|
 Environmental sampling is
an important tool for determining the presence
of Bacillus anthracis spores
in indoor environments. Sampling can help assess
the extent and degree of contamination and the
risk of exposure to building occupants and responders.
Sample results also contribute to informed decisions
on medical treatment and decontamination options
and are ultimately used to determine the effectiveness
of decontamination.
| General Sampling Plan |
 The purpose of a sampling
plan is to create a road map to achieving defined
sampling objectives. An effective sampling
plan will provide confidence that the results
obtained are valid and indicative of the contamination
present. The sampling plan should address each
of the following questions:
In evaluating methods that
will achieve the sampling objectives, several other
factors should be considered:
- Laboratory capability (such as the
laboratory's ability to handle the proposed
sampling media),
- Collection efficiency of the method,
number of samples, and need for quantification,
- Suitability of the sample-collection
and analytical methods,
- Cost effectiveness and efficiency of
the sampling plan in meeting stated objectives,
and
- Utility of the sampling method to the
owner and/or federal agency having jurisdiction
over the project.
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| Specific Sampling Objectives |
There are a variety of
potential sampling objectives. The specific sampling
objectives that may be applicable for an anthrax
response may include:
- Real-time Monitoring: Determine,
in real-time, whether a release of spores is
occurring or has occurred in a facility. Real-time
instruments may be used to detect biological
agents as they are released.
- Preliminary Assessment
of a Facility (Screening): Determine
qualitatively whether any spores are present. Typically,
composite samples of large areas and air
volumes are obtained to maximize the likelihood
of finding contamination.
- Identification of Spores
in a Bulk Material: Determine
qualitatively if a bulk material, such as
a powder in an envelope, is contaminated
with anthrax. On-site analysis may be
used for preliminary assessment, but laboratory
analysis provides confirmation.
- Determination of Contamination
of an Article: Determine whether
the surface of a small article is contaminated. Typically,
composite surface samples of large articles
and/or individual samples of small articles
are collected.
- Extent and Location of
Contamination (Site Characterization): After
anthrax is positively identified, further
sampling is necessary to determine how far
the contamination has spread. Sampling
is performed to determine qualitatively,
and if possible, semi-quantitatively, the
extent and magnitude of contamination.
- Effectiveness of Decontamination
(Process Verification Sampling): Determine
whether decontamination has reduced spores
to a safe level.
- Post-Decontamination Sampling
(Re-occupancy Verification Sampling): Final
post-decontamination sampling is conducted
inside and outside of the exclusion zone
to verify that the originally contaminated
environment has been sufficiently decontaminated
to allow re-occupancy of the area without
the use of personal
protective equipment (PPE).
- Special Re-occupancy Considerations: There
may be site-specific circumstances where additional
sampling should be considered.
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| Sampling Approach |
After the sampling objectives
have been identified, a logical approach and
schedule must be developed to carry out the sampling
tasks. The approach and schedule will depend
on the objectives.
For Assessment/Characterization:
- Targeted Sampling Strategy: If
a building or area becomes contaminated from
a known source and the source is quickly isolated,
the sampling approach will center on the source
area and move outward away from the source
to define the extent of contamination.
- Statistically-Based Strategy: If
contamination is likely to be present in a
building or area and the source has not been
identified, sampling will focus on areas that
are statistically the most likely to be contaminated. The
objective of statistical sampling is to maximize
the probability of detecting contamination.
- Air Movement and HVAC
Considerations: Spore-bearing particles
less than 10 microns in size, or spores themselves,
may remain suspended in the air for long
periods of time (such as hours or days).
In such cases, spores may spread throughout
an air space and into adjacent areas on localized
air currents, such as those created by people
walking by, and also through generalized
airflow created by HVAC systems. To
fully assess the extent of contamination,
the investigator should extend beyond the
targeted approach to sample areas on projected
contaminant pathways, such as those associated
with air movement, dust collection, or work
process flow.
For Verification Sampling
Prior to Occupancy:
- Post-Decontamination Surface
Sampling: The effectiveness of
decontamination should be confirmed by post-decontamination
environmental sampling in ambient air; and
in areas and on surfaces that were previously
contaminated. Verification sampling
prior to occupancy should include surfaces
and air in areas outside of the exclusion
zone (for example, the area where decontamination
activities were conducted) to ensure that
the outside areas continue to remain free
of spores.
- Air Sampling: Aggressive
air sampling techniques have been developed
for recent anthrax responses that model EPA
guidance for clearing facilities for re-occupancy
after asbestos decontamination. While
the area is under negative pressure, all surfaces
are aggressively agitated and air is continuously
disturbed while samples are collected. An
air sampling method that maximizes the likelihood
of detecting contamination should be used.
Sampling Quality Assurance
and Quality Control (QA/QC) Considerations:
QA/QC measures must be incorporated into the sampling
approach to insure the legitimacy of sampling results. The
sampling objectives will dictate the rigor of the
QA/QC program for a given site or task. QA/QC
includes four key elements:
- QC of field activities
- Sample documentation and management
- Sample handling and shipment
- Data validation and management
|
| Sampling
Techniques |
There are various sampling
techniques available for determining the presence
of anthrax in air and surface samples.
 The following techniques are available and have
proven useful for surface
and bulk sampling:
- Wet wipe,
- Wet swab,
- High-volume vacuum with "HEPA" sock
(Alsock), and
- Bulk sampling.
 The following techniques are available and have
proven useful for air
sampling:
- Gelatin filter (low volume),
- Mixed cellulose ester (MCE) filter,
- Anderson air sampler and single-stage
impactors with seetle plates,
- Open agar plate,
- Dry filter unit (high-volume air sampler
with polyester 1-micron filter), and
- Liquid impingers.
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| Analytical
Methods |
The following analytical
methods are available for the detection of Bacillus
anthracis:
- Immunoassay Tests: Hand-held
assays, sometimes referred to as "smart
tickets," are sold commercially for the
rapid detection of Bacillus
anthracis and other biological agents. These
assays are intended only for screening environmental
samples. Due to their low and questionable
sensitivity, they cannot be relied on for a
determination that anthrax is or is not present.
- Polymerase Chain Reaction
(PCR): PCR is a technique that amplifies
DNA and compares sequences to known test
probe standards for Bacillus
anthracis. PCR can be used in
the field or the laboratory; in either case,
the samples must be cultured to confirm that
the bacteria is Bacillus
anthracis and that it is viable. Field
PCR systems are very selective, but do not
work well with heterogeneous environmental
samples, and the probes are very expensive. PCR
has been shown to work best as a final confirmation
of positive samples taken from plated colonies.
-
 Culturing: Samples
may be analyzed for Bacillus
anthracis using a traditional
lab culturing technique. The sample
is appropriately prepared for elution
and plating, after which it is inoculated
onto plates containing sheep blood agar. The
plates are allowed to incubate for several
hours and are then examined for growth
of suspicious colonies.
The laboratory selected for the sample analyses
should be consulted by the team developing the
sampling plan. The lab should be authorized for
work with Bacillus anthracis and
laboratory procedures must conform to guidance
provided by CDC or the Association of Public Health
Laboratories (APHL). The lab should also be a part
of the Laboratory Response Network (LRN), which
was developed by CDC, APHL, and the FBI to help
public health labs prepare for and respond to acts
of terrorism, including bioterrorism.
The following links provide additional information
regarding analytical methods:
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| Packaging and Transportation of Samples |
 There are strict requirements
for packaging and transporting anthrax samples
to ensure that the general public and workers
transporting the samples are protected from exposure. These
requirements include:
- Rigorous packaging designed to withstand
rough handling and prevent leakage,
- Appropriate marking and labeling that
identifies the contents of the package,
- Documentation of the hazardous contents
of the package and emergency point-of-contact,
and
- Training of transportation workers
on how to handle the contents in the
event of an emergency.
Packaging and transporting
anthrax samples are subject to various regulations
established by the DOT, CDC, USPS, OSHA, and the
International Air Transport Association (IATA). It
is also important to consult with the analytical
laboratory receiving the samples to determine whether
they have additional packaging or shipping requirements.
Additional details on packaging and shipping procedures
for anthrax samples, according to CDC guidelines,
can be found at:
- Packing Critical Biological Agents. Centers for Disease Control and Prevention,
Public Health Emergency Preparedness and Response. Protocols for
packing, shipping, and transport of biological agents/diseases.
- Interstate
Shipment of Etiologic Agents. Centers for Disease Control and Prevention,
Office of Health and Safety, Biosafety Branch.
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| Interpretation of Results |
 Since analytical methods
are not fully validated for Bacillus
anthracis, the following considerations
are recommended for interpretation of analytical
results:
- Use a multi-disciplinary team of technical
experts to interpret the analytical results.
- Consult with field investigators and
laboratory personnel during the interpretation
process. This will provide the best
insight into sample collection and recovery.
- Consider the analytical method limitations
and use professional judgment in interpreting
any positive or negative findings as
well as quantitative or semi-quantitative
results.
- In the case of preliminary assessment
sampling, positive findings will usually
require more extensive sampling in that
particular area.
- Laboratories will sometimes report "colony
counts" with positive results. In some
cases, colony counts can help the data interpretation
teams decide how much more extensively to sample
in that area, prior to establishing additional
isolation and starting decontamination. However,
quantitative methods have not been validated
for a variety of sample collection methods
and the use of "colony count" data
is not yet accepted as a reliable indicator
of large populations of colony forming units.
- The multi-disciplinary team described above
should establish a decision tree for evaluating
the adequacy of the sampling and decontamination
to coordinate its efforts in reaching a unified
conclusion.
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| Coordination
with Affected Parties |
Before, during, and after
a response, the sampling team should coordinate
with the following affected parties:
- Facility manager,
- Union officials, and
- Other individuals who might have site-specific
knowledge of the contamination.
The sampling plan should
indicate the following:
- Employees working within the affected sampling
area need to be informed of the sampling objectives
and methods.
- The method of providing results to employees
working within the affected sampling area.
- Before sampling begins, employees should be told when, where, and why sampling may occur and they should also be advised when plans are changed.
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