoclaving procedures that
we're looking at.
And
to test the hypothesis a bit further, we picked up on the experiments of David
Asher's going back, I think, to the 1980s.
I think he was among the first to observe that with scrapie-like agents,
if the materials are dried onto surfaces, they become extremely difficult to
inactivate.
Here,
we have an infected brain homogenate simply autoclaved and then injected into
mice, and in this case, one of the eight animals went down. If on the other hand, we took the
homogenate, dried it onto a slide, autoclaved it and then reconstituted it,
scrape the top of the infectivity again and try to challenge animals, 100
percent of these recipient animals went down.
That,
again, would be compatible with the idea that this thin sheet of material on a
microscope slide in autoclave would be subject to very rapid and efficient heat
fixation. And there is one more
experiment that we carried out with this in mind where we knew that dry heat at
160 for an hour would not inactivate the agent, but that autoclaving without
any other processing was effective. We
dry heated, which would heat fix, and then autoclaved and, again, we had
substantially more survival of infectivity.
So
my interpretation is that the effects that we're seeing of smearing and drying
of tissue in tubes in autoclaving experiments may well be down to heat
fixation. I make no apology for the
fact that many of the experiments that I have done have used brain macerate and
not brain homogenate, that these have all been more scarce conditions, because
many have been driven by public health concerns funded by the Department of
Health who do actually want to know what happens under worst case circumstances
that could reflect conditions relating to tissues dried on instruments,
etcetera.
In
terms of concern over instruments, the Department of Health has funded quite a
number of studies relating to decontamination, disinfection. They are quite interested in the combined
hydroxide and heating effect, and one of the concerns is what effect does this
have on stainless steel instruments and devices?
So
one of the studies being carried out in Edinburgh is to look at test pieces
made of stainless steel before and after various hydroxide treatments. It is mainly facilitated by collaboration
with the engineering department who have a scanning white light interferometer
where you can compare the roughness indexes of surfaces before and after
various treatments. It will print out
different graphs giving you the roughness indexes.
And
here, just to the naked eye are test pieces, which on the left hand side are
all untreated. These are different
grades of stainless steel. On the right
hand side are pieces that have been subjected to autoclaving at 121 centigrade
for 24 hours. And as you can see, in
some cases, there is hardly any difference, but in some cases, there is a
darkening of the testing piece.
As
I understand it, this is due to precipitation of chromium salts and what I'm
unaware of is whether you can clean these chromium salts off and start again
with a pristine surface. This is what
was started before I left the unit, so I'm not quite sure of the current state
of play.
I
will finish off with just three slides containing anecdotal information, which
may be of some interest. One is that a
low formalin fixed tissue is incredibly difficult to inactivate, and one would
recourse usually to incineration for its disposal. We did find that the hot hydroxide process, when applied to
infected brain tissue fixed in formalin was, in fact, effective at removing
that.
And
I will finish up with two more overheads, if I may. One goes back to the GME study, and the figures may have changed
slightly, but the principles are nevertheless the same. It was discussed how the superimposing of
our sodium hydroxide step, especially to the ossein material that remains after
the acidic extraction process where there was a significant amount of
infectivity surviving, at that point.
If you then applied -- I'm sorry, this should be hydrochloric acid up
here. If you then apply .3 molar sodium
hydroxide for two hours at ambient temperature, there was no infectivity
detectable in the resulting gelatin.
This
clearly suggests that earlier studies using infected brain suggested that one
more sodium hydroxide is quite effective and not completely so, these studies
suggest that when you get the circumstances in an environment such as ossein
where you are largely devoid of any extraneous lipids or proteins, that the
hydroxide process is much more effective.
And
I will leave you with some recent data from a commercial study, which I have
some sketch information for you from.
This involves a process where raw materials exposed to saturated lime
calcium hydroxide, and then it goes on to hot lime at 80 degrees centigrade,
here we are, sorry, and thereafter onto even hotter lime at greater than 140
degrees centigrade under pressure, much of the same conditions if not higher conditions
than those described that are completely effective.
Now,
the thing to be on your mind here is that the pH of the lime, the maximum pH of
lime is significantly lower than that of one molar sodium hydroxide. What we seem to be finding here is that
after the exposure to saturated lime for three hours at 80, we do have some
titre loss. The expectation then might
be that when you go on to this very high pressure, high temperature process,
that you might lose all the infectivity, but, in fact, you do not.
To
me, this demonstrates potentially two things.
One is that molarities of hydroxide lower than one molar may not be
truly effective under the high pressure conditions and/or separately than any
surviving infectivity from this stage, which is carried out at 80 degrees
centigrade, the heat fixation, which goes on in here, all the surviving
infectivity may, in fact, render it more resistant to inactivation at this
level. So these are speculative
comments, but they all contribute to the general arguments about heat and
hydroxide. And I will leave it
there. Thank you.
CHAIR
PRIOLA: Are there any questions? Oh, please, Dr. Edmiston.
DR.
EDMISTON: I have a comment, which I
want to direct to the speakers, the previous speaker and Dr. Taylor, and also a
general comment to the members of the panel in terms of how this applies to
surgical instruments in the operating room.
I
am not surprised that you haven't achieved complete inactivation, because as a
rule, it's a general trend we sort of adhere to, is as long as there is
biological material present or, I should say, as long as the organic component
is still there, it's unlikely you're going to see complete inactivation.
From
a surgical perspective, one needs to recognize, and I'm not quite clear on what
my colleagues are doing in Europe, but at least from the U.S. perspective, we
just don't take surgical instruments and put them into an autoclave. There is a pretreatment facility, which
reduces organic comment, and I know the next speakers will address that
probably in some detail.
Actually,
I am heartened by some of the data you have shown in terms of inactivation,
especially in the presence of high organic content. The fact that in these high carbon environments, you are able to
reduce the number of viable particles, so I think we need to think about this
two step process as we procedure through the next day and a half in that we're
just not talking about instruments being directly sterilized. We're talking about a process in which
instruments are being rendered sterile by virtue of not only a sterilization
process itself, but also the removal of organic material prior to
sterilization.
DR.
TAYLOR: Yes. Could I make one comment now?
There is, at least in the UK, what I consider to be a worrying trend,
and that is that traditionally in a very common sense fashion, it was common in
wards and even theaters for certain instruments to be washed in the sink before
they went on for washing in the Central Sterilization Department.
That
process is increasingly being discouraged for health and safety reasons. It's resulting in an increasing number of
instruments reaching the Central Sterilization Department with absolutely dried
on blood, tissue and whatever, and this is the problem that, I think, you are
referring to. The washing processes as
they exist at the moment, at least in the UK, are largely incapable of dealing
with the situation where you have material that is absolutely dried or baked
on.
DR.
EDMISTON: And I think the recommendations
and this Committee needs to anticipate the fact that that is a problem. Therefore, the recommendations not only from
this Committee, but from other professional organizations such as APEC and
others would suggest that pretreatment of these instruments is mandatory.
CHAIR
PRIOLA: Yes, Bob?
DR.
ROHWER: Yes. I am aware that that's how it's done in the hospital
setting. I work in a hospital, but the
problem that we have with that is the potential for cross-contamination at the
level of the cleaning, and especially in a laboratory environment at least,
that would be a disaster for us to spread this stuff around in our sinks and
cleaning stations before it ever got to the autoclave.
So
we want to make absolutely sure that we know that our instruments are
contaminated. You don't necessarily
know that yours are, and so we want to make absolutely sure that everything is
gone before we even handle them, and we do that by going to these extreme
measures.
On
the other hand, the only thing that I find encouraging about what you do is
what standard practice is in the hospital, is that slide that Dr. Asher showed
in his introduction. There really isn't
any evidence that instruments cleaned and sterilized in the way that is
specified by current practice are causing CJD infections, and I think we have
to give a lot of weight to that.
On
the other hand, I think it's also very important to think about the cleaning
step and what kind of potential that poses for having a major accident if you
don't contain that particular environment, as well, because I consider that a
high-risk environment.
DR.
EDMISTON: Well, you need to know that
most of us have had a high threshold interest in this for several years, and
more and more of neurosurgical instruments are being triaged and actually being
treated separately in separate kits.
And for the most part, and I will say for the most part, because there
are exceptions, are not getting into the main surgical instrument stream.
And
we're spending a lot of time and effort with our neurosurgical colleagues to
first of all identify potential patients or suspected patients, but overall, I
can tell you most surgical departments, most hospitals, will have unique
surgical, neurosurgical kits, and this is becoming more and more common for the
reason that you just mentioned.
DR.
ROHWER: I guess the thing is I would
like to know more about how that segregation takes place, because it's not
particularly comforting to me to know that the neurosurgical instruments are
being segregated. Neurosurgery is
potentially the biggest hazard in terms of passaging the disease, so you run
through a set of CJD exposed instruments, and then that is followed by a set of
cleaned instruments, you know, coming from a normal patient or something like
that. How do you assure yourself that
you're not getting cross-contamination at the level of neurosurgical
instruments in that type of environment?
DR.
EDMISTON: I won't go into a lot of
detail on this, because I know my colleague over here will discuss it, but when
patients are identified, those instruments are quarantined and sequestered, so
that they are treated entirely separate from the rest of the general
instruments. So that is the policy that
most of us have developed over the years in dealing with these suspected
patients.
Now,
the other issue is well, how about all of the other neurosurgical patients,
which you find out about anecdotally?
Now, that is an important process to discuss, but in terms of those that
we identify or we suspect, those instruments are quarantined and they are
triaged and segregated out of the system.
DR.
ROHWER: Okay. Can I say anything more?
Are you tired of hearing me? I
guess my rebuttal to that would be that my guess is that the greatest part of
the risk comes from people that you will never, ever identify as even carrying
the disease, and that's the greatest part of your exposure. You will never know about it, and what I see
a need for is some way to actually effectively sterilize the cleaning
environment between uses.
CHAIR
PRIOLA: Are there other questions for
the speakers? I have one quick one for
Dr. Taylor about the experiment you showed where you exposed material to dry
heat at 160 degrees, and you had complete transmission. And then you took the material, exposed it
to dry heat and then, if I remember, you autoclaved following the dry heat, and
that dropped to almost 50 percent survival or you get 50 percent survivors.
What
implications do you think that has or does it have any implications for
multiple rounds of autoclaving, say wet autoclaving or multiple rounds of
sterilization and getting rid of that residual activity?
DR.
TAYLOR: It's difficult to answer your
question within the context of infectivity dried onto surfaces. All I can say is that I have done one
experiment where I didn't make any attempt to smear and dry, but just using
standard samples with the hamster agent where after one round of sterilization,
and I am quoting figures very crudely here.
In
the first round of a standard autoclaving procedure, I lost something like 4
logs, somewhere about there. And when
that material was taken and just reautoclaved, the loss on the second round was
about 1.7 logs. So the second
autoclaving, even under these conditions, was certainly not very efficient, and
I suspect would have been even poorer if this had been agent that partially
survived after smearing and drying.
CHAIR
PRIOLA: Dr. Gambetti?
BOARD
MEMBER GAMBETTI: Listening to all these
presentations, of course, are very informative. One, however, wish that experiments were available in which
decontamination of surgical instruments is monitored under more realistic
conditions. For example, one wished
that there would be some data on decontamination of surgical instruments used
experimentally in a more surgical operation on a CJD brain, and then see how
this level of contamination that is a classic level of contamination that you
may expect from a CJD brain in surgery, how the decontamination is effective on
those particular conditions.
Vice
versa, one would like to know how much decontamination is achieved on
contaminated surgical instruments after the routine sterilization that the
surgical instruments undergo, as I said, under routine conditions. Those are the data that I would like to know
whether they are available at all. I
have never seen, so I think those would be very useful data to have for this
discussion.
CHAIR
PRIOLA: Dr. Taylor, do you have a
response to that?
DR.
TAYLOR: Yes, I have an experiment that
I started before I retired, and I'm going to throw the buck right over to
Robert Somerville here as my successor.
In this experiment, the very question that you're asking was asked. In other words, how realistic or how
appropriate are the inactivation we're achieving to real life situations?
Now,
we weren't doing neurosurgery on human patients, but we were daily doing
surgical interventions within the brains of infected animals. So we took deliberately infected instruments
that had been deliberately traumatized into animal brain, subjected them to
routine washing procedures, and then proceeded to reuse these instruments again
neurosurgically or in subsequent animals.
My
take on things before I left, and I haven't looked at the data since, was that
even the washing processes in the lab, which were not anything up to the
Central Sterilization Department were having a useful, if not complete effect. But by the time we got to reuse of these
instruments on animals, they weren't, as measured at that time, producing any
significant levels of infections in the animals. I don't know if Robert can add anything to these data or are they
still lying buried?
DR.
SOMERVILLE: I think they are still
lying buried, David. I don't have
access to the data at present. What I
would say to Professor Gambetti though is that attempts, which I think
Professor Weissmann is addressing the Committee about tomorrow, I think model
the kind of situation that you are trying to -- the kind of question that
you're asking, and that is the implantation of contaminated surgical
instruments, stainless steel. Professor
Weissmann has already done some studies with contaminated wares and our lab is
also hoping to initiate this kind of system with different grades of stainless
steel.
One
of the problems that you have to appreciate is that surgical instruments made
out of various kinds of stainless steel, and that is one of the challenges of
actually set up these kinds of experiments, is how you model the different
kinds of surfaces that will be involved in real life. But to summarize, I think the best way of testing your question
is through this kind of model.
CHAIR
PRIOLA: Okay. I think we'll move on to the next speaker who, as Dr. Asher
mentioned, was supposed to be Dr. Philippa Edwards, but she has taken ill and
is unable to attend, so Dr. Pedro Piccardo from the FDA has graciously agreed
at the very last minute to give her presentation. Dr. Piccardo?
DR.
PICCARDO: Thank you. Well, obviously, Dr. Edwards could not
attend, and yesterday I was given somehow the daunting task of presenting the
information that she provided. I will
try to do this as objectively as I can.
Next
one. Okay. It has been settled already iatrogenic transmission of
Transmissible Spongiform Encephalopathies from person to person has occurred in
non-Variant CJD, and this has instated already, and here are the numbers that
were provided by a publication called Brown and Neurology in the Year 2000. And as you see, the bulk goes to growth
hormone treatment and dura mater grafting.
However,
there are here five cases implicating which neurosurgery, meaning contaminated
instruments, have been implicated. On
top of that, we have a few cases following treatment with gonadotrophin,
chromium transplants and, of course, electrodes here.
The
next, please. However, one of the big
problems came when in 1986, Bovine Spongiform Encephalopathy was described in
the UK, and the problem became humongous when in 1996, vCJD was described in
humans. As you see here, I mean,
obviously, there are a numbers of barriers that have been established to try to
prevent the transmission of vCJD, the agent, from animals to humans. However, the big question here is humans are
being exposed. Humans died with vCJD,
and the question is we don't know how many people has been exposed, how many
people could be infected.
The
next one, please. And, of course, we
don't know how many people may be asymptomatic, at this time and carriers.
The
next one, please, the next one, the next one.
Okay. The next one, please. The next one, please, the next one, the next
one. So due to great uncertainties,
risk assessment has been considered.
What happened? Oh, okay. Here we go.
Due to great uncertainties, risk assessment has been considered. The risk assessment has considered a wide
range of scenarios.
And
why the risk assessment was done?
Basically, for two reasons. One
was to determine the risk of transmission of vCJD through surgical instruments,
and the second one to indicate what measures could be the most effective to
reduce the risk.
The
next one, please, the next one, next one, next one, next one. The guidance follows the assumption that an
average of 10 milligrams of material could remain in instruments, and this
information I gather from the document that was provided by the CJD Incidence
Panel.
Next
one, please. Go ahead again,
again. The risk could be calculated for
different scenarios, and the effect of different actions could be estimated.
Next,
please. Next, please. Okay.
Improving the standards of decontamination is one of the main objectives
of the UK policy, and single-use instruments have been considered, for example
for extraction of CSF, and the idea was to use as much as possible single-use
instruments without compromising the clinical standards, of course. And a pilot program was established to use
single-use instruments for tonsillectomies.
Next
one, please. Go ahead. Why tonsillectomies? Why was this chosen? Because infectivity is present in vCJD in tonsils. I mean, PrP has been found in tonsils from
patients with vCJD. The other thing was
the relatively large number of operations and the other thing is the young
patients usually with long life expectancy go through this type of surgery, and
these are instruments that can be identified.
Next
one, please. Okay. But there were some adverse reactions. I mean, why there were problems? One was you cannot probably think that the
problems raised from the quality of the sets, the surgeon preferences and the
other problems were unrelated to the use of single-use instruments. So at this time, there is an audit on this
situation.
Next
one, please. So what Dr. Edwards tried
to convey, the message that she tried to convey with this cartoon is that while
trying to solve one problem, you create another.
The
next one, please, again. The best
decontamination available cannot be guaranteed to remove all sorts of
infectivity, and single-use instruments definitely is a situation that is not
possible for all kinds of surgery, so we must bear that in mind.
Next
one, please. Okay. Here, we have on this panel, a tissue
forceps, the tip of a forceps that has been routinely decontaminated. Here, we have electromicroscopy, and this
what we can see here in green is material that remains on the tip of that
forceps. This is the kind of material
that remains. This is florescent
staining for protein, and this is the superimposition of these two images gave
this image. So, obviously, there is a
significant amount of material that remains, a lot of which is protein
following routine decontamination.
The
next one, please. So to reduce the risk
of transmission of TSE from person to person, the Department of Health seek
guidance from the Advisory Committee.
Next,
please. Next, please. And a first version was done in 1998 and
now, there is a revised version, June 2003.
Next,
please. This presentation concentrates
mainly on the risk arising from the care of patients.
Next,
please. Yes, okay. So well, one of the issues is dealing with
symptomatic patients. I mean, when
dealing with patients with CJD, there are three types of definition. One is a definite case. By definite we mean something that has been
clinical and pathologically confirmed.
Probable case, which is has clinical, but on top of that usually, there
is electron encephalographic analysis and there is MRI imaging analysis, and
possible CJD when usually is by clinical presentation.
Next,
please. Now, when we are dealing with
asymptomatic patients, when we talk about risk in the case of -- when we talk
about asymptomatic patients and we talk about risk, we have to consider two
situations. One is in the case of
inherited diseases, and by inherited diseases, we consider that there are two
or more blood relatives are affected by a prion disease or one or more blood
relative showed genetic testing, show a mutation in the prion protein
gene. Usually what is done is PCR sequence,
the open reading frame of the protein, and then from there you can detect mutations.
Next,
please. Now, the other is the
iatrogenic risk, and this case already was mentioned treatment with hormones,
dura mater grafts, and that is why, I mean, obviously, the Department of Health
seeks advice.
Next,
please. This table is based on what we
currently understand about the distribution of infectivity in sporadic CJD or
in non-Variant CJD, and obviously, when we talk about tissue infectivity as it
has been said already many times, the highest amount of infectivity is here in
the CNS or retina and low medium type of infectivity in the eye and olfactory
epithelium.
Next,
please. Now, when we talk about risk of
different tissues in Variant CJD, the situation varies, because we have to
introduce into the medium risk tissue lymphoid tissues. The rest remains the same.
Next,
please. So we don't have a problem here
and we don't have a problem here, because this is by genetic testing or what
was said already, is we can know who these people are, and we understand who
these people are. But the problem is
when we deal with sporadic or when we deal with variant, people that are
asymptomatic, at this time, but might have infectivity.
The
next, please. Yes, okay. So the problem comes or starts when,
obviously, a CJD patient is diagnosed, and immediately the question should be
has that patient had surgery or donated blood, etcetera, and then try to assess
what is the risk to other patients that have been exposed to instruments that
have been used on this patient.
Next,
please. And the risk, basically, will
depend on the type of tissue that we are talking about, because we said already
that there are tissues with high levels of infectivity and in this case of
vCJD, the lymphoid tissue corresponds with tissues with medium levels or medium
risk.
Next,
please. So this graph is an estimate
that comes from animal studies, so this is an estimate that comes from animal
studies. And, obviously, the paren of
tissue infectivity in vCJD probably could follow this, and this is the onset of
clinical symptoms. So if we go, let's
say that the surgery was done way before the development of clinical symptoms,
probably the amount of infectivity will be very low, and because we are dealing
with vCJD in this graph, we have two parameters or two tissues to
consider. One is the CNS and the other
is lymphoid tissues.
Next,
please. So, as I said already, the risk
depends basically on the type of -- I mean, depends once again on the type of
tissue where the surgery is performed, and if there is variable time between
surgery and onset of disease. Well,
this basically refers to the previous graph.
Next,
please. Okay. One of the issues here is that the risk depends -- let me
see. In the document provided by the
CJD Incidence Panel, it is stated that the first washing and autoclaving would
achieve at least a 105.4 reduction of infectivity, and this was
already mentioned by Dr. Taylor before, and that subsequent cycles of
decontamination reduce the infectivity, but it's much less effective.
The
next one, please. So at this time, the
Department of Health is in discussion with manufacturers of surgical
instruments. I mean, the discussion is
based on what is the probability of using single-use instruments or to replace
parts or to provide instruments that could be easily decontaminated.
Next,
please. So what are the aims of the CJD
Incidence Panel? Well, it seems to be
quite obvious, which is to protect the patients. Let me see.
Next,
please. The aims. Go ahead again, again. Next, please. No, yes, yes, here we go.
Obviously, the aims are to protect the patients and to inform
potentially exposed patients, and to inform the public and to increase the
knowledge.
Next,
please. So in management of risk, I
mean, what is being done is quarantine the instruments during the risk
assessment, and instruments that have undergone less than 10 cycles of
decontamination should be incinerated.
Next,
please. Again, next, please. Okay.
So the patients will be contacted to alert them of their possible
exposure and to take health protective actions. So these are the patients that should be contacted under these
circumstances. If the index patient
goes through, I mean, the material went through high-risk procedures, the first
six patients that follow that first surgery should be contacted. This is for tissues with less amounts of
infectivity, and these are the amount of patients that should be contacted.
Next,
please. So this is sort of the kind of
data that has been gathered during the last two years of experience, and this
is the incidence reported to August of 2002, and definitely we have 39 cases
implicated in Variant CJD, 39 cases implicated in sporadic CJD, and that there
are few that correspond to familial or non CJD or unclear, cases that could not
be determined.
The
next one, please. The type of surgery
is 131. So obviously, before were 87, I
believe. Yes, 87, and now, we are
talking about 131 surgeries, and the reason is that some patients went through
more than one surgical procedure. And
what we see here is that the GI surgery takes the bulk followed by obstetric
and gynecology and here we have neurology, neurosurgery.
Next
one, please. So in 76 incidents,
tracing was sought. Some or all were
traceable, that means in 34. In 18, it
was not possible to trace them. And in
24, there is incomplete information, at this time.
The
next one, please. So instruments that
have been quarantined are 48 and that have been not quarantined are 39.
The
next one, please. The fate of the
quarantined instruments, we have 21 that have been returned to use, because it
was assessed that the risk was not higher than the usual risk for the UK
population. Here, we have four that
have been completely destroyed, the whole panel was destroyed, because it was
not possible to identify exactly which of the instruments was involved. And in 23 cases, the hospital directly
decided to take care of the instruments and destroy them.
The
next one, please. So, obviously, this
is a very difficult task and there are a number of dilemmas and difficulties,
and there are a number of scientific uncertainties, and it is very difficult to
trace back instruments and sometimes patients and, of course, there are ethical
issues that are involved.
The
next one, please. Go ahead, yes. And these are the websites that Dr. Edwards
suggested consulting for further information.
I think this is it. Thank you.
CHAIR
PRIOLA: Dr. Khabbaz?
DR.
PICCARDO: Oh, before -- excuse me,
sorry. Yesterday, after I learned that
I had to give this presentation, I called the UK immediately, right away, and
the first question I asked before you ask me the question was what do I do with
-- how do I handle the questions?
So
the thing is we will make clear note of your questions, and then we will
forward the questions to the UK, and Dr. Edwards has been kind enough to review
them and, hopefully, if she feels well enough, to provide the answer tomorrow. So we are in business. Anyhow, if you want to ask the question, go
ahead.
CHAIR
PRIOLA: So that gets you off the hook,
doesn't it, Pedro? Yes.
BOARD
MEMBER KHABBAZ: You may know the
answer. I had actually a couple of
questions. One has to do with the
adverse events related to single-use of instruments for tonsillectomy. Do you have any idea what types of adverse
events would occur?
DR.
PICCARDO: Yes, the answer is bleeding,
bleeding. That is the answer to that.
BOARD
MEMBER KHABBAZ: Thanks. The second question is I don't think -- I
may not have understood you correctly.
I think when you talked about the various types of CJD, you mentioned
for inherited and iatrogenic not concerned for infectivity?
DR.
PICCARDO: Sorry, come again. I mentioned --
BOARD
MEMBER KHABBAZ: No infectivity for
inherited and iatrogenic versus sporadic and Variant CJD. Is that in terms of how they got it or for
peripheral tissues?
DR.
PICCARDO: Let me see. Can you pose the question again? I have a problem hearing, too.
BOARD
MEMBER KHABBAZ: Okay.
DR.
PICCARDO: Listening to the
question. Yes, go ahead.
CHAIR
PRIOLA: Yes. I think you're referring to the slide you showed where you had
sporadic patients, iatrogenic and inheritable.
And are you asking if there was infectivity associated with those
patients?
BOARD
MEMBER KHABBAZ: I didn't understand the
statement that there is no infectivity related to inherited and iatrogenic.
CHAIR
PRIOLA: Oh, I don't think that's what
you said.
DR.
PICCARDO: No, well, I would be happy to
review Dr. Edwards' slide. However, my
answer to that is that there is no difference between inherited and
sporadic. We probably put them in the
same box. We will do a difference when
we deal with Variant CJD, because that is when we have tonsils and we have the
lymphoreticular system involved that we tend not to have in sporadic or other
forms of CJD.
CHAIR
PRIOLA: Right, and I think that was one
to identify, patients at risk, right, prior to use of instruments. If someone has inheritable mutation, then
that is a patient that you identify as being at risk of possibly transmitting
to somebody else. I think that's what
that --
DR.
PICCARDO: Right, in terms of risk. I mean, if you have a patient that,
obviously, comes from a family and has the mutation, etcetera, etcetera, you
know that patient is at risk already, so it's very easy to recognize that
patient. It's also very easy to
recognize a patient that, let me see, that went through surgery that has a dura
mater graft. It could be a patient at
risk. However, if you say well, let's
take sporadic CJD, maybe I am incubating sporadic CJD and I don't know and no
one will know.
CHAIR
PRIOLA: Dr. Nelson?
DR.
NELSON: How would you classify
cerebrospinal fluid that, let's say, has lymphocytes or is an inflammatory
cerebrospinal fluid? Would that be the
same as blood being low-risk or would it be closer to CNS tissue?
DR.
PICCARDO: Well, I would like someone
else, probably Dr. Asher, to attend that.
Before we go ahead with that, Dr. Edwards made clear that they provide
disposable instruments for CSF extractions, so now, it's single-use. Go ahead.
DR.
ASHER: Yes. In the NIH series, about 15 percent of spinal fluids from
subjects with mostly sporadic CJD did transmit disease to primates. So the risk of spinal fluid is comparable to
the risk of some non CNS solid tissues, lymphoid tissue, liver, kidney, spleen,
lung.
DR.
NELSON: But it's definitely higher than
blood?
DR.
ASHER: Higher, definitely higher than
blood.
CHAIR
PRIOLA: Yes, Dr. Gambetti?
BOARD
MEMBER GAMBETTI: I believe that the
experiment that, David, you are quoting included not only sporadic, but also
Kuru patients, and three of 37 or so, spinal fluid tested transmitted the
disease. Do you know whether some of
the CSF that transmitted the disease were actually from Kuru patients, rather
than sporadic case? Do you know that?
DR.
ASHER: I think it's in the '93 article,
but I don't remember.
CHAIR
PRIOLA: Okay. I think we'll move on.
Thank you, Dr. Piccardo. We'll
move on to our last speaker before the break if there are no other questions,
and that is Dr. Bill Rutala who is going to discuss TSE agents and infection
control in U.S. hospitals.
DR.
RUTALA: Thank you very much and good
afternoon. What I would like to do very
quickly, and certainly by looking at the next slide, review the recommendations
on and the practices in U.S hospitals as it pertains to the prevention of cross
transmission from medical devices contaminated with prions and, hopefully, have
a few minutes to discuss how important methodology is, and how important
methodology is from the standpoint that we can fail to inactivate even easy to
kill microorganisms like bacteria with FDA cleared sterilization processes
dependent upon the methodology that is employed to include the absence of
cleaning.
Next
slide. Let's begin with the rationale
for the U.S. recommendations, and these recommendations have existed for
decades, the recommendations in infection control literature, surgical
literature, certainly, essential processing literature and so forth.
But
let's look at the next slide. As we
know as far as the epidemiology of prion transmission, we know that it's not
spread by contact. It is not spread by
airborne. It is not spread by
environment, but we are concerned about the iatrogenic spread.
Next
slide. We can see here that
contaminated medical instruments have been implicated in disease transmission,
and we'll discuss that in just a minute.
Next
slide. Well, let's look at this issue
of prion transmission via surgical instruments.
Looking
at the next slide, we see, essentially, the two confirmed cases that have
already been mentioned. Those two
confirmed cases, of course, were reprocessed by a method that we never use in
U.S. hospitals, a combination of benzine, alcohol and formaldehyde vapor, and
then we also have four suspected cases.
Those
four suspected cases are involved with CJD that has occurred in persons
following brain surgery. However, only
one of the four had an index CJD case identified. These cases occurred before 1980 and there has been no known
failure of steam sterilization to date.
Next
slide. How about the infectivity of
human tissue as we discuss this rationale?
As we already know by looking at the next several slides, we used
epidemiology data and, of course, we used experimental data and infectivity
data. We know that there is evidence of
transmission via eye and brain from an epidemiology standpoint, and we know
that experimentally we can inoculate animals, susceptible animals, and
demonstrate that certain body fluids and tissues transmit CJD.
And
we have already discussed the contents of the next slide, which is that there
are certain tissues that are considered high-risk, certain tissues are
considered low-risk and, of course, some tissues that are considered no risk.
Next,
we see the issue of removing microbes by cleaning, something that, certainly,
we need to discuss a little bit more as it pertains to the methodology and how
methodology is so important in this issue of prion inactivation.
In
the next slide, we will see, essentially, something that has already been
mentioned by one of the panelists. The
issue that effectiveness should not consider only the effectiveness of the
disinfection of sterilization procedure, but also has to consider the
effectiveness of removal by cleaning.
And, of course, the probability of a device remaining capable of
transmitting disease is related to not only the initial concentration of that
prion on the surgical instrument, but also it is related to the effectiveness
not only of disinfection and sterilization, but also cleaning.
And
there are literally dozens of studies in the literature, which show how
effective cleaning is. Cleaning will
reduce anywhere from 4 to 6 logs of microorganisms by a manual or a mechanical
cleaning procedure. We don't have as
much data regarding protein reduction, but there are a few papers in the
literature that demonstrates there is, approximately, a 2 log reduction of
protein by the various cleaning procedures.
In
the next slide, we see the prion inactivation studies. We don't need to go over this very
much. We're just going to very quickly
go through a few slides. We could possibly
put a question mark up here with prions.
The question mark I would put up there is related to the fact that maybe
the studies that have been done are artifactual in nature and, essentially, a
reflection of the methodologies that are employed, and I think I can show you
data that would be supportive of that.
And
then also, we would see here that other microorganisms fall below possibly
prions and spores and it pertains to the susceptibility to disinfection and
sterilization procedures. And in just a
minute, I'm going to show you some data where bacteria will survive FDA cleared
sterilization processes, because precleaning did not precede the sterilization
process.
Next
slide. We know, of course, there are
many procedures that are ineffective or partially effective.
Next
slide. We know also that there are some
gaseous sterilization procedures and, of course, physical procedures that are
also ineffective or partially effective processes.
Next
slide. We can see, of course, that
there are some effective disinfectants and, of course, by effective here, we're
saying a 4 log reduction decrease in the ID50 within one hour and,
certainly, among them include sodium hydroxide and sodium hypochlorite.
The
next slide will just tell us what the effective processes are as it pertains to
sterilization, and this is what we use in U.S. hospitals. We, of course, use sterilization primarily
by steam sterilization with a prevacuum sterilizer at 134 for 18 minutes. Sometimes, the combination of sodium
hydroxide and steam sterilization is employed, but it's not widely employed
because of some of the deleterious issues associated with the combination of
sodium hydroxide and steam, deleterious, of course, to the instruments,
deleterious to the sterilizer and, of course, the vaporization of sodium
hydroxide to staff. But we certainly
recognize the effectiveness, and that is an option for hospitals to choose.
Next
slide. As it pertains to risk
associated with instruments, let's see what we have with that.
Next
slide. I just wanted to mention,
essentially, that there are certain categories of instruments in every health
care institution, not only in the United States, but in the world. There are certain instruments that we
consider must be sterile. They are
instruments that have contact with sterile tissue or the vascular system. We consider, of course, then to be very critical.
There
are other instruments like in endoscopes that have contact with mucous
membranes or skin that is not intact, and we have a very high level of
disinfection associated with those instruments. And the other instruments are noncritical, only have contact with
intact skin and, essentially, are not involved in disease transmission.
The
reason for mentioning that is seen in the next slide and that is, essentially,
in a minute we're going to develop, essentially, the scheme for how we
disinfect and sterilize instruments in health care setting in the United
States.
As
it pertains to surgical instruments, a question was just asked. What is the microbial load associated with
surgical instruments? Actually, we do
know the microbial load associated with surgical instruments. A few studies have actually evaluated the
microbial load. Of course, it's not for
prions. It's for other microorganisms,
and microbial load 80 percent of the time is less than 100 organisms. Rarely does it exceed 1,000 organisms. Many surgeries, many different investigators
have made that observation.
Next
slide. Well, this is how we decide how
to, essentially, employ special prion precautions in U.S. hospitals. We, essentially, assess the patient, assess
the tissue and assess the device. Of
course, we consider whether it's a high-risk patient, a high-risk tissue and a
high-risk medical device, again, those critical and semicritical devices.
Next
slide. As far as that is concerned,
most U.S. hospitals then would do special prion reprocessing, and that would be
those higher temperatures or a combination of sodium hydroxide with steam,
special prion reprocessing when it's a high-risk tissue, a high-risk patient
and a high-risk medical device, and for all other situations with one possible
exception, it would just be conventional disinfection and sterilization.
The
one possible exception would be a high-risk patient, a critical and
semicritical device and low-risk tissue.
Some hospitals do treat low-risk tissues from a high-risk patient,
critical and semicritical device as instruments requiring special prion
reprocessing, so possibly this would go into that category.
Next
slide. So the conclusions of this, of
course, is that from an epidemiology standpoint, we have two cases of disease
transmission that are definitive, possibly four other cases. The guidelines that we have discussed and
are used in the U.S. are based upon epidemiological evidence, tissue
infectivity, the risk of disease associated with certain medical devices and,
of course, inactivation data, and the risk assessment is based again on
patient, tissue and device. And only
when there is critical and semicritical devices, contacting high-risk tissue
and possible low-risk tissue from high-risk patients do we require,
essentially, special prion reprocessing.
Now,
the next slide, we see, essentially, what those special reprocessing procedures
are. We have already mentioned that
this is the preferred procedure that many hospitals use, the 134 at 18 and a
prevacuum sterilizer. There is no low
temperature sterilization technology that is recommended and we know,
essentially, that there are some disinfectants that have activity against CJD.
Next
slide. So this is what we're talking
about here. We're talking about used
instruments, and this is true for all used surgical instruments. They are kept wet. They are not allowed to dry.
They are, essentially, cleaned before they are sent to central
processing. We don't let tissue and
fluid dry on them. When they get to
central processing area, an area where all instruments are received for a
quality control standpoint, the instruments generally go into a mechanical
washer disinfector. In the case of
special prion reprocessing, there would be a special steam sterilization cycle,
and that instrument would be returned to health care.
Now,
we have already mentioned, essentially, the rationale. The last thing I want to do is very quickly
look at methodology and how methodology affects results. And I am going to show you some slides, and
let's begin with the next slide where, essentially, we can fail to kill easy to
kill microorganisms by methodological manipulations, and I will call them
manipulations, because all we do is we don't add cleaning to, essentially, the
process. And, of course, we have
already seen how important cleaning is.
Next
slide. This is the issue. I don't want to get too involved in this,
but, essentially, the point needs to be made that there are a number of studies
that have been done, and most of those studies are done in a worst case
scenario, of course, and, of course, we try to achieve sterilization by using
appropriate reprocessing procedures.
And
there are no studies, including the studies that have been published involving
cleaning, that reflect the reprocessing procedures in a clinical setting. We use enzymatic cleaners. We use mechanical sterilizers in a closed
system and we use mechanical washer disinfectors in a closed system.
Next
slide. Now, I want to talk briefly how
factors affected sterilization, and many factors affect sterilization, but I am
only going to choose a couple. First,
let's look at protein and salt.
Next
slide. If we just put, essentially,
some microorganisms on a penicylinder like this right here, next slide, and
then we let it dry for 30 minutes, next slide, and then we put those
penicylinders that are inoculated with easy to kill microorganisms, such as E.
coli and pseudomonas and enterococcus faecalis in a FDA cleared sterilization
process like ethylene oxide or hydrogen peroxide gas plasma, in the absence of
serum or salt, you get 100 percent kill.
In
the presence of serum or salt, you get 40 percent failure, as well as in this
case, 63 percent failure, a significant amount of failure, because cleaning did
not precede the sterilization procedure.
Now, the amount of salt and serum is not really that high. The amount of salt is .65 percent. The amount of serum is 10 percent, but
failure in the absence of any cleaning.
Next
slide. If we use, essentially, a lumen
device and do the same experiment, next slide, we see, essentially, 60 percent
failure. Again, we're failing to
sterilize instruments that are contaminated with easy to kill microorganisms,
because we failed to clean them. And
it's not really the lumen device that is causing the problem, because we see
here in the absence of serum or salt, those organisms are killed. So the failure to clean allows the survival
of easy to kill microorganisms in a sterilization process, such as low temperature
sterilization.
Next
slide. So all technologies have
limitations. Salt and serum provide
protections for spores and bacteria, and salt and serum with a lumen carrier
even provides extraordinary protection.
Next
slide. Now, let's look at the issue of
cleaning and let's look at spores.
Next
slide. Let's just put some spores on a
stainless steel scalpel and we'll see on the next slide. We're going to put about 106
geobacillus stearothermophilus spores on this stainless steel scalpel, and then
we're going to put that scalpel in a low temperature sterilization technology,
such as hydrogen peroxide, gas plasma in the absence of cleaning, and we can
see complete failure here. 60 out of 60
positive stainless steel scalpels.
Now,
let's look at the next slide where the only thing we did is we again put the
spores on the stainless steel scalpel.
And
then the next slide. All we did was
place the stainless steel scalpel into either distilled or tap water for 60
seconds, just placing it there for 60 seconds, taking it out and then putting
it into low temperature sterilization technology.
Next
slide. You can see that there is
complete success, a complete ability to kill microorganisms to include spores
just because of a static soak. Of
course, you can see here a very light rinse also was successful.
Next
slide. Here, we are going to,
essentially, try to identify why this is happening, and what we're really
looking at is what is going on as far as chloride, protein and spore
concentration by just doing that static soak.
In
the next slide, we see, essentially, in a matter of seconds we see the salt,
protein and spores released from the fetal bovine serum dried on stainless
steel blades and placed into deionized water at room temperature. In a matter of seconds, essentially, you get
significant reductions in salt, proteins and spores.
Next
slide. Now, so what we found is,
essentially, inorganic, organic and microbial contaminants on the device are
dramatically reduced during washing and, of course, there is a significant
reduction of spores.
Next
slide. Well, let's see if that is
effective for steam sterilization.
Right now, we're really talking about these low temperature
sterilization technologies. Does the
same thing happen with steam sterilization?
This is a study from Doyle and Ernst in 1967 where, essentially, all
they did was monitor the effect of spore occlusion and calcium carbonate
crystals in inactivation in steam dry heat and ethylene oxide sterilization
processes.
They
were just inoculating 103 or 8 times 103, bacillus
subtilis spores, and let's see what happened because of the spore occlusion and
calcium carbonate. Here, we see steam
at 121 degree centigrade in the unoccluded spores, the biological challenge. It only takes 10 seconds. You can kill 104 in 10 seconds,
no time. But in the presence of the
calcium carbonate, to kill that 104 took 150 minutes. For dry heat, it's three and a half
hours. In the presence of the calcium
carbonate, it's 50 hours.
Next
slide. So, essentially, a number of
things have been found, that is contact with water or cleaning for just a short
period of time rapidly leads to the dissolution of crystals, of course, removed
microorganisms and, of course, also has an effect on protein elimination. And, of course, minimal cleaning eliminates
the effects of these salts, which effect the effectiveness of sterilization
processes. And simulated use tests that
do not include washing would not represent conditions that exist in clinical
use situations.
Next
slide. And this is, essentially, what
you see in electron micrograph. If you
look at, essentially, .75 sodium chloride in the presence of spores, you see
the salt crystal, essentially, occluding the microorganisms from exposure and,
essentially, cleaning dramatically effected those results.
Next
slide. The point that we need to make
for all these studies that have involved prion inactivation is that you can
clean without sterilization, but you never can sterilize without cleaning. That is a point, a principle, that is known
to every professional in health care, certainly, every professional that is
involved in reprocessing instruments.
Next
slide. So the conclusions would be all
sterilization processes are effective in killing spores. Salts favor crystal formation and impairs
sterilization not only for low temperature sterilization, but also high
temperature sterilization. Cleaning
removes salts and proteins and must precede sterilization. Failure to clean or ensure exposure of
microorganisms to the sterilant could affect the effectiveness of the
sterilization process. We say
repeatedly if the organism does not have exposure to the germicide or the
sterilant, inactivation will not occur.
And, of course, these salts and protein materials and possibly other
environmental conditions to include surfaces affect that exposure. And lastly, CJD inactivation studies should
be consistent with actual clinical practice.
I
think we have done what we said we were going to do. We have looked at the recommendations from the U.S. We have talked a little bit about
methodology and how methodology affects results not only for hard to kill
organisms such as spores, but easy to kill microorganisms such as bacteria.
Next
slide. I thank you very much for your
attention.
CHAIR
PRIOLA: Okay. Thank you, Dr. Rutala.
Are there are questions from the Committee or from Dr. Taylor? Would you like to make a comment?
DR.
TAYLOR: Just a couple of quick
comments. A very nice talk and very
much to the point, I think. Two
comments, one is mentioned that generally, the washing procedure is usually
pretty effective in taking off bacteria and spores, etcetera. The one comment here is that one might
anticipate that TSE infectivity might, nevertheless, be somewhat more adherent
to instruments because of the hydrophobicity of the PrP protein.
The
other comment is that there are concerns about damaged autoclaves by hydroxide,
but that is certainly not inevitable, because it depends on the grade. The commercial company that we asked where
do you dispose of animal carcasses, in reactor vessels with hot hydroxide, have
had vessels running for many years now and they have certainly subjected these
to x-ray analysis, etcetera, and they are absolutely fine.
DR.
RUTALA: To your two points, I certainly
agree with the first point. Certainly,
the data that I presented, of course, are non-prion proteins, as well as
microorganisms and, certainly, the same type of analysis needs to be done with
prion proteins, and I support that work.
In
regard to the second point, the effect of sodium hydroxide on sterilizers,
certainly some sterilizer manufacturers have threatened the owner of the
sterilizer that in the hospital, if they use sodium hydroxide in the
sterilizer, they will nullify the warranty, which, of course, affects the
utilization of sodium hydroxide.
But
there are ways, as you probably know better than I do, to limit that
vaporization and contain the vaporization with, for example, containers that
have lids. And certainly, Dr. Asher
knows more about that, and possibly that can be discussed.
CHAIR
PRIOLA: I have one very quick question
before Dr. Rohwer makes a comment, and that is you stress very strongly that
instruments are always kept wet, so that you don't have this problem of
material drying on the instrument and then perhaps adversely affecting its
ability to be sterilized.
During
a surgery -- I mean, I know that when I do my little surgeries on mice that as
you're doing it, stuff does dry on the instrument just as you're poking around,
so how is that dealt with?
DR.
RUTALA: Well, many times -- it is dealt
with in different ways. Many hospitals,
essentially, don't even take the surgical instrument and put it on a dry tray. They very commonly place it in a basin,
which has, for example, saline or water or possibly even a germicidal agent, so
it doesn't go into a setting, which is going to allow dry fluids and tissues to
be achieved.
And
then, of course, the other issue is that there is sometimes precleaning before
it is sent to central processing.
Central processing does not want instruments that are contaminated with
tissue and blood. They won't accept
instruments that are contaminated with tissue and blood. Sometimes, there is also a washer sterilizer
that, essentially, is a precleaning procedure before it goes to central
sterilization.
So
different hospitals do different things, but the one thing in common is there
is an effort to keep it wet, and there is an effort to keep it clean, because
most central processing areas won't accept surgical instruments that have dried
tissue on them or bloody instruments.
CHAIR
PRIOLA: You know, I understand
that. I guess my point was more during
the procedure as you use the instrument.
Just as you're using it, it's going to air dry, because it is exposed to
the environment, and so you can't keep it wet the entire time. I mean, you're just going to have some dried
material that will probably be taken care of possibly by the cleaning and
whatnot.
DR.
RUTALA: Yes.
CHAIR
PRIOLA: But some drying will occur no
matter what you do.
DR.
RUTALA: And by immersion in the bath,
but some drying, depending upon the level, of course, will occur.
CHAIR
PRIOLA: Okay. Dr. Rohwer, what was your comment?
DR.
ROHWER: Yes, I have a couple of
comments and also would like to get Dr. Rutala's opinion on something, and that
is first, I would highly advise not autoclaving with the lid on the vessel, and
I am also mystified by this concern about sodium hydroxide vapors. As far as I know, sodium hydroxide has no
measurable vapor pressure, and a properly operating autoclave should not be
aerosolizing it either, because it shouldn't boil on the way down. That is my first point.
But
what I would like to have you address is this issue of cross-contamination at
the level of washing, because that is our major objection with that
approach. We're talking about an agent
that is very difficult to get rid of.
If it gets spread around the laboratory and the environment then, we
just couldn't tolerate that. You know,
we don't want it in our sinks. We don't
want it on our surfaces, etcetera. And
so how is it that you deal with the eluates and the washers that come off of a
set of instruments, which you know have been exposed to a Creutzfeldt-Jakob
Disease patient, for example? How are
the washers sterilized? How do you
dispose of that?
It
seems to me you create a cascading level of problems that have to be dealt
with, and I am absolutely willing to concede all the points that you are
making, except that in the case of this particular agent, it is very difficult
for me to accommodate this idea of spreading this stuff around, exposing myself
to it willingly before it has actually been decontaminated.
DR.
RUTALA: Well, to your point, certainly,
this practice has been employed for a number of years, and I think it has been
practiced for the following reasons.
One, of course, in a decontamination area in central processing, the
persons in that area wear personal protective equipment. They wear gowns, gloves, protective
masks. Second is that nearly every
hospital where decontamination takes place in central processing, it's a closed
unit. It's a washer disinfector
completely closed. That is there is no
aerosolization of droplets, of fluid that are related to the washing procedure. It's a closed procedure.
The
closed procedure, of course, has many steps to it. Some of them are the use of enzymatic detergents, high
temperatures, rinses and so forth. And
I guess the third point related to that, protective apparel, a closed system,
is the issue that while there is a recognition that some prion proteins may go
down the drain connected to a sanitary sewer, we don't believe that that's the
only source of prions reaching, essentially, the sanitary sewer system.
And
to that point also, prions, of course, are unlike many other infectious
diseases, are not transmitted by direct contact, indirect contact, droplets
spread, airborne or the environment. So
I am not sure what the level of concern is as far as transmission. It certainly wouldn't be, as I look at the
issue, transmission to health care workers, it could possibly be contamination
of the environment, but I believe that there is other forms of contamination in
the environment outside that setting.
CHAIR
PRIOLA: Dr. Bailar?
BOARD
MEMBER BAILAR: Very nice talk. I do have a question about this very simple
straightforward slide you had on the decreasing order of resistance of
microorganisms disinfected in sterilants.
You didn't go into the details of that, but I suspect it's ranked on the
basis of things we know are effective at the susceptible end, that is the
bacteria and the enveloped viruses.
Is
anybody looking at other kinds of agents, you might call them unconventional
agents, that may not be very effective at that end, but might be pretty good
with prions?
DR.
RUTALA: As far as that slide is
concerned, you know, it is a general slide.
There is an exception to that slide as it pertains to germicidal agents
and particular groups of microorganisms, but it is a general slide. As your question intimates, most of the
data, and there is literally hundreds of papers that support that slide, and
most of the data is with the types of germicides that you have seen today and
in my presentation and in other presentations.
That is the conventional, not the nonconventional germicides and
sterilization processes, products such as the alcohols, the phenols, the
ethylene oxide, the steam sterilization and dry heat and so forth. Does that answer your question?
BOARD
MEMBER BAILAR: Yes, it leaves me
wondering whether it might be worthwhile for somebody to try some of these
things, but I am no expert in this field.
DR.
RUTALA: Well, to your point, I think
that there are people interested in that.
CHAIR
PRIOLA: If there are no more questions,
we have two more speakers, but I think we should take maybe a 10 minute
break. We're about 15 minutes behind,
but let's take a 10 minute break and reconvene at 5:10.
(Whereupon,
at 5:06 p.m. a recess until 5:18 p.m.)
CHAIR
PRIOLA: If I could have the Committee
members take their seats, so we can conclude this session. Okay.
Our next speaker is Captain Edward Rau, Environmental Health Officer for
NIH, and he is going to discuss infectivity of air emissions and the
incineration of scrapie tissue.
CAPTAIN
RAU: Thank you very much. Unfortunately, our only TSE expert in our
group, Paul Brown, couldn't be here today.
He is on some kind of a hardship assignment in southern France at the
beach right now, so he has left that up to me to take care of.
The
other disclaimer is that the results that I am going to present here are really
very preliminary. Our experiments are
still in progress. The paper is not
written. None of the data is
published. So with that, we'll go ahead
here.
Could
I have the next slide, please? I don't
need to elaborate on all of the difficulties there are in inactivating the
prion agents, and that the resistance to thermal inactivation is, of course,
highest under conditions of dry heating.
And that has led some concerns about even incineration being an
effective technology to dispose of certain TSE waste.
Sitting
here this afternoon, I think I have a new definition for incineration. It's that process, which incorporates all of
the things that make inactivation difficult to do. We start out with a material that has been smeared and mashed
around by all the handling of the medical waste process. It has not been precleaned. In fact, it is, in some cases, pure
dirt. Then we're going to take that
into a process, which begins by a drying and probably fixation step, and maybe
melt a few varnish like materials over it before we really get into the
combustion process.
Next
slide, please. As you are aware,
incineration is still the technology of choice for disposing of most medical
waste that contain TSEs, and it is also being used to dispose of large volumes
of animal products, contaminated carcasses and so forth, some of which are
still in large quantities in storage.
The potential for TSEs being in emissions from combustion processes is
of public concern, and has received very little investigation so far.
Next
slide, please. We have published some
previous experiments documenting the unprecedented level of resistance to
thermal inactivation, both crude brain tissue and purified PrP from the 263
scrapie infected hamsters. That
included a partial inactivation after heating for 300 degrees for 15 minutes,
and several transmissions after actually ashing brain material at 600 degrees
C. And at those higher temperatures,
there were similar patterns of resistance in both formalin fixed and non- fixed
tissues.
Next. The objectives of our experiments in
progress are first to confirm the results from our previous study. Some people were a little bit skeptical
about the transmission after 600 degrees C.
Others might have even labeled it science fiction. And secondly, we wanted to investigate the
potential for transmission via the air emission that might come from a medical
waste incinerator.
Our
previous experiments were rather primitive in that we merely headed brain
tissue macerate in vented crucibles.
The new experiments, we are actually going to simulate the conditions of
humidity and the gas mixtures and so forth that occur in two types of
incinerators.
The
first are the reducing environment or starved air incinerators. These are the most commonly used type of
incinerator in the United States. A
synonym for that is the controlled air incinerator. The other situation we wanted to look at was an oxidizing
environment or referred to as a normal or excess air incinerator. And in this study, we repeated the
temperatures that were used in the previous study, 600 degrees and 1,000
degrees C.
Next. Materials, our tissues samples were cooled,
hamster brain tissue macerates from terminal animals with the 263 scrapie
strain, about 10 logs of infectivity per gram, and for controls we had tissue
from normal animals. The incineration
situation was referred to as a Lindberg Furnace in a quartz reactor tube, the
removal of the specimen crucible and holder.
The gas supply coming into the incineration unit was normal air or
nitrogen with flow and humidity controls.
And then coming out of the unit was an impinger train and terminal
filter for collection of the air emissions.
Next
slide. This is a photograph of the main
part of the simulator. It all fit in a
large chemical fume hood. The incoming
gases come into the furnace here. The
quartz tube is contained inside of the Lindberg Furnace. This is a pyrometer. And then the outflow coming out of the
combustion process goes into this impinger system, a series of collectors, the
first one being ice water bath. The
second one, dry ice, and then out through a terminal filter.
Next. This schematic gives an idea of the inside
of the reactor tube, inside of the furnace.
The tube is, approximately, one inch diameter, all quartz construction. At this end, we can remove the plug and
insert into a thermocouple to directly measure the temperature or we can insert
the sample on a glass rod, which is housed right here.
The
gas flow comes from this direction in the top.
It passes the sample. Exists
through a ball joint and then on to the impinger train. In designing this, we tried to ensure that
all the components in the system were inert.
We used quartz and teflon joints as the materials.
Next. This schematic shows a little bit more
information about the impinger system.
It's a rather challenging design, because we really didn't know what we
were trying to trap coming out of this process. Again, we have the ice water bath and the crushed dry ice bath
following into a cartridge filter and exhausting into the hood.
Next. The methods began by introducing a one gram
sample of the brain tissue into the reactor tube. We incinerated that for 15 minutes at either 600 or 1,000 degrees
in either normal air or starved air conditions. Following the process, we collected the air and air emission
samples separately from each run, and then replaced the impinger train between
each run. We didn't have enough reactor
tubes to use one for each experiment, so we disinfected those with bleach after
each test.
Next. So each experimental run gave us three
different samples. We had the ash
residue that was collected in the crucible.
The small amount of residue that formed in the reactor tubing as it
exited the quartz reactor and came out cooled down at the border of the
furnace, and then the emissions collected in the impinger traps.
Next. This gives you the array of samples that we
collected. We ran both normal and
infected tissues in the two different gases, two different temperatures, and
for the infected material, that gave us three different samples. We combined samples for two of the normals
and we did not run some of the exit tubes and traps on those. The reason for that is simply
economics. We're dealing with about 500
animals to be maintained here, a great cost and time.
Next
slide, please. The bioassay method, we
concentrated the samples of the ash and emissions from each test into,
approximately, a one milliliter volume in saline, and that was intracerebrally
injected into, approximately, 30 Wingling hamsters. That is about 3 hundredths of a milliliter per animal. So the entire emission from each test was
injected. We're not taking a sub sample
out of those emissions. The entire
emission from each burn was injected into animals.
Animals,
of course, were segregated by test group.
We observed them over 12 months for symptoms and then examined all of
the brains for the presence of prion proteins by Western Blot testing. That testing is still in progress on the
negative appearing animals.
Next. Results, we had no transmissions from the
controls. There was some possibility,
my commentor thought, we might be able to get some kind of symptoms as
artifacts of this trauma and injecting into the animals, this residue. We just wanted to rule that out. We had no transmissions from any of the
materials collected at the 1,000 degree C burn, and we had no positives from
any of the residues collected in the impingers or the end of the reactor tube.
However,
we did get two transmissions, and these were after very long incubation time
from the ash from the crucible from the 600 degree group in normal air. The asymptomatic animals, again, we're still
testing those for silent infections.
We're not finished with that, so we have to call our results
preliminary, at this point.
Next. Conclusions from the experiment. First, that the results were very similar
from the previous study showing that there is, apparently, a threshold
transmission from tissues at about 600 degrees C. The low transmission rate and very long onset time for the
symptoms suggest, again, that that is the extinction temperature or very near
it. We found no evidence of infectivity
in the air emission samples.
Next. Speculate a little bit about what the
environmental implications of this are.
First, we did not see any evidence of transmission in the air emissions,
so it's probably unlikely that will have a possible emission to the air from a
properly operated medical waste incinerator.
It is possible that some survival of the agent could occur in ash if there
is not enough penetration of the temperature and time of exposure in the ash
bed.
I
wanted to kind of put this in perspective a little bit though, because I think
it's a very low potential for transmission.
First off, as Dr. Taylor said, we don't see environmental sources in
transmission going on. Secondly, what
we are seeing these two positives on is a simulated burn load that is pure
material with 10 logs of infectivity per gram, and we're collecting virtually
of the residue from that and injecting it into, approximately, 100
animals. So there is a very low level
of infectivity in that material that is coming out.
Probability
of survival in ash not only depends on a lot of factors, the load density, the
turbulence, the type of equipment, other operational factors. And as we heard earlier today about the
importance of context, I really can't imagine a more complicated context than a
medical waste incinerator and all the combustion and mixing and reactions and
things that are going on inside of that process.
Next. Our colleagues at EPA felt that these
factors probably would be the most important in increasing the potential for
prion survival in ash from medical waste incinerators. Under normal conditions, there are certain
design factors, particularly in the grate oriented designs that might allow
some of the material to not be treated for 15 minutes. It falls through the grates or it somehow
gets passed on through the system faster than the nominal residence time for
the solids.
Particularly,
as things are just inserted into the incinerator, you tend to get a boil off of
some of the material, a flash burn.
That can be carried over very quickly into the second chamber. The other factor is that the ash bed
temperatures often may run 100 degrees C lower than the actual air temperature.
Reported
temperatures for incinerators refer to the air. That is what is being monitored, and not the actual temperature
in the ash. Under abnormal conditions,
a lot of things can really go wrong, cold start up conditions, overloading,
inadequate control of the under fire air flow.
Next. We tried to compare what our experimental
conditions were with the conditions in actual medical waste incinerators and
some other types of processors used for bone meal products, and the most common
incinerator in the U.S., the controlled air or starved air type of incinerator,
in that primary chamber, you have temperatures of about 760 to 980. If you subtract about 100 degrees from that,
allowing for some cooler temperatures in the ash, we're right on that threshold
of survival that we saw in our experiments.
The
secondary chamber, which mostly sees the pyrolysis products and not the ash is
usually well up into that 1,000 degree temperature. I don't have much information on the temperatures in the excess
air incinerators. They are probably
quite variable because of the way that process is run. The secondary chamber is, again, quite hot,
sufficient probably to inactivate prions.
There
is some information on the other types of burn units that are being used in
Europe for disposal of the meat by-product material. One of the articles had indication that there is actually a
measurement of temperature at the ash grate in this 800 to 1,000 degree range,
which looks pretty good for inactivation.
Next. There are other possible incineration
options. If we get into a situation
where we have a large amount of material to dispose of, the mass burn municipal
waste incinerator in the United States operates at about 1,000 degrees, so that
would likely inactivate prions.
Western
Europe is looking at a variety of other types of industrial incinerators, fuel
burners and so forth. Again, they have
some high temperature and residence times.
In one system, a holding time of 30 to 40 minutes, which is very
encouraging.
I
think that's the last slide. Next. I believe that is the last slide. Again, our results are very preliminary. So far, all of the testing on the negative
appearing animals is confirming that's the case, but we're not quite finished
with that yet. Any questions?
CHAIR
PRIOLA: Any questions from the
Committee? All right. Thank you very much, Captain Rau.
CAPTAIN
RAU: Thank you.
CHAIR
PRIOLA: Our final talk of the -- oh,
I'm sorry. You had a question? Oh, sorry.
Yes, go ahead.
DR.
ROHWER: One issue I had with the
original study was that the thermocouples were not actually in the sample, and
the sample was loaded wet, and it wasn't clear here whether you're starting
with wet tissue or dry tissue, and where the temperature measurement is actually
being made, vis-a-vis, the sample. And
the reason I bring this up is that a wet sample will not spend as much time at
600 degrees as a dry one, because you got to boil off the water first, and that
could actually take some time.
CAPTAIN
RAU: We did start with wet samples, wet
tissue samples. The burn time is 15
minutes, however, in here, so I think we're probably spending most of that time
at temperature. With regard to the
thermocouple, before each sample was inserted, it was measured, then the
thermocouple was withdrawn. We still
have a pyrometer on the outside of the tube that we're confirming temperatures
with, and that is really the best we could do.
There is just not a way to have the thermocouple in there and be able to
insert and withdraw a sample out of there.
I agree with your boil off concern, but that is also real world, what's
happening in the incinerator.
DR.
ROHWER: In the original experiment,
there was a thermocouple between the crucibles, so I took that to mean that
there are thermocouples that can survive those kinds of temperatures. Is that incorrect?
CAPTAIN
RAU: Yes, the problem is getting the
output out of the burn chamber. It was
a design issue. But in the first
experiment, the thermocouple was right adjacent to the crucible and we were
able to measure that in the muffle furnace.
CHAIR
PRIOLA: Okay. Thank you. I think we'll
move on to our last speaker who is Dr. David Asher from the FDA, as well as Dr.
Brown, Dr. Stanley Brown. Oh, actually,
you're going to start.
DR.
BROWN: Actually, the last team is -- I
am the rigger coming in from CDRH.
Could I have my first slide, because it tells the whole story? Let's see, it worked on my computer. It was created on Terry's computer.
Well,
anyway, I'll start. I'm Stan
Brown. I am an engineer from the Center
for Devices. I will present the first
half of the team effort between myself and David Asher's group, which was
funded by the FDA Office of Science, and these data, some have been published. Some are preliminary.
Some are in manuscripts in preparation, and this is not good news on the
screen.
Basically,
what we were doing in my side of town was to look at four questions. The first question is can you safely
autoclave in sodium hydroxide without wrecking your autoclave? The second question is what are the effects
of the WHO protocols on surgical instruments?
The third part of that was to develop an experimental instrument that
could be used in a simulated instruments contamination study that would be
compatible with the hamster model that David Asher has. And the fourth was to do some -- there we
go. Okay. Let's click through here.
First
of all, instruments from CDRH, we were thinking about primarily reuse, as you
have heard, or reusables, but there is also growing concern about these things
called SUDs or single-use devices, and with the law we are now reevaluating how
we assess the reprocessing and validation of some of the reusables,
particularly those that are neurological or other type of tissue contact. From the CBER point of view, as you know, we
are talking about contact of instruments that you use for tissue preparation.
Next. Disclaimer.
We developed these methods, because they fit within the financial
constraints and the laboratory constraints.
In our laboratories, these presentations do not constitute a regulatory
endorsement for these methods. They are
simply methods we thought would get answers that we're after.
Next. You know all about this story and we're
primarily interested in the sodium hydroxide autoclaving phase and the soaking
in sodium hydroxide or bleach.
Next. So if you go to the CDC website, you will
see that there are a couple of warnings.
One is that if you autoclave in sodium hydroxide, you wreck your
autoclave and two, if you soak in bleach, you will wreck your instruments. And these are based on the studies that we
started a few years ago, and I think CDC may actually have now inserted the
reference from our work on that.
Next. Again, there are the four questions that we
are trying to answer.
Next. The first one has just been published in the
American Journal of Infection Control with Kathy and myself.
Next. She did the work and I wrote the paper, so I
got to be first author. And again, the
question is the autoclave manufacturer said if you autoclave in sodium
hydroxide, you don't have a warranty on your autoclave. Knowing, of course, this must be done in a
gravity displacement, so it doesn't fit in the standard central storage big
prevacuum type autoclave. It has got to
be controlled with a liquid cycle.
Next. Two approaches. One was we would put a liter of sodium hydroxide in a pan and
cover it. Two, we would put some sodium
hydroxide in a beaker and put the beaker in a pan and cover it. And then we put it in an autoclave. We did repeat one hour sterilizations, and
we did these at 134C just to be a little more extreme, and we did them for an
hour, up to five cycles, and we put pH paper inside, outside, everywhere. We put pH meters inside and outside, and it
is a closed system for the little tabletop with six liters of water
reservoir. We put it through five
cycles to see what happens to the water in the reservoir to see, again, what
happens to the autoclave.
Next. And we got thinking about pans and
lids. This one probably, the condensate
will get on the top and drip down and wreck your autoclave. This one, it might wreck your autoclave and
it might drip inside. Some of them have
little nipples or construction bars. To
me, that is a Black Iron Dutch Oven where these drips, so that they roast while
you're doing it. And then you have got
some that actually have gutters that contain the lid within the pan. So part of this is what kind of pan and lid
design you have.
Next. The two that we used successfully was a
Nalgene Instrument tray shown here.
This was filled with a liter of sodium hydroxide and closed.
Next
slide. And if you look here, you can
see this is a drain on that gutter, so the condensate goes back into the
pan. It doesn't go out.
Next. And there you see the lid that is fully
contained within the gutter and, of course, this has been used for years. It's for control of human waste and
biohazards.
Next. The other type of pan was a Lid (D), which
has a lip on the lid. It also has
crossbars that act as condensate drip spots.
Next. And the results of this were no pH changes
outside the containment. Inside the
lids were very caustic. The bottom of
the pans were very caustic. There was
lots of vaporization, condensation going inside, but it was all contained
within the containment vessel. So we
conclude that if you use this kind of -- if you use the right kind of pans and
lid, you can do it without wrecking your autoclave. And, of course, those of you who have been doing this in the lab
for years know that. Obviously, hot
caustic is hot and you have to be careful.
It probably cannot be done in a standard central storage autoclave and it
may require larger approved type pans.
Next
question. What do these things do to
your instruments? Next. What we did was we bought surgical
instruments from Roboz, which is a medical device supplier in Rockville, and we
bought lab stuff from VWR. Some of them
are labeled Germany with CE marks and some of them were labeled Pakistan, and
there are some members in the audience who will appreciate this. We put them through repeat cycles of the WHO
including autoclave and water.
Next. And here you see some carbide tipped needle
holders. This one has been through five
times autoclave and sodium hydroxide, five hours in sodium hydroxide. There is a little bit of blackening in the
box joint. This one was soaked for one
hour in Clorox, and you can see there is a tremendous amount of corrosion going
on at the box joint and around the carbides.
Next. These had beautiful gold handles, high
quality instruments, and that is one hour in bleach. So if you got gold handles, don't bleach them. This is five times gold handles autoclaved
in sodium hydroxide. It looks fine.
Next. This is a German pair of scissors versus a
Pakistani pair of scissors, five times in sodium hydroxide. You can't photograph shiny, but this is
shiny. It just looked great, and this
looked really dark and dingy.
Next. This is Germany versus Pakistan, and you
notice the Roboz label on this thing as stainless steel, and this tubing clamp
around this weld really took it with the Clorox. This tubing clamp after five hours in bleach, this is the 6
percent, which is what, 2,800 parts per million. It looked fine. So some
go, some are fine.
Next. So the conclusion of this, and I didn't show
you any pictures. Titanium really does
not like sodium hydroxide, and this is well-known in the material science
literature, as well. Soaking in sodium
hydroxide, they couldn't care less, none of them. Soaking in bleach, some did fine, some didn't. The problems were the gold handles and the
welds. But the important message here
is if it's going to corrode, it's going to do it first time. So you don't have to do a long
experiment. If you put it in Clorox and
it comes out rusty, you know it's going to rust and you go on and find a better
instrument.
Next. So Part 3.
We wanted pins and this is part to lead into an animal model of the
simulated instrument for David's studies.
He was using a 25 gauge needle on a half cc syringe in his animal work
for injection, and we wanted to make pins instruments that were like that, but
we also wanted to be able to suspend them over 96 weld plates, so we could do
serial dilutions of bacteria, viruses in brain homogenate, and the system
needed to be autoclavable.
Next. So there is the syringe needle that he was
using. That is a good old copper penny,
and what we did was we took Eppendorf tips.
My wife is a microbiologist, and so is Kathy. We took half millimeter stainless steel pins. We took some epoxy. We used the Eppendorf to draw the epoxy up
into the tip, stuck the metal pin into it, hung it in a rack, put it in an oven
to cure and now, you have things that can be autoclaved, and it's the same size
as the needle that is used. So from an
ergonomics point of view, it's something that he would feel comfortable with, I
think.
Next. And here, you see the setup. This is your standard Eppendorf rack with
modified stainless screws to adjust the height, and the pins were sitting in
the wells of a 96 weld plate, and Kitty, that after a little bit of practice,
she could actually get all the needles into all the holes, right?
Next. Finally, we wanted to do some preliminary of
adhesion of both blood and tissue and looking at WHO protocols, and one
question was what about damage and adherence?
So we were using stainless steel pins and we also made pins out of piano
wire, which really did not have a good time in Clorox.
Next. Pins are placed in a rack and stuck into a
slab of liver for an hour, and then we left to dry as a worst case. The pins were stuck in a 96 weld plate in
sheep's blood for an hour, and then they were left to dry. We went through ultrasonic cleaning, which
the standard protocol is 60 degrees C with an enzyme cleaner. We autoclaved in sodium hydroxide. We soaked in bleach and we got unclean
controls.
The
results were the unclean ones, that the protein was more adherent from liver
than it was from blood, and the amount was using Bradfords reagent. It's about the equivalent of -- our minimum
was one microliter detection limit.
Damaged pins did not seem to be more adherent, so that the blackening in
the box from autoclaving and sodium hydroxide is probably not a problem. Repeat exposure did not show accumulative
effect.
Next. So then Kathy wanted to do some
bacteriology, and she soaked them in a solution of staph epi. for 24 hours, let
them dry and did the same kind of cleaning things, and then stuck them into an
agar in a test tube and incubated for 24 hours.
Next. Lo and behold, autoclaving in bleach killed
everything. So we had to try some
modified WHO. So we dropped the sodium
hydroxide autoclave and the ultrasonic cleaning was done at room temperature.
Next. So what we found was that only the pins and
bleach showed no growth, but, of course, we don't know if we cleaned it or we
just killed them. The other produce
showed fewer protocol than the untreated control, but bacteria was still
present. And then the question is are
we cleaning or are we just killing?
Next. And we tried some SEM work, and you can see
a little small column of staph epi. here on the uncleaned tip. It was very unconvincing in terms of whether
we really were cleaning or we were just killing.
Next. So our conclusions were, first of all, yes,
you can safely autoclave in sodium hydroxide with the right pan and lid. Some WHO protocols can damage some
instruments. Discoloration does not
seem to impair function or cleaning.
The bacteria leave a lot of unanswered questions, and the questions for
prions, of course, we don't touch them in CDRH, that is David's role.
So
next, and I will turn the podium over to David to talk about his hamster
studies.
DR.
ASHER: Thanks. You can go right to the next slide,
please. Thank you. I can't believe that it's almost 6:00 and
we're still giving talks. Quite a few
people in CBER participated in TSE related activities. These studies are really involved only
people in CBER in my own little group, and especially Kitty Pomeroy who I think
is still here in the back. Without her
holding the whole enterprise together, there is no way that we could have done
this.
Next
slide, please. And among the staff at
CDRH, of course, Kathy Merritt and Stan Brown, who has just spoken, and I don't
believe that we would have gotten the funding to do what we have been able to
do without Stan's efforts.
Next
slide, please. We have developed two
simple methods to evaluate methods for decontamination of TSE agents dried on
surfaces. In this talk, I am going to
concentrate on two regimens that more or less replicate recommendations of the
World Health Organization consultation.
We used two general models for both of which 263K scrapie was the agent.
The
first model was a modification of a method for evaluating virucides that was
described years ago by John Chen of the Environmental Protection Agency. He dried viral agents onto glass cover
slips, treated them and then assayed residual virus.
The
second method was stimulated by two reports from Charles Weissmann's group, and
Professor Weissmann will speak here tomorrow morning. They dried scrapie agent onto steel wires implanted into mouse
brains. We didn't do that, but as you
saw from the devices that Stan showed you, our model was very similar.
Years
ago, we had done a couple of experiments with model squirting scrapie through
actual hypodermic needles, but it was very cumbersome, and we never followed up
on it. We have used the first method.
Next slide, please. The first method for many years, simply
dropping suspensions of scrapie infected hamster brains onto glass cover slips.
Next
slide, please. Then they are dried in a
petri dish in a hood.
Next
slide. And then they can be exposed to
any number of disinfectant or decontamination regimens, here potassium
permanganate solution that can be autoclaved, that can be soaked. You can do all kinds of things with it.
Next
slide, please. Then they can be ground
up and supernatant fluid assayed. Now,
we used plastic pipettes and tubes for the whole procedure, because they are
disposable. We don't have to worry
about potential carryover of infectivity.
Although, the original method described by Chen used 10 brack tissue
grinders of the kind seen here. So the
slips are ground up in an ml of diluent.
Next
slide, please. The glass is allowed to
settle out, and then the supernatant fluids are assayed for infectivity by
intracerebral injection of hamsters.
Next
slide. If the hamsters get sick, their
brains are removed and then they are checked for protease resistant prion
protein as evidence that scrapie agent was present and was not eliminated by
the decontamination regimen.
Next
slide, please. For reasons that may
become clear at the end of this talk, I have about six minutes left, we think
that it might be useful to do immunohistochemistry on some of these brains, as
well. Although, we haven't done that
yet.
Next
slide, please. One advantage of the
method is that you can rid of residual toxic disinfectants, Robert Somerville
talked about that problem this morning, by simply rinsing the cover slips in
distilled water to get rid of things like Clorox, which is really terrible for
assay animals.
As
you see here, this is sort of an upside down dose response curve. It's hamster survival times plotted against
the dilution of supernatant fluid. And
here are three curves, one for unsoaked or dipped slides, one that has been
soaked in water, and the other that was soaked in water and then dipped in
water a second time. And you can see
that they are virtually superimposable.
None of the infectivity appeared to come off on this exposure to
distilled water.
Next
slide, please. And it was using this
method that we first demonstrated the resistant fraction of infectivity that
survived drying on glass and then prolonged exposures to steam autoclaving at
elevated temperatures.
Next
slide. To investigate some of the
performance characteristics of the model, you can imagine doing many, many
assays in hamsters is extremely cumbersome.
We tried drying specimen samples of polio virus and two other viruses
suspended in brain onto glass, and then titrating multiple samples. The results summarized here suggest that the
agreement from test to test and day to day was reasonably good, but that there
was enough variability, so that a controlled titration really should be done in
each test. And it might even be
reasonable to consider putting in a test decontamination, a reference
decontamination treatment. Although, we
have never had enough resources to support that.
Next
slide, please. When we began to work
with the scrapie brain suspensions dried onto the steel needles that Stan Brown
has just showed you, we decided first to do some similar preliminary studies
with conventional virus suspended in saline containing 10 percent brain
extracts to get some idea of how the scrapie agent might be expected to behave,
so we wouldn't waste months and months on a model that wouldn't get enough
infectivity on.
But
we were surprised when neither polio virus nor porcine parvo virus suspended in
brain appeared to stick to the steel needles at all or at least we couldn't
detect any of them in cell culture assays.
We had no trouble getting them to stick to glass, but we couldn't find
detectable porcine parvo virus or polio virus dried onto steel needles.
So
now wanting to take a chance that we would set up our test with the limited
funding available and the find that we hadn't had enough challenge agent stuck to
the steel needle, so we decided to suspend the 263K scrapie hamster brain in a
normal brain paste, and we used a calf brain as the source of the normal brain
material.
Next
slide, please. I must say we decided to
check out our concern. We had enough
hamsters to do a rough titre of 263K scrapie diluted in phosphate-buffered
saline without any additional brain material, and it appears that the scrapie
infectivity in the saline suspension did stick to the steel wires. You will notice we get positives out to a
dilution of 10-5, so that the behavior of the polio virus and the
porcine parvo virus does not appear to have predicted the behavior of the
scrapie agent.
But
the studies that I am going to summarize in the next few slides use scrapie
infected brains as a paste to charge the needles. The glass was charged in the way that I described previously with
saline suspensions.
Next
slide, please. So let me summarize for
you the general design of the efforts.
We looked at two variations of two kinds of decontamination regimens
that generally resemble those recommended by the WHO consultation, and then
after that, I will add some other results that we thought you might find of
some interest.
First,
as we have mentioned, infected brain is dried on the objects, either a saline
suspension on glass slides or tissue paste onto steel needles. Then come the decontamination steps, which
are either a chemical soak in one normal sodium hydroxide in the autoclave for
30 minutes or a soak in sodium hypochlorite, full strength chlorine bleach from
the grocery store at room temperature for 60 minutes followed by an autoclaving
at 121 celsius for 30 minutes or at 134 celsius for 90 minutes. The autoclaving with sodium hydroxide is in
the sodium hydroxide. The autoclaving
with bleach is after it is in water.
Following
that, all these materials were cleaned in an ultrasonic cleanser using a
laboratory proprietary detergent with a pH of 9.45. All of them got this, because, as pointed out by Dr. Rutala, it
is important to try and replicate the conditions under which these things would
be done in a hospital. The sonic
cleaner was cranked up the highest temperature that it would take, which was
over 60 degrees, although, somewhat variable, and for the longest time the
timer kept, which was for 90 minutes.
This
was done by putting each object into a separate tube filled with the cleaner,
and then the tubes were immersed in more cleaner in the chamber. Following that, there was a water rinse and
then, finally, a terminal sterilization in the autoclave at 121 celsius for 20
minutes in order to model what we took to be standard hospital practice.
Next
slide, please. As I mentioned, the
sonicator was set at maximum temperature and time. We only did the one set of conditions. We made no effort to select a better cleaning solution. I am sure there are many others, that many
others are available.
Okay. Next slide, please. First, the assay technique for residual
infectivity on the glass slips. Each
experiment, a positive control consisting of 10 slips each holding .1 ml of
dried on scrapie infected, 10 percent hamster brain dried down, not exposed to
any decontamination regimen, ground to a powder and a ml of PBS glass allowed
to settle. Fluids were then pooled,
tenfold dilutions performed in phosphate-buffered saline, each dilution assayed
in four hamsters, the same volumes that Dr. Rau showed you, .03 ml each
intracerebrally into the left frontal lobe.
That is simply so that we would know in each test how much infectivity
had been used.
Next. Hang on a second. Let me finish. Each of 10
slips then was exposed to some decontamination regimen, and then also ground to
a powder in phosphate-buffered saline, the fluid assayed as for the controls,
so that each experiment on glass involved 10 slips and 40 hamsters.
As
David Taylor had mentioned to you earlier, we deducted incidental deaths. We took 45 days as the cutoff between
considering it a death incidental.
Perhaps we shouldn't have done that.
Next
slide, please. For the steel needles,
the positive controls were tenfold, dilutions of infected hamster brain as a 10
percent paste in normal calf brain, and then serial dilutions were done with
normal calf brain paste. Four needles
were charged for each dilution, dried and then a separate hamster was assayed
for each needle. For the actual test,
40 needles charged with 10 percent hamster brain and normal calf brain, dried,
tested and then assayed as for the control above. Less incidental deaths occurring before four days.
Next
slide, please. This is just to show you
what a titration on glass looked like, the interim score here at eight and a
half months. Notice that the last
positive animal so far as those inoculated with a dilution of 10-8
calculated from the original brain tissue.
Next
slide, please. And a similar titration
for scrapie dried onto steel needles, also positive to a reasonably high
dilution. Actually, somewhat higher
than we got with the saline suspension, so we weren't sorry that we had used
the brain paste.
And
you might notice that there is one negative at the lowest dilution. That was an animal that died at 55
days. Brain was negative, and that is
why we're wondering whether 45 days might have been the best date to estimate
incidental deaths.
Next
slide, please. Before I move on to the
actual results, we were interested to see what ultrasonic cleaning in hot
alkaline detergent alone without any other treatment would do, so we did a
titration from that and found a substantial reduction, both of the infectivity
on glass and on steel needles from the hot ultrasonic cleaning alone. The log reduction factor is slightly over 5
logs. Although, for both models there
was some residual infectivity left on the surface.
Again,
we made no effort to optimize, to modify or optimize the procedure. We presume that most of the infectivity
probably went into the liquid, but we haven't made any attempt to find out
whether that is true.
Next
slide, please. So here are the WHO
studies. After exposure of glass slips,
there are the glass slips, to sodium hydroxide or to sodium hypochlorite with
autoclaving at either 121 celsius or at 134 celsius, there was obviously a
dramatic removal of infectivity, but darn, one of the animals assaying material
exposed to one normal sodium hydroxide at 134 autoclave, 134 celsius for 90
minutes has come down positive.
Next
slide, please. And similar experiments
with steel yielded relatively similar results, at least two, maybe three of the
assay animals have had positive Western Blots.
We are going to have to check those out. Obviously, these stray positives have been seen before, and we
have to convince ourselves as to whether they are really positives or whether
it's inadequately digested PrP in the Western Blot or whether it's real.
So
the methods are, obviously, highly effective.
They saved almost all the hamsters and removed so much infectivity that
most of the objects assayed didn't show evidence of contamination. Remember that each of these objects was charged
with at least a million lethal doses of scrapie agent, but we can't say that
they are perfect.
Next
slide, please. We have seen similar
results in the past using single chemical soaks. These are all done with the Chen glass test. And, again, we have frequently seen, these
are sodium hydroxide soaks at various temperatures, an occasional stray
positive.
Next
slide. Some tests have found no
positives at all, but remember with the Chen glass test, we sample only about
12 percent of the supernatant fluid from each slip, so that these results are
not necessarily different from the ones that show single positives. There is a substantial sampling problem when
you're dealing with very small amounts of infectivity.
Next
slide, please. Here is another result
with sodium hypochlorite where we had no positive animals. I marked these all as interim, because we
haven't finished all the Western Blots even though some of these are not new
experiments.
Next
slide, please. And finally, I would
like to say that some other chemical agents are probably also very
effective. Here are some results using
concentrated formic acid, which is used to treat tissues for histology and
immunohistochemistry. Note that there
is only a single positive animal out of 37 tested. Reports of a commercial phenolic disinfection, at least
temporarily unavailable here in the United States is reported to be very
effective, and we have heard that there are other decontamination regimens in
development not yet ready to share with the FDA or the public that are showing
promise.
Let
me conclude now by, next slide, please, just summarizing that methods developed
to evaluate the effects of virucides are adaptable to evaluate decontamination
of TSE agents. Studies with two models
both suggested that exposure to 263K scrapie agent dried on surfaces to
solutions of sodium hydroxide, sodium hypochlorite with simultaneous or
sequential autoclaving and ultrasonic cleaning in hot alkaline detergent
markedly reduced amounts of infectivity, and the risk that any object would
retain detectable amounts of agent.
Other
chemical treatments may also be effective, but uncertainties remain. One, the reliability of the decontamination
procedures, not only the fact that we see stray positives, but also there is a
theoretical concern that the predictive value of these results, the results
from such models, may not adequately predict the behavior of decontamination
regimens in the actual health care or manufacturing setting, concern that there
may be sanctuaries of the kind that Bob Rohwer and David Taylor have discussed
that might occur in manufacturing processes or health care setting that would
impair the ability of otherwise effective decontamination regimens to act.
It
is quite late, but I am happy to answer questions for anybody who has got the
energy still to ask them. Thank you.
CHAIR
PRIOLA: Are there any questions for Dr.
Asher or Dr. Brown? All right. If not, I -- okay, Dr. Somerville has one
for you.
DR.
SOMERVILLE: I'm just going to make a
brief comment about the first part of the talk, and that is that in our
experience, in our survey, we find that the various different grades of
stainless steel are used from the manufacturer of surgical instruments and with
various different finishings, and they have different responses to the kinds of
treatment that Stan Brown was trying on the instruments.
The
one brief question I have is have you tried anything other than visual
inspection to see what the degree of damage is being done to the instruments?
CHAIR
PRIOLA: Can Dr. -- oh, he is coming up
there. Dr. Brown can answer that.
DR.
BROWN: The answer, at this point, is we
have done nothing other than visual, and part of the next generation of study
is to be looking at some of the different alloys, some of the different
corrosion test methods. Some of these
effects are so blatant that why both to -- I mean, I cut up the gold handles
and put them in the SEM just to make sure it really was gold.
And,
in fact, there was gold on those handles, but no, we haven't gone any
further. But one of the questions is
are the different grades, you know -- in talking with the instrument
manufacturers and the people who do chemical analysis of instruments, there are
a whole wide range of grades of stainless steels, but the manufacturers will
tell you what probably is the most important is actually the mechanical
treatment in terms of how they make them, coworking, etcetera. And it may not be a matter of chemistry, but
it's a matter of mechanical parts.
The
finger rings very typically are attacked by Clorox, and that is an area where
there has been a lot of mechanical cowork to form the rings. Whereas, elsewhere on the same instrument,
the surface looks fine. So it's not
just the chemistry, but it's actually the mechanical processes used in the
forming or fabricating. And again, if
it's going to go, it's going to go the first time you throw it in bleach.
BOARD
MEMBER HOGAN: Dr. Brown, before you
leave, I have one more question. Dr.
Brown, could you get rid of the black deposit that formed on the sodium
hydroxide instruments?
DR.
BROWN: First of all, we didn't do any
other cleaning. We just over and over
and over, autoclave and bleach. We
didn't use what do they call it, milk, the cleaning milk that is used in
standard central storage.
BOARD
MEMBER HOGAN: So you didn't try? Is that it?
DR.
BROWN: So what I did actually on some
of them is I did a bit of gentle scrubbing in the box joints to see if it would
come off. It wouldn't come off much by
general scrubbing. Actually, if you
reuse them, you can begin to wear off the blackening. But it's really a very superficial kind of blackening, and then
the thing with the protein adherence with the piano wire, they really did
corrode and at least the serum protein stuff we did didn't show any difference.
CHAIR
PRIOLA: Dr. Edmiston?
DR.
EDMISTON: I know it's late and I don't
want to hold anybody up, but I really want to commend Dr. Asher and Dr.
Brown. You are heading in the right
direction. The question that I have is
do you contemplate looking at this in devices that have larger bores in terms
of if you're looking at a hollow device, are you looking at other devices that
may have a larger bore where the cleaning process may be expedited, normal
cleaning process may be expedited on the basis of having a larger internal
diameter?
DR.
BROWN: These are solid pins.
DR.
EDMISTON: These are solid pins?
DR.
BROWN: These are solid pins. They were not needles.
DR.
EDMISTON: Okay.
DR.
BROWN: So the idea was that David had
been using a needle. He was used to the
feel of that size needle, and I made solid pins to match. So these were not hollow.
DR.
EDMISTON: So you don't know what would
happen with a hollow bore device?
DR.
BROWN: No. One can sort of guess, but I think --
DR.
EDMISTON: Right.
DR.
BROWN: You know, this term of, you
know, the nooks and crannies and the hiding places, I think that's the next
generation of the studies. Polymer
coated, we have got some that, apparently, are even teflon coated that are part
of the next step in the study.
DR.
ASHER: Yes. As I mentioned, years ago I did some standard hypodermic needles,
just squirting suspensions of scrapie through and letting the needles dry and
autoclave. You know, you're not
surprised to hear that they were not sterilized.
DR.
EDMISTON: I think our experiences have
been that, especially in the case of neurosurgery, that those patients who fall
into that risk category, a lot of us are moving towards the use of disposable
biopsy needles.
CHAIR
PRIOLA: Dick, did you have a
question? I'm sorry, can you what?
BOARD
MEMBER JOHNSON: Can we leave our papers
behind?
CHAIR
PRIOLA: I think you --
SECRETARY
FREAS: If you want it tomorrow morning,
I would really recommend you take it to your room. I do have a couple of quick announcements. This morning, we passed out about 200
Conflict of Interest questionnaires and we got about five of them back. I would like to encourage you to look at the
questionnaires and if you could drop them off on your way out, we'll pass out
another 100 tomorrow and, hopefully, we'll got some back.
Also,
somebody left behind a Palm Pilot. It
looks like it's a very expensive Palm Pilot, and if you can identify it, it's
yours. Tomorrow morning, we'll be
seeing you at 8:00 sharp.
CHAIR
PRIOLA: Okay. I would like to thank all of the speakers for presenting
published and unpublished data to the Committee, and we're adjourned until 8:00
a.m. Thank you.
(Whereupon,
at 6:18 p.m. the meeting was adjourned.)