Received: 13 September 2016
Revised: 13 September 2017
Accepted: 10 October 2017
DOI: 10.1002/rmv.1961
REVIEW
An overview: Rabbit hepatitis E virus (HEV) and rabbit providing an animal model for HEV study Lin Wang
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Lin Liu
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Ling Wang
Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China Correspondence Ling Wang, Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, 38 Xueyuan Road, Haidian District, Beijing 100191, China. Email:
[email protected];
Summary Hepatitis E virus (HEV) is a single‐stranded, positive‐sense RNA virus and the causative agent of hepatitis E. The virus belongs to genus Orthohepevirus in the family Hepeviridae, which contains 4 major genotypes closely relating to humans. Genotypes 1 and 2 only infect humans whereas genotypes 3 and 4 HEV are harbored in a wide range of animal species worldwide and are zoonotic to humans. Recently, a novel animal strain of HEV has been isolated in farmed rabbits in China, and subsequently more strains were discovered in the rabbit populations in at least 7 other countries. Due to high sequence similarity to genotype 3 HEV, rabbit HEV (rHEV) has been assigned to genotype 3. Experimental study showed that rHEV could infect non‐human primate
Funding information National Science Foundation of China, Grant/ Award Number: 81772175; Beijing Natural Science Foundation, Grant/Award Number: 7162103
and human, which pose a direct threat to human. Further pathogenesis studies showed laboratory rabbits infected with rHEV and genotype 4 HEV could present similar signs of acute and chronic hepatitis E along with extra‐hepatic replication as observed in humans. High mortality and vertical transmission were reproduced in rHEV infected pregnant rabbits. Furthermore, rabbit model was also found suitable for evaluating HEV vaccine efficacy in order to manage zoonotic transmission. These data showed laboratory rabbits could serve as an alternative animal model for HEV study under the current circumstances that HEV propagation is limited in vitro. In general, this review aims at presenting comprehensive up‐to‐date information about rHEV strains and rabbit model for HEV studies. KEY W ORDS
animal model, hepatitis E virus, rabbit
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and HIV/AIDS patients.8 This observation, along with extra‐hepatic
I N T RO D U CT I O N
manifestations in the context of pre‐existing HEV infection including Hepatitis E virus (HEV), the causative agent of epidemic or sporadic
neurological disorders,9,10 musculoskeletal,11,12 and hematological
hepatitis E, has an estimated 1% to 3% case fatality rate worldwide.1
manifestations,13 has aroused attention globally.
However, the mortality is up to 20% in pregnant women infected with
HEV is a non‐enveloped virus with positive‐sense, single‐stranded
HEV.2-4 HEV is generally transmitted via fecal‐oral route.5 It was a
RNA that belongs to genus Orthohepevirus in the family Hepeviridae. To
well‐accepted consensus that hepatitis E is an acute self‐limiting dis-
date, at least 7 major genotypes of HEV within species Orthohepevirus
ease; however, HEV‐associated chronic infections and cirrhosis were
A were recognized.14 Genotypes 1 and 2 (HEV‐1 and HEV‐2) are
recently reported in immunocompromised patients, such as recipients
restricted to humans and primarily dominated in Africa and Asia
of solid‐organ transplants,6 cancer patients receiving chemotherapy,7
causing large epidemics. Water contaminated with patients' excretion is the main medium facilitating their transmission. Genotypes 3 and 4 HEV (HEV‐3 and HEV‐4) are zoonotic with numerous animal hosts
Abbreviations used: HEV, Hepatitis E virus; HEV‐1, genotypes 1 HEV; rHEV, rabbit HEV; sHEV, swine HEV; HVR, hypervariable region; SPF, specific‐ pathogen‐free; wpi, week post‐inoculation; ALT, alanine aminotransferase; i.v., intravenously; GE, genome equivalent; AST, aspartate transaminase; IHC, immunohistochemistry
Rev Med Virol. 2018;28:e1961. https://doi.org/10.1002/rmv.1961
other than humans, including pig,15 wild boar,16 deer,17 and yak.18 The consumption of raw or undercooked contaminated meat is the main source of infection. Transfusion‐associated infection of HEV‐3 has also been reported recently.19
wileyonlinelibrary.com/journal/rmv
Copyright © 2017 John Wiley & Sons, Ltd.
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The first strain of rHEV was isolated in the farmed rabbits in Gansu 20
ET AL.
widespread rHEV circulation in local rabbit population.21 In France,
21
rHEV has been detected in the farmed rabbits, with a positive rate at
France,22 Italy23,24 Germany,25,26 the Netherlands,27 Korea,28 and
7.0%, lower than the wild rabbits at 23.0%.22 A single case of detecting
province, China, in 2009. 29
Then, researchers in the United States,
also discovered evidence of rHEV infections in different
rHEV in a pet rabbit's liver was reported in Turin, Italy.23 The isolated
rabbit populations. According to the International Committee on the
rHEV strain from the liver sample was found closely related to the
Taxonomy of Viruses Hepeviridae Study Group, the rHEV is
human sequence (TLS‐18516‐human, GenBank No: JQ013793)
temporarily assigned to HEV‐3 based on that the intermediate nucleo-
isolated from a French patient. It was the first report of detecting rHEV
tide and amino acid distances were between those observed within
in Italy and in a pet rabbit.23 A recent serological survey conducted in
HEV‐1, HEV‐3, and HEV‐4, and distances between genotypes.14 Sub-
Italy has found an anti‐HEV antibody seroprevalence of 3.40% in 206
sequent studies have shown that rHEV can transmit across species
farmed rabbits and 6.56% in 122 pets, suggesting that HEV is
barrier to infect SPF pigs30 and non‐human primate,31 thus suggesting
circulating in rabbits in Italy.24 A retrospective study in Germany tested
Canada
32
In a very
13 serum samples of wild rabbits hunted in 1989 around the city of
recent study, researchers in France have found the direct evidence of
Greifswald. Four out of 13 (31%) samples were positive for anti‐HEV
rHEV infection in human.33 As rabbit, like with pig, is becoming a note-
antibodies and a rHEV strain (7.7%) was isolated. The sequence has a
worthy reservoir of HEV, rHEV gains increasing attention regarding its
close relationship to rHEV sequences from France, which indicates a
epidemiology and pathogenesis. This review aims to provide a compre-
long‐existing circulation of rHEV in Europe.24 Another study conducted
hensive up‐to‐date knowledge of rHEV and rabbits for HEV study.
in Germany found that 2.2% (14/624) of the European brown hares and
the potential risk of zoonotic transmission to humans.
37.3% (47/126) of the wild rabbits were tested positive for anti‐HEV. No HEV RNA was detected in European brown hares but 17.1%
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(28/164) of the wild rabbits were HEV RNA‐positive.26 Researchers
PREVALENCE OF RHEV
from the Netherlands, Korea, and Canada have reported detecting Rabbit HEV was first isolated in 2009 in Gansu province, China. Anti‐
HEV RNA in the local rabbit populations, with prevalence ranging
HEV and HEV RNA were detected in the serum samples of farmed
from 0.9% to 60%. The overall prevalence of HEV in rabbits
rabbits, and the positive rates were 57.0% (191/335) and 6.9%
worldwide is summarized in Table 1.
20
Soon afterwards, another study reported a
Different prevalence of anti‐HEV and HEV RNA in rabbits was
different strain of rHEV in the fecal samples of Rex Rabbits in Beijing.
observed in studies around the world (Table 1). In China, rabbits in
(23/335), respectively.
The detection rates of anti‐HEV and fecal HEV RNA were 54.62%
Inner Mongolia have the highest prevalence of anti‐HEV IgG (57%)
(65/119) and 6.96% (8/115), respectively. The detection of HEV
and viraemia (72%).36 In the US study, rabbits in Farm A had a higher
RNA in fecal samples indicated the possibility of fecal‐oral transmis-
prevalence of HEV RNA in serum and fecal samples (48.0% and 40%)
sion of rHEV from rabbit to rabbit or to other species of animals.34
than in farm B (3.3% and 5.0%).21 The reason remains unknown though
A large‐scale investigation involved 10 counties in China, and a total
plausible explanation may be the age of rabbits, sampling scale, and
number of 1094 serum samples were collected from various breeds
hygienic conditions as well as housing practices. The rabbits in Inner
of rabbits. In all samples, 169 (15.4%) were found positive for anti‐
Mongolia were caged in groups of 2 to 336 but 2 to 9 in Farm A in
HEV antibody and 22 (2.01%) were positive for HEV RNA. The
Virginia.21 The recent study involving 3 regions of China showed an
35
estimated infection rate varied from 3.03% to 53.40%.
Another
overall positive rate of fecal HEV RNA at 1.0%, and all rabbits were
survey conducted in Inner Mongolia tested 211 farmed 4‐month‐old
caged individually, which, in a way reduced the possibility of fecal oral
rabbits with positive rates of anti‐HEV IgG and HEV RNA at 57.3%
transmission between cage mates.37
(121/211) and 71.6% (151/211).
36
In a recent study, which involved
So far, isolations of rHEV strains have been reported in 8
17 mammal species and 24 avian species in several regions of China,
countries, and the virus could induce infection in multiple breeds of
111 farmed rabbits in Beijing were tested for fecal RNA with a
rabbit (Table 1), which suggests that rHEV is widely spread in the world
positive rate of 4.6% (5/111). Fecal samples of 285 farmed rabbits
and rabbit will become the second largest reservoir of HEV next to pig.
in Shandong province and of 96 farmed rabbits in Henan province were also tested, but no HEV RNA was detected.37 Besides the farmed and wild rabbits, recent reports have found both anti‐HEV 38,39
and HEV RNA in SPF rabbits.
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GENETIC VARIABILITY OF RHEV
The seroprevalence for HEV was
7.5% (25/332) and 72.5% (58/80) in the SPF rabbits in Beijing and
To investigate the genetic variability of rHEV, 52 complete genomes
Shaanxi, respectively. HEV RNA was also detected in the SPF rabbits
including 17 rHEV strains were retrieved from GenBank and analyzed
of both cities, with the positive rates of 4.8% (16/332) in Beijing and
phylogenetically. By using MEGA 6.0 software package (version 6.0,
46.15% (24/52) in Shaanxi. These studies demonstrated that, in
www.megasoftware.net), a phylogenetic tree was constructed using
China, rabbit is an important reservoir of HEV second only to pig.
reference sequences of the 7 major genotypes of HEV strains in
The presence of rHEV has been further confirmed by studies on
species Othorhepevirus A and a novel genotype of HEV (most likely
farmed and wild rabbits in the United States and France. The
the HEV‐8) strains isolated from Bactrian camels in Xinjiang, China.40
prevalence of anti‐HEV antibody in rabbits is high in USA, with
The 17 rHEV full‐length genomes clustered into an independent clade
36.5% (31/85) positive rate and the detection of rHEV RNA in serum
distinct from other known genotypes (Figure 1) but share a long
(16.5%,
branch with HEV‐3. The percent nucleotide sequence identity was
14/85)
and
fecal
(15.3%,
13/85)
samples indicates
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TABLE 1
The prevalence of rabbit HEV antibodies and RNA worldwide
Country
Region
Year
Rabbit Species
Sample
Anti‐HEV (%)
HEV RNA (%)
China
Gansu Beijing Jilin Beijing Hebei Shanxi Hubei Zhejiang Guangxi Inner Mongolia Beijing Shaanxi
200920 201034
Serum Serum and feces
201236 201638 201739
Rex Rex Rex New Zealand white Rex Japanese white, chinchilla Rex Rex Rex Rex Laboratory Laboratory
Serum Serum and feces Serum and feces
57.30 54.62 9.42 42.1 7.3 7.7 53.4 10.7 3.03 57.30a 7.5 72.5
7.50 6.96 0 1.9 1.3 1.9 11.6 0 0 71.60 4.8 46.15
Virginia
201121
Multiple
Serum and feces
36.50a
22.0
Farmed, wild
Bile (farmed) Liver (wild)
NA
7 (farmed) 23 (wild)
The United States
201135
22
Serum
France
Western
2012
Italy
Turinb
201523
Pet
Serum, liver
NA
NA
Multiple
201624
Farmed, pet
Serum, feces
3.40 (farmed) 6.56 (pet)
0
Greifswaldc
201525
Wild
Serum
31
7.7
Multiple
201726
European Brown hare, wild rabbit
Serum and liver
2.2 (hares) 37.3 (wild)
0 (hares) 17.1 (wild)
The Netherlands
Multiple
201627
Pet, farmed and wild
Feces, liver
NA
23 (pet) 60 (wild)
Korea
Multiple
201728
Farmed
Feces
NA
6.4
Canada
Ontario
201729
Companion, commercial
Feces
NA
5 (companion) 0.9 (commercial)
Germany
a
The anti‐HEV tested here was anti‐HEV IgG.
b
A single case report with only one rabbit tested.
c
The study published in 2015 with the tested samples collected in 1989.
Abbreviation: NA, not applicable.
determined using the BioEdit v 7.0.9 (Carlsbad, CA, USA). The
There was no evidence of natural cross‐species transmission between
complete
68.6.1%–73.4%,
rabbit and pigs.41,42 To clarify the possibility of natural cross‐species
68.5.9%–72.8%, 71.9%–78.6%, and 70.0%–74.4% similarity with
transmission of sHEV to rabbits, pigs with HEV‐4 infection were
HEV genotypes 1 to 4, respectively. The rHEV strains were distantly
farmed along with SPF rabbits in the same enclosed space. Five of
related to HEV genotypes 1 to 8 with sequence identity less than
10 rabbits had seroconversion for anti‐HEV antibody from the third
80.0% but much closer to HEV‐3 (Table 2).
week after mix breeding. However, HEV RNA was undetectable in
sequence
of
rHEV
strains
shared
The full‐length genomes of rHEV also contain 3 ORFs, and
feces, serum, liver, and bile of the 10 rabbits.43 This study suggests that
interestingly, in the X domain of the ORF1 in all rHEV strains isolated
rabbits naturally exposed to sHEV might only lead to an inapparent
worldwide, a 31‐amino acid insertion was found, which was not present
infection in SPF rabbits by fecal‐oral route.
in any known strains of HEV genotypes 1 to 4.32 The function of this
Researchers in the United States successfully induced HEV
observed insertion in rHEV strains may indicate a possible difference
infection in SPF pigs,30 using a feces‐derived US rHEV strain
between rHEV and HEV‐3, which needs further investigation.
(GenBank No: JN383986) and a serum‐derived Chinese rHEV strain.20 An additional rat HEV strain and a sHEV‐3 strain were also used in the study. Each HEV strain was inoculated into four 6‐week‐old
4 C RO S S‐ S P E C I E S TR A N S M I S S I O N O F RHEV |
SPF pigs. In the US rHEV strain group, fecal shedding was observed in only 1/4 pigs at 5 weeks post‐inoculation (wpi) and detection of transient viraemia in 2/4 pigs. In pigs inoculated with the Chinese
More genetically diverse HEV strains have been isolated from multiple
rHEV strain, 2/4 had fecal shedding at 6 wpi without viraemia
animal species, and it seems that the host range of HEV is still
detected. No pigs showed any signs of infection in the group of rat
expanding. Therefore, it is crucial to understand whether an animal
HEV. The positive control group, in which the pigs were inoculated
strain of HEV is transmissible to other animal species, especially
with swine HEV‐3, showed fecal shedding at 1 wpi and viraemia in
humans. The discovery of rHEV aroused concern, and a series of
the next week. Anti‐HEV IgG was only detected in pigs inoculated
studies were conducted to determine the possibility of rHEV in
with sHEV‐3. This study showed that rHEV could cross the species
cross‐species transmission.
barrier to infect pigs, which raised concern of transmission of rHEV
In natural settings, no rHEV was found in pig populations so far, even in the areas where pig and rabbit farms were located closely.
to pig and then to humans, although pig was less susceptible to rHEV than sHEV.
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FIGURE 1
Phylogenetic tree of full‐length genome of hepatitis E virus of species Othohepevirus A and D. Phylogenetic tree was constructed by the neighbor‐joining method and evaluated by the maximum composite likelihood method with the aid of the MEGA 6.0 software package (www.megasoftware.net). One thousand re‐samplings of the data were used to calculate percentages of the branches obtained. The sequences signed with the black diamonds are rabbit HEV strains In another study, 2 cynomolgus macaques were inoculated with a
rHEV strain (CHN‐BJ‐R14, GenBank No: JX109834) which was
no direct contact with rabbits. This research showed direct evidence that rHEV can transmit to humans.
isolated previously from the farmed rabbits in Beijing suburb.31 All
Taking all these findings into account, rHEV has been proved to
monkeys showed fecal shedding and viraemia at 5 to 10 wpi, and
cause infections in other species of animals, including a non‐human
elevation of serum alanine aminotransferase (ALT) was observed.
primate and human patients, therefore posing a direct threat to human
During the acute phase, the monkeys manifested jaundice, malaise,
beings. Close attention should be given in the future.
and other symptoms that occur in hepatitis E. The partial sequences of the PCR products from the two monkeys' feces shared 99% to 100% nucleotide identity with the original inocula. Anti‐HEV IgM and
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PATHOGENESIS OF RHEV
IgG seroconverted at 6 to 7 wpi and IgM level dropped down when more durable IgG levels appeared, which coincides with the natural
In the first pathogenesis study, 42 SPF rabbits were randomly divided
infection history that appears in humans. This experimental infection
into 11 groups with 1 group serving as negative control.44 Several
of rHEV in the non‐human primate provides strong evidence that rHEV
strains of rHEV (GenBank No: FJ906895, FJ906888, FJ906890,
may infect humans.
FJ906896,
FJ906893)
isolated
from
Gansu
were
inoculated
A human strain of HEV has been isolated in France (TLS‐
intravenously (i.v.) into these SPF rabbits. The inocula were diluted
18516‐human, accession number: JQ013793), which shared 80.3%
from 101‐107 genome equivalents (GE). Rabbits became infected after
22
to 85.0% identity to 3 rHEV strains isolated from China and France.
HEV inoculation, fecal shedding could be detected at 1 to 2 wpi, and
A 93‐nt insertion in the X domain of ORF1 was found in this human
viraemia occurred at 4 wpi. The fecal and serum HEV RNA remained
strain. The finding reinforces the zoonotic risk of rHEV but lacks
detectable in some of the rabbits by the end of this study at 14 wpi.
detailed epidemiological data of this patient. Very recently, rHEV infec-
Elevated ALT levels were also observed during the late period of fecal
tion in human has been confirmed in France.33 By analyzing 919 HEV
shedding. The value peaked at 9 to 11 wpi with a 4‐fold elevation
strains obtained from patients in France infected during 2015 to 2016,
from baseline level. Liver histology was detected for pathological
5 rHEV strains (0.5%) were isolated from 5 patients. One infected
signs of HEV infection. Multifocal lymphohistiocytic infiltrates and
patient was immunocompetent, and 4 were immunocompromised. All
local hepatocellular necrosis were observed. All rabbits inoculated
immunocompromised patients developed chronic HEV infection. How-
with non‐passaged rHEV strains seroconverted by 3 months post‐
ever, the source of infection was unclear because all patients reported
inoculation. Rabbits are susceptible to rHEV strains, and the severity
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TABLE 2
Nucleotide sequence identities of complete genome among HEV strains from rabbits and other HEV strains Nucleotide Sequence Identity (%)
rHEV Isolate (Region)
HEV‐1
HEV‐2
HEV‐3
HEV‐4
HEV‐5
HEV‐6
HEV‐7
HEV‐8
FJ906896 GDC46 (Gansu, China)
71.5–72.5
70.9
76.2–77.5
72.1–73.0
71.4
71.6–71.8
72.8
73.1–73.2
FJ906895 GDC9 (Gansu, China)
71.5–72.4
70.9
76.5–77.6
72.1–73.3
71.6
71.5–72.2
73.1–73.2
73.5–73.6
GU937805 ch‐bj‐n1 (Beijing, China)
72.3–73.2
71.6
76.8–78.4
72.4–73.9
72.5
72.1–72.4
73.7–74.0
73.4–73.8
JQ768461 CHN‐BJ‐rb14 (Beijing, China)
71.7–72.5
71.1
76.4–77.9
72.4–73.6
72.7
72.1–72.6
73.7–74.1
73.7–74.2
JX109834 CHN‐BJ‐R14 (Beijing, China)
71.7–72.6
71.1
76.4–77.9
72.4–73.4
72.5
72.0–72.5
73.7–74.1
73.3–73.8
JX121233a CHN‐BJ‐R14 (Beijing, China)
72.5–73.3
71.9
77.5–78.4
72.8–73.9
72.7
72.3–72.6
73.9–74.3
73.6–74.0
KJ013414b CHN‐BJ‐r14(8) (Beijing, China)
68.7–69.8
68.6
72.0–73.3
70.0–70.8
69.7
69.7–69.8
70.7
70.5–70.9
KJ013415b CHN‐BJ‐r14(9) (Beijing, China)
68.6–69.7
68.5
71.9–73.1
70.0–70.7
69.5
69.6–69.8
70.6–70.7
70.5–70.9
JX565469 CMC‐1 (Virginia, USA)
72.1–73.3
71.7
77.0–78.6
72.5–73.9
72.5
72.3–72.4
73.6–73.8
73.3–73.7
AB740222 rbIM004 (Inner Mongolia, China)
71.9–72.6
71.6
76.8–78.1
72.3–73.5
72.4
71.6–72.4
73.1–73.2
73.1–73.3
AB740221 rbIM022 (Inner Mongolia, China)
72.5–73.4
71.8
77.1–78.2
72.7–74.2
72.5
72.3–72.4
73.9–74.3
73.6–73.9
AB740220 rbIM199 (Inner Mongolia, China)
72.1–73.1
72.0
77.1–78.1
72.5–73.4
72.1
72.2–72.4
73.2–73.7
73.4–73.8
JQ013791 W1‐11 (France)
72.6–73.2
72.8
77.3–77.9
73.0–74.4
71.8
71.7–72.0
73.5–73.7
73.5–73.7
JQ013792 W7‐57 (France)
72.5–72.8
72.8
76.6–77.0
72.4–73.7
72.0
71.3–71.4
73.5
JQ013793c TLS‐18156 (France)
72.2–73.0
72.7
77.1–78.0
72.4–73.6
72.1
71.8–72.3
3.5–73.9
73.3 73.5–73.6
KX227751 CHN‐SX‐rHEV (Shaanxi, China)
71.9–72.8
71.6
76.6–78.3
72.8–73.7
72.5
72.5
73.5–74.3
73.6–73.7
KY496200 KOR‐Rb‐1 (Korea)
71.7–72.7
71.2
77.0–78.3
72.3~73.7
72.1
72.3~72.9
73.8
73.6~73.9
a
Strain isolated from an experimentally infected Cynolmolgus monkey.
b
Strains isolated from two experimentally infected rabbits at 34 and 35 weeks post‐inoculation.
c
Strain isolated from a hepatitis E patient.
Reference sequences are Bur82 (M73218), Uigh179(D11093), Yam67(AF459438), Sar55(M80581), Morocco(AY230202), Abb‐2B (AF185822), FHF (X98292), Mexican (M74506), US1(AF060668), swJ570(AB073912), SwineHEV(AF082843), JBOAR‐1Hyo04 (AB189070), JRA1 (AP003430), Osh‐205 (AF455784), swArkell (AY115488), T1(AJ272108), CHN‐XJ‐SW13(GU119961), bjsw1(GU206559), HE‐JA1(AB097812), HE‐JA2(AB220974), IND‐SW‐ 00‐01(AY723745), swGX40(EU676172), swGX32(EU366959), swCH25 (AY594199), hb‐3(GU361892), CHN‐XJ‐SW33(GU119960), swCH31(DQ450072), JBOAR135‐Shiz09 (AB573435), wbJOY_06 (AB602441), wbJNN_13 (AB856243), DcHEV‐178C (KJ496143), DcHEV‐180C (KJ496144), BcHEV‐12XJ (KX387865), BcHEV‐48XJ (KX387866), and BcHEV‐62XJ (KX387867).
of the disease in rabbits is dose dependent. Another study45
into 2 naïve SPF rabbits, and rabbits inoculated with 106 copies of
reproduced the findings of the previous one. The rabbits inoculated
rHEV subsequently developed persistent infection of HEV. One rabbit
i.v. with rHEV strains (GenBank No: JQ065065, JQ065068) showed
showed a 40‐week infection and the liver tissue of this rabbit
similar manifestations of acute hepatitis E. In addition to i.v.
presented with sign of portal fibrosis and chronic inflammatory cell
inoculation, the authors also administrated the rHEV strains orally in
infiltration. These studies suggested that the development of chronic
15 rabbits, but the infectivity was low with only 2 rabbits showing
HEV infection is also dose dependent and demonstrated that rabbits
fecal shedding and seroconversion. The pathogenesis of rHEV in
may be a desirable animal model for chronic HEV infection, although
rabbits is similar to acute HEV infection observed in human, with
many specific details that are crucial to the chronicity should be
fecal shedding of HEV RNA, viraemia, seroconversion, overt
investigated thoroughly in the future, especially the factors associated
histopathological changes, and elevated ALT levels, although an i.v.
with the host immune status.
administration with high dose of virus is required. These data suggest that rabbit could be used as a model of acute HEV infection.
Another important issue about the human HEV infection is the pathogenesis during pregnancy. An up to 20% mortality rate has been
Interestingly, another study found that rabbits inoculated i.v.
reported in HEV infected pregnant women, and the underlying
with rHEV isolate CHN‐BJ‐rb14 showed a 9‐month fecal shedding
mechanism is still unclear.2-4 Researchers used the rHEV isolate
of virus RNA.46 Liver histopathology results showed chronic
CHN‐BJ‐R14 to investigate its pathogenesis and effects in pregnant
inflammatory cell infiltrations and obvious portal fibrosis, which indi-
rabbits.49 HEV infection was successfully established in 6 pregnant
cated the chronicity of HEV infection in rabbits. The prolonged
rabbits. Two of 6 infected animals miscarried, and 3 of the rest died.
viraemia and fecal shedding in rHEV infected rabbits corresponded
All rabbits had fecal shedding of virus from 3 days post‐inoculation,
to human chronic HEV infection. All rabbits seroconverted at
with persistent or transient viraemia. Elevations of ALT and aspartate
approximately 5 wpi and the high antibody level maintained till the
transaminase (AST) were observed. The seroconversion to anti‐HEV
end of the study, except one rabbit seroconcerted at 22 to 25 wpi
antibodies occurred at 3 to 7 wpi. Both positive and negative strands
and became undetectable thereafter. The same observation has been
of HEV RNA were detected in the placental tissues of the infected
47
and it may be related to the compromised
rabbits and positive staining for HEV antigen was observed in placental
host immune status. In a further investigation,48 inocula with 104,
tissue by immunohistochemistry (IHC) staining. Furthermore, vertical
105, and 106 copies of CHN‐BJ‐rb14 were, respectively, injected i.v.
transmission was discovered in the offspring with the newborns
reported in human, too,
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seroconverting at 3 months of age, which suggested that the anti‐HEV
an optimal strategy for large‐scale vaccination of rabbits. Another
antibodies were more likely induced by infection rather than passively
group vaccinated the rabbits with a different candidate vaccine, HEV
acquired from their infected mothers. The study for the first time
p179 (Changchun Institute of Biological Products Co. Ltd, Chinese
reproduced the severe outcome in pregnancy‐associated human HEV
National Biotech Corporation, Changchun, China) with 3 doses of
infection in an animal model, including high mortality rate, miscarriage,
20 μg each. The vaccinated rabbits produced anti‐HEV which
and vertical transmission.
completely protected the rabbits against 105 GE HEV‐4 infections.45
Extra‐hepatic replication has also been observed in rabbits
These studies provide the idea that rabbit may serve as an animal
infected with rHEV. Positive/negative HEV RNA and HEV ORF2
model for vaccine evaluation not only for evaluating the vaccine effi-
antigen have been detected in rabbits' brain, heart, lung, stomach,
cacy for human, but also for investigating the management of zoonotic
intestine, kidney, and placenta.31,46,49 In a recent study, rabbits
transmission. Besides, the rabbits may also have potential to be an
chronically infected with rHEV presented with kidney injury.48
appropriate model for testing antiviral agents.
Inflammatory changes in kidney tissues were observed along with
So far, the published studies regarding rabbit model for HEV
the detection of positive/negative HEV RNA and the HEV ORF3
study showed promising results that rabbit could be an optimal alter-
proteins in the kidneys, indicating the lesions seen were induced by
native for HEV infection study.53 However, recent surveys showed
HEV replication in the organ. HEV RNA was also detected in the urine
that the SPF rabbits were naturally infected with rHEV.38,39,54 Thus,
and subsequent injection of this urine sample to naïve rabbits has
a strict screening for HEV in SPF rabbits before relevant studies is
48
induced successful infection.
On the basis of these findings, the high
warranted.
pathogenecity of rHEV in SPF rabbits is further confirmed.
7 6
|
ANIMAL MODEL FOR HEV STUDY
As a novel animal model of HEV infection, the susceptibility of rabbits to multiple HEV genotypes is of great importance. To date, studies have shown that rabbit is susceptible to rHEV and HEV‐4 strains, although rHEV has a more severe pathogenecity in rabbits than HEV‐4.44,46 Only one study tried to infect rabbits with HEV‐1 but was not successful. HEV RNA was not detected in the 9 rabbits inoculated with HEV‐1; however, 6 of them seroconverted to anti‐ HEV IgG.44 Although rHEV has been assigned to HEV‐3,14 experimental infection of rabbits with human strains of HEV‐3 was not successful.45,50 This observation indicates that rHEV may have different biological characteristics compared to swine or human HEV‐3. More strains of human or swine HEV‐3 should be studied in future experiments in order to clarify the susceptibility of HEV‐3 in rabbits and this will also provide more insight into the relationship of rHEV and HEV‐3. As mentioned earlier, the previous pathogenesis studies suggest that rabbit can serve as a useful small animal model for the pathogenesis study of acute and chronic hepatitis E. Moreover, in the infected SPF rabbits, extra‐hepatic replication of HEV was detected in brain, heart, lung, kidney, small intestine, spleen, and 31,46,49
placenta.
Many extra‐hepatic manifestations have been
reported in HEV‐infected patients, and the rabbit model may allow us to study the potential mechanism of HEV‐associated extra‐hepatic
|
CO NC LUSIO N
It has been 8 years since the first discovery of rHEV in rabbits, and the further employment of the SPF rabbits in HEV infection studies has made great progress. There are still several aspects of this virus that should be investigated more deeply. More surveys should be conducted to understand the global distribution and possible genetic variability of rHEV. Meanwhile, evidence from both bench and bedside studies of rHEV showed capability of cross‐species infection in humans. The risk of zoonotic transmission of rHEV to human should not be neglected and surveillance on rHEV‐associated hepatitis E in humans must be enhanced. Rabbit is the natural host of rHEV and is susceptible to human HEV with similar pathogenesis observed in human HEV infection. Hence, given the conditions that the cell culture system of HEV is limited, rabbit model may have more potential in studying the pathogenesis of human HEV infection. ACKNOWLEDGEMENTS We thank Prof. Youchun Wang from Division of HIV/AIDS and Sex‐transmitted Virus Vaccines, National Institutes for Food and Drug Control, Beijing, China, for giving critical advice and Prof. Paul Griffiths for proofreading the manuscript. This work was supported by the National Science Foundation of China [grant number 81772175] and the Beijing Natural Science Foundation (grant no. 7162103).
manifestations. Rabbit has also been proved suitable for vaccine evaluation. A former study showed the HEV 239 vaccine could confer protection
CONFLIC T OF IN TE RE ST All authors declared no potential conflict of interest.
on rabbits against rHEV and HEV‐4 infection.51 However, the vaccine dosage used in the study is designed for humans, and it is costly for large‐scale immunization in animals. Therefore, another study was performed to explore a more cost‐effective immunization strategy to protect rabbits against HEV infection,52 and the results demonstrated that two 10‐μg doses could not only protect rabbits against rHEV and HEV‐4 infection, but also to be more economical and thus provide
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How to cite this article: Wang L, Liu L, Wang L. An overview: Rabbit hepatitis E virus (HEV) and rabbit providing an animal model for HEV study. Rev Med Virol. 2018;28:e1961. https:// doi.org/10.1002/rmv.1961
