|
|
Dispatch
Human and Porcine Hepatitis
E Virus Strains, United Kingdom
Malcolm Banks,*
Richard Bendall,† Sylvia Grierson,* Graham Heath,* Jonathon Mitchell,‡
and Harry Dalton†
*Veterinary Laboratories Agency, New Haw, Addlestone, Surrey, United Kingdom;
†Royal Cornwall Hospital, Treliske, Truro, Cornwall, United Kingdom; and
‡Kings College Hospital, Denmark Hill, London, United Kingdom
Suggested citation
for this article:
Banks M, Bendall R, Grierson S, Heath G, Mitchell J, Dalton H. Human
and porcine hepatitis E virus strains, United Kingdom. Emerg Infect
Dis [serial on the Internet]. 2004 May [date cited]. Available
from: http://www.cdc.gov/ncidod/EID/vol10no5/03-0908.htm
We describe a case
of acquired infection of a strain of hepatitis E virus (HEV) with a
100% amino acid identity to the analogous region in strains of HEV circulating
in a United Kingdom pig herd. This case further supports the theory
that autochthonous HEV infection in industrialized countries is zoonotic.
Hepatitis E virus (HEV) is a common cause of acute hepatitis in many
developing countries (1). HEV is often waterborne and
causes an illness similar to hepatitis A. During the icteric phase, patients
may be viremic (2). In the last few years, sporadic cases
of hepatitis E have been reported in industrialized countries in the absence
of foreign travel or other known risk factors. Nucleotide sequence analysis
of some of these viruses has indicated a high degree of sequence identity
with the sequences of HEV detected in pigs (3–5) and
in pig meat purchased in retail outlets in Japan (6).
More recently, the most direct evidence of zoonotic transmission yet available
was described in a case study from Japan. In this case, members of a family
group were infected following consumption of uncooked Sika deer meat.
The virus identified from the patients was shown to be identical to that
recovered from uneaten quantities of meat from the same deer (7).
In the United Kingdom, autochthonously acquired hepatitis E has been reported
rarely and a short (98-bp) section of the virus genome has been sequenced
from one such patient (8). Serologic evidence of HEV
infection has been detected in U.K. pigs in a preliminary study (9)
In two pigs from separate farms, an HEV genome was detected by reverse
transcription–polymerase chain reaction (RT-PCR) and sequenced (strains
1-40 and 14-P354). These sequences identified the porcine strains as nearly
identical to the 98-bp U.K. human HEV sequence in HEV genotype III (9).
Further study is underway to determine the prevalence and strain diversity
of HEV infection in U.K. pigs.
The Study
In June 2000, a 58-year-old woman, who worked as a shop assistant, was
seen at a rapid access "jaundice hotline" clinic with a 5-day
history of myalgia and jaundice. She had not traveled outside the United
Kingdom for 10 years and had no contact with farm or domestic animals.
She was not a vegetarian and, although she admitted to eating raw sausage
and bacon in the past, she claimed not to have done so in the 3 months
before her illness. Examination confirmed jaundice and tenderness in
the right upper quadrant. Her liver function tests showed elevated levels
of bilirubin, 46 mmol/L; alanine aminotransferase, 2,421 IU/L; and alkaline
phosphatase, 200 IU/L. Blood drawn at this time was positive for anti-HEV
immunoglobulin (Ig) M; no serologic evidence indicated active infection
with Epstein-Barr virus, cytomegalovirus, or hepatitis A, B, or C. Results
of a liver ultrasound were normal, and serologic evidence did not indicate
autoimmune or metabolic liver disease. One month later, she felt better,
was no longer jaundiced, and results of her liver function tests were
generally normal. A diagnosis of acute hepatitis E was made.
Total RNA was extracted in Trizol (Sigma, Poole, UK) from a blood sample
drawn from the patient at the time she sought treatment. Template cDNA
from the blood extraction was prepared by a reverse transcriptase step,
according to standard protocols. A nested PCR was used to detect HEV from
the prepared cDNA samples. This assay used degenerate oligonucleotide
primers to amplify a 348-bp fragment of open reading frame 2 (ORF2) of
HEV as described by Meng et al. (1997) (10).
Cycling parameters were the same as those described by Meng et al. (10),
except for annealing temperatures of 60°C and 55°C for external and internal
primer pairs, respectively, and an extension time of 1 min. RT-PCR products
were subjected to electrophoresis on a 2% agarose gel containing ethidium
bromide and visualized under UV light.
Amplicons of correct size were excised from 2% agarose gels, purified
with QIAquick Gel Extraction Kit (Qiagen Ltd., Crawley, UK), and sequenced
with Big Dye Terminator Cycle Sequencing Ready Reaction (Applied Biosystems,
Warrington, UK). Sequences comprising 304 bp of HEV ORF2 were assembled
by SEQMAN (DNASTAR) and phylogenetic analysis was performed using PHYLIP
(ver 3.6) with NEIGHBOR. Bootstrap confidence values were calculated by
using SEQBOOT and CONSENSE (available from http://evolution.genetics.washington.edu/phylip.html).
|
Figure
1 |
|
|
 |
|
|
Click to view
enlarged image
Figure 1. Outline of dendrogram of selected
partial nucleotide sequences of ORF-2 region of swine and human
hepatitis E virus (HEV) isolates (300 nt)...
|
|
|
|
|
|
Figure
2
|
|
|
 |
|
|
Click
to view enlarged image
Figure 2. Human United Kingdom isolate (AY362357)
is shown in bold and compared with closely related swine and human
hepatitis E virus isolates... |
|
Analysis of the 304-bp nucleotide sequence obtained between the primer
sequences showed 16 and 32 nucleotide differences from the two U.K. pig
strains 1-40 and 14-P354, respectively. However, all of these differences
were silent when the gene sequence was translated into amino acids; the
putative amino acid sequence of the virus detected in the patient had
100% identity with that of the two U.K. pig strains. Based on the nucleotide
data the phylogeny of the three strains was determined (Figures
1 and 2), showing a very close relationship
between the three strains in the genotype III lineage. In Japan, high
sequence similarities have been demonstrated between human and pig HEV
strains within genotype III and genotype IV (5).
Conclusions
HEV was discovered in pigs as recently as 1997 (10),
despite evidence from several countries that it had been present since
at least the early 1990s. The lack of recognized clinical disease in pigs
(11) was undoubtedly a factor in this late discovery.
The similarity of the human strain detected in this case to the two U.K.
pig strains is suggestive of zoonotic transmission, as is the finding
of higher prevalence of anti-HEV antibodies in those who work with pigs
(12) and the observation that in 9 of 10 hepatitis E
patients in Japan, the disease developed within 2–8 weeks of consumption
of grilled or undercooked pork (6). Consideration is
being given within the U.K. Department for Environment, Food and Rural
Affairs to research proposals to gain better understanding of HEV in the
U.K. pig herd. Screening patients who present with acute hepatitis for
evidence of HEV infection is a simple way to improve the assessment of
human HEV infection. Rapid access jaundice hotline clinics (13)
are an ideal venue for such testing because patients may present while
still viremic, allowing molecular characterization of infecting viruses
as in this case. Work is currently in progress to assess the incidence
of hepatitis E in a cohort of over 650 acutely jaundiced patients who
have contacted the jaundice hotline in the past 5 years.
Dr. Banks has been
a research virologist at the Veterinary Laboratories Agency for over
20 years. He has a background in the control of animal herpesviruses,
working with the European Union and Office Internationale des Épizooties.
His current research interests lie in the pathogenesis of porcine multisystemic
wasting syndrome and the incidence and prevalence of animal hepatitis
E virus in relation to zoonoses.
References
- Labrique AB, Thomas DL, Stoszek SK, Nelson KE. Hepatitis
E: an emerging infectious disease. Epidemiol Rev 1999;21:162–79.
- Zhang JZ, Im SW, Lau SH, Chau TN, Lai ST, Ng SP. Occurrence of hepatitis
E virus IgM, low avidity IgG serum antibodies, and viremia in sporadic
cases of non-A, -B, and -C acute hepatitis. J Med Virol 2000;66:40–8.
- Wu JC, Chen CM, Chiang TY, Sheen IJ, Chen JY, Tsai WH, et al. Clinical
and epidemiological implications of swine hepatitis E virus infection.
J Med Virol 2000;60:166–71.
- Okamoto H, Takahashi M, Nishizawa T, Fukai K, Muramatsu U, Yoshikawa
A. Analysis
of the complete genome sequence of indigenous swine hepatitis E virus
isolated in Japan. Biochem Biophys Res Commun 2001;289:929–36.
- Nishizawa T, Takahashi M, Mizuo H, Miyajima H, Gotanda Y, Okamoto
H. Characterization
of Japanese swine and human hepatitis E virus isolates with 99% identity
over the entire genome. J Gen Virol 2003;84:1245–51.
- Yazaki Y, Mizuo H, Takahashi, M, Nishizawa T, Sasaki, N, Gotanda Y,
et al. Sporadic
acute or fulminant hepatitis E in Hokkaido, Japan, may be foodborne,
as suggested by the presence of hepatitis E virus in pig liver as food.
J Gen Virol 2003;84:2351–7.
- Tei S, Kitajima N, Takahashi K, Mishiro S. Zoonotic
transmission of hepatitis E virus from Sika deer to human beings.
Lancet 2003;362:371–3.
- Wang Y, Levine DF, Bendall RP, Teo CG, Harrison TJ. Partial
sequence analysis of indigenous hepatitis E virus isolated in the United
Kingdom. J Med Virol 2001;65:706–9.
- Banks M, Grierson SS, King DP, Gresham A, Girones R, Widen F, et al.
Evidence for the presence of Hepatitis E virus in pigs in the UK. Vet
Rec 2004;154:223–7.
- Meng XJ, Purcell RH, Halbur PG, Lehman JR, Webb DM, Tsareva TS, et
al. A
novel virus in swine is closely related to the human hepatitis E virus.
Proc Natl Acad Sci U S A 1997;94:9860–5.
- Meng XJ, Halbur PG, Haynes JS, Tsaeva TS, Bruna JD
Royer RL, et al. Experimental
infection of pigs with the newly identified swine hepatitis E virus
(swine HEV), but not with human strains of HEV. Arch Virol 1998;143:1405–15.
- Meng XJ, Wiseman B, Elvinger F, Guenette DK, Toth TE, Engle RE, et
al. Prevalence
of antibodies to hepatitis E virus in veterinarians working with swine
and in normal blood donors in the United States and other countries.
J Clin Microbiol 2002;40:117–22.
- Mitchell J, Hussaini H, McGovern D, Farrow R, Maskell G, Dalton H.
The
"jaundice hotline" for the rapid assessment of patients with
jaundice. BMJ 2002;325:213–5.
|
