Avian Hepatitis E Virus Research Articles

Hepatitis E Virus in Livestock-Update on Its Epidemiology and Risk of Infection to Humans.

Hepatitis E virus (HEV) is a public health problem worldwide and an important food pathogen known for its zoonotic potential. Increasing numbers of infection cases with human HEV are caused by the zoonotic transmission of genotypes 3 and 4, mainly by consuming contaminated, undercooked or raw porcine meat. Pigs are the main reservoir of HEV. However, it should be noted that other animal species, such as cattle, sheep, goats, and rabbits, may also be a source of infection for humans. Due to the detection of HEV RNA in the milk and tissues of cattle, the consumption of infected uncooked milk and meat or offal from these species also poses a potential risk of zoonotic HEV infections. Poultry infected by avian HEV may also develop symptomatic disease, although avian HEV is not considered a zoonotic pathogen. HEV infection has a worldwide distribution with different prevalence rates depending on the affected animal species, sampling region, or breeding system.

Inflammatory response triggered by avian hepatitis E virus in vivo and in vitro.

Hepatitis E virus (HEV) is relevant to public health worldwide, and it affects a variety of animals. Big liver and spleen disease (BLS) and hepatitis-splenomegaly syndrome (HSS) associated with avian HEV (aHEV) were first reported in 1988 and in 1991, respectively. Here, cell culture-adapted aHEV genotype 3 strain, YT-aHEV (YT strain), a typical genotype isolated in China, was used for basic and applied research. We evaluated liver injury during the early stages of infection caused by the YT strain in vivo. Both in vivo and in vitro experimental data demonstrated that viral infection induces innate immunity, with mRNA expression levels of two key inflammatory factors, interleukin-1β (IL-1β) and IL-18, significantly upregulated. The YT strain infection was associated with the activation of Toll-like receptors (TLRs), nuclear factor kappa B (NF-κB), caspase-1, and NOD-like receptors (NLRs) in the liver and primary hepatocellular carcinoma epithelial cells (LMH). Moreover, inhibiting c-Jun N-terminal kinase, extracellular signal-regulated kinase (ERK1 or 2), P38, NF-κB, or caspase-1 activity has different effects on NLRs, and there is a mutual regulatory relationship between these signaling pathways. The results show that SB 203580, U0126, and VX-765 inhibited IL-1β and IL-18 induced by the YT strain, whereas Pyrrolidinedithiocarbamate (PDTC) had no significant effect on the activity of IL-1β and IL-18. Pretreatment of cells with SP600125 had an inhibitory effect on IL-18 but not on IL-1β. The analysis of inhibition results suggests that there is a connection between Mitogen-activated protein kinase (MAPK), NF-κB, and the NLRs signaling pathways. This study explains the relationship between signaling pathway activation (TLRs, NF-κB, MAPK, and NLR-caspase-1) and viral-associated inflammation caused by YT strain infection, which will help to dynamic interaction between aHEV and host innate immunity.

Combined inhibitory effect of interferon and antiserum on avian hepatitis E virus

Epidemiologic investigations in recent years have shown that the detection rate of avian hepatitis E virus (HEV) in chicken flocks is increasing in China. Nevertheless, effective prevention and control measures are still lacking. In this study, specific pathogen-free (SPF) chicken serum against HEV was prepared using recombinant HEV open reading frames (ORF2 and ORF3) proteins as immunogens. An SPF chicken infection model was established by intravenous inoculation of chick embryos. Swab samples were collected at 7, 14, 21, and 28 d of age and used to detect avian HEV load, along with other indicators, by fluorescence quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) assay. The therapeutic effects on blocking vertical HEV transmission were observed, by using the methods of antibody application alone, mixed, or combined application of each of the 2 antibodies with type I interferon. The results showed that type I interferon alone or in combination with antiserum reduced the positive rate of HEV from 100 to 62.5% and 25%, respectively. However, the avian HEV-positivity rate was reduced to 75, 50, and 37.5% after type I interferon was used alone or in combination with antisera against ORF2 and ORF3, respectively. The inhibitory effect of type I interferon alone or in combination with an antiserum, on HEV replication was more significant in cells than in vivo. In this study, the inhibitory effect of type I interferon alone or in combination with an antiserum on avian HEV replication was observed in vitro and in vivo, providing the necessary technical reserve for disease prevention and control.

Application of ORF3 Subunit Vaccine for Avian Hepatitis E Virus.

Avian hepatitis E virus (HEV) is the main etiologic pathogen of chicken big liver and spleen disease which is widely prevalent in China in recent years. However, due to the lack of a highly effective culture system in vitro, a genetically engineered subunit vaccine is the main direction of vaccine development. In this study, ORF3 genes of VaHEV strain from laying hens and YT-aHEV strain from broilers were amplified, respectively, and ORF3 protein was successfully expressed by Escherichia coli prokaryotic expression system. The serum samples were collected periodically to detect avian HEV antibodies by indirect immunofluorescence after specific pathogen free chickens immunized with the two proteins and their mixed proteins, the results showed that all serum samples were positive for antibodies to avian HEV. The antibody-positive chickens were artificially challenged with the cell-adapted strain YT-aHEV strain. The chickens from the immunized control group were infected successfully; no fecal detoxification was detected in the immunized group. In this study, two representative strains of ORF3 subunit vaccines of laying hens and broilers were prepared by prokaryotic expression, the immune effects of different proteins of these were evaluated through immunization and challenge studies in vivo, which provided a new technical possibility for prevention and control of avian HEV.

Research Note: The prevalence and vertical transmission of avian hepatitis E virus novel genotypes in Tai'an city, China

To investigate the prevalence of avian hepatitis E virus (HEV) in chickens and gather evidence of viral vertical transmission, we collected 288 cloacal swabs and 288 yolks samples from 12 farms with clinically healthy chickens in 4 different areas in Tai'an City, Shandong Province, China (i.e., Daiyue District, Xintai City, Feicheng City, and Ningyang County). We also collected 240 samples from 2 breeder farms (from each of which 30 chicks, 30 dead embryos, 30 live embryos, and 30 hatching eggs were taken). PCR detection revealed that the positive rates of cloacal swabs and yolks were 6.25% (18/288) and 4.51% (13/288), respectively. Besides, avian HEV was detected with higher positive rates in the chicks (11.67%), hatching eggs (10.00%), live embryos (13.33%), and dead embryos (26.67%) from 2 breeder farms. Sequence and genetic evolution analyses revealed that the nucleotide homology of the isolated strains was 76.4to 83.9% compared with 4 reported genotypes, but the isolated strains were located in a separate branch, indicating they were potential novel genotypes. In conclusion, those results indicate that the latent infection of avian HEV novel genotypes has been widespread in chicken farms in Tai'an City, and provide reliable evidence of the possible vertical transmission of avian HEV.

Emergence and Molecular Characterization of an Avian Hepatitis E Virus From Donglan Black Chicken in Southern China.

Avian hepatitis E virus (HEV) is a major pathogen associated with hepatitis splenomegaly syndrome in chickens and has been reported in China. Phylogenetic trees, Bayesian analysis, positive selection sites screening, and recombination analysis were first used to comprehend the global avian HEVs. All the avian HEV strains, including a new isolate named GX20A1 got from Donglan Black chicken in Guangxi, China, were uniformly defined into four genotypes, and GX20A1, belongs to Genotype 3. The topology of the phylogenetic tree based on the sequences of a 339-bp fragment (coding the helicase) in open reading frame (ORF) 1 of the avian HEVs was consistent with that based on the full-genome sequence. The estimated evolution rate of avian HEVs is 2.73 × 10−3 substitution/site/year (95% confidence interval (CI): 8.01 × 10−4−4.91 × 10−3), and the estimated genetic diversity of the strains experienced a declining phase from 2010 to 2017 and stabilized after 2017. It was further found that the Genotype 3 HEVs, including isolates from Hungary and China, likely originated in the 1930s. Notably, GX20A1 was gathered in the same branch with a Genotype 3 Guangdong isolate CaHEV-GDSZ01, which appeared earlier than GX20A1. In addition, two positive selection sites were identified, one for each of ORF1 and ORF2. Overall, the study revealed that avian HEVs were uniformly defined into four genotypes, and a 339-bp fragment in ORF1 of the viral genome could be used for the classification. A Genotype 3 isolate GX20A1 was first found from Donglan Black chicken and most likely originated from Guangdong.

Avian Hepatitis E Virus ORF2 Protein Interacts with Rap1b to Induce Cytoskeleton Rearrangement That Facilitates Virus Internalization.

ABSTRACTAvian hepatitis E virus (HEV) causes liver diseases and multiple extrahepatic disorders in chickens. However, the mechanisms involved in avian HEV entry remain elusive. Herein, we identified the RAS-related protein 1b (Rap1b) as a potential HEV-ORF2 protein interacting candidate. Experimental infection of chickens and cells with an avian HEV isolate from China (CaHEV) led to upregulated expression and activation of Rap1b both in vivo and in vitro. By using CaHEV capsid as mimic of virion to treat cell in vitro, it appears that the interaction between the viral capsid and Rap1b promoted cell membrane recruitment of the downstream effector Rap1-interacting molecule (RIAM). In turn, RIAM further enhanced Talin-1 membrane recruitment and retention, which led to the activation of integrin α5/β1, as well as integrin-associated membrane protein kinases, including focal adhesion kinase (FAK). Meanwhile, FAK activation triggered activation of downstream signaling molecules, such as Ras-related C3 botulinum toxin substrate 1 RAC1 cell division cycle 42 (CDC42), p21-activated kinase 1 (PAK1), and LIM domain kinase 1 (LIMK1). Finally, F-actin rearrangement induced by Cofilin led to the formation of lamellipodia, filopodia, and stress fibers, contributes to plasma membrane remodeling, and might enhance CaHEV virion internalization. In conclusion, our data suggested that Rap1b activation was triggered during CaHEV infection and appeared to require interaction between CaHEV-ORF2 and Rap1b, thereby further inducing membrane recruitment of Talin-1. Membrane-bound Talin-1 then activates key Integrin-FAK-Cofilin cascades involved in modulation of actin kinetics, and finally leads to F-actin rearrangement and membrane remodeling to potentially facilitate internalization of CaHEV virions into permissive cells.IMPORTANCE Rap1b is a multifunctional protein that is responsible for cell adhesion, growth, and differentiation. The inactive form of Rap1b is phosphorylated and distributed in the cytoplasm, while active Rap1b is prenylated and loaded with GTP to the cell membrane. In this study, the activation of Rap1b was induced during the early stage of avian HEV infection under the regulation of PKA and SmgGDS. Continuously activated Rap1b recruited its effector RIAM to the membrane, thereby inducing the membrane recruitment of Talin-1 that led to the activation of membrane α5/β1 integrins. The triggering of the signaling pathway-associated Integrin α5/β1-FAK-CDC42&RAC1-PAK1-LIMK1-Cofilin culminated in F-actin polymerization and membrane remodeling that might promote avian HEV virion internalization. These findings suggested a novel mechanism that is potentially utilized by avian HEV to invade susceptible cells.

The diagnosis and molecular epidemiology investigation of avian hepatitis E in Shandong province, China

BackgroundAvian hepatitis E virus (HEV) is the pathogenic agent of big liver and spleen disease (BLS) and of hepatitis-splenomegaly syndrome (HSS) in chickens, which have caused economic losses to the poultry industry in China. In this study, 18 samples of BLS chickens were collected to reveal the molecular epidemiological characteristics of avian HEV in the province of Shandong, China.ResultsGross and microscopic lesions of clinical samples were observed; then, virology detection and genetic analysis of avian HEV were performed. The results showed that there was significant swelling and rupture in the liver and that the spleen was enlarged. Microscopic lesions demonstrated obvious hemorrhage in the liver, with infiltration of heterophilic granulocytes, lymphocytes, and macrophages, as well as the reduction of lymphocytes in the spleen. Eleven of the 18 samples were positive for avian HEV, with a positive rate of 61.11%. More importantly, all avian HEV-positive samples were mixed infections: among these, the mixed infections of avian HEV and chicken infectious anemia virus (CIAV) and avian HEV and fowl adenovirus (FAdV) were the most common. Furthermore, the genetic evolution analysis showed that all avian HEV strains obtained here did not belong to the reported 4 genotypes, thus constituting a potential novel genotype.ConclusionsThese results of this study further enrich the epidemiological data on avian HEV in Shandong, prove the genetic diversity of avian HEV in China, and uncover the complex mixed infections of avian HEV clinical samples.

Antigenic cross-reactivity among human, swine, rabbit and avian hepatitis E virus capsid proteins

Hepatitis E virus (HEV), a zoonotic virus, infects many animal species, including humans. Capsid proteins of human, swine, rabbit and avian HEVs share 48 %–100 % amino acid identity. In the present study, antigenic cross-reactivity among human, swine, rabbit and avian HEV capsid proteins were analyzed in detail using indirect and blocking enzyme-linked immunosorbent assays (ELISAs). The C-terminal 268 amino acids of genotype 1 human, genotype 4 swine, genotype 3 rabbit and genotype B3 avian HEV capsid proteins served as coating antigens for ELISA. Hyperimmune rabbit antisera (against four HEV capsid proteins) and human, pig, rabbit and chicken clinical sera were as primary antibodies. Closely correlated and statistically indistinguishable results were obtained for detection of anti-HEV antibodies in human and pig sera using human, swine and rabbit HEV capsid proteins as coating antigens. Moderately correlated differences in detection of anti-HEV antibodies in rabbit sera were obtained using the three capsid proteins. Statistically significant differences with no correlations were obtained for anti-HEV antibodies detection in chicken sera between avian HEV capsid protein and human, swine and rabbit ones. Blocking ELISA results demonstrated that two common epitopes among the four species HEVs were immunodominant in avian HEV, but were non-immunodominant in human, swine and rabbit HEVs. Nevertheless, three epitopes common to human, swine and rabbit HEVs were all immunodominant epitopes for the three species HEVs. Collectively, these results demonstrate that anti-HEV antibodies in human and pig sera can be detected using human, swine and rabbit HEV capsid proteins. By contrast, for optimal detection of anti-HEV antibodies in rabbit and chicken sera, the respective rabbit and avian HEV capsid proteins should be used. These results provide insights to guide future development of serological assays for diagnosing HEV infections in various animal species.

Isolation, identification and genome analysis of an avian hepatitis E virus from white-feathered broilers in China

Avian hepatitis E virus (HEV) is the major causative pathogen of the big liver and spleen disease, hepatitis-splenomegaly syndrome, and hepatic rupture hemorrhage syndrome. Until now, there are 6 different avian HEV genotypes that infect chickens have been reported worldwide. Epidemiologic investigations of the avian HEV demonstrated that avian HEV has been widely spread in China in recent years. In this study, an avian HEV named YT-aHEV was obtained from white-feathered broilers using LMH cells by virus isolation assay in Shandong province, China. The complete genome consists of 6656-nt excluding the poly(A) tail. The isolate was highly similar to the CaHEV strain and segregated into the same branch belonging to avian HEV genotype 3. Indirect immunofluorescence using capsid protein-specific polyclonal antibodies confirmed that YT-aHEV could establish productive infection and replicate stably in LMH cells. Furthermore, an in vivo avian HEV infection model was established successfully in specific pathogen-free chicken embryos by intravenous experiments. In the present study, we demonstrate an avian HEV infection associated with liver lesions of hemorrhage and swelling by LMH cells for the first time in a white-feather broiler flock in China. This research also provides a new diagnosis method for detection of avian HEV, which laid a foundation for the understanding of pathogenicity and molecular biology of this virus for further study.

Cell Division Control Protein 42 Interacts With Hepatitis E Virus Capsid Protein and Participates in Hepatitis E Virus Infection.

Hepatitis E Virus (HEV) causes viral hepatitis in humans worldwide, while a subset of HEV species, avian HEV, causes hepatitis-splenomegaly syndrome in chickens. To date, there are few reports on the host proteins interacting with HEV and being involved in viral infection. Previous pull-down assay combining mass spectrometry indicated that cell division control protein 42 (CDC42), a member belonging to the Rho GTPase family, was pulled down by avian HEV capsid protein. We confirmed the direct interaction between CDC42 and avian and mammalian HEV capsid proteins. The interaction can increase the amount of active guanosine triphosphate binding CDC42 state (GTP-CDC42). Subsequently, we determined that the expression and activity of CDC42 were positively correlated with HEV infection in the host cells. Using the different inhibitors of CDC42 downstream signaling pathways, we found that CDC42-MRCK (a CDC42-binding kinase)-non-myosin IIA (NMIIA) pathway is involved in naked avian and mammalian HEV infection, CDC42-associated p21-activated kinase 1 (PAK1)-NMIIA/Cofilin pathway is involved in quasi-enveloped mammalian HEV infection and CDC42-neural Wiskott-Aldrich syndrome protein-actin-polymerizing protein Arp2/3 pathway (CDC42-(N-)WASP-Arp2/3) pathway participates in naked and quasi-enveloped mammalian HEV infection. Collectively, these results demonstrated for the first time that HEV capsid protein can directly bind to CDC42, and non- and quasi-enveloped HEV use different CDC42 downstream signaling pathways to participate in viral infection. The study provided some new insights to understand the life cycle of HEV in host cells and a new target of drug design for combating HEV infection.

Research Note: A putative novel subtype of the avian hepatitis E virus of genotype 3, Jiangxi province, China

In recent years, the avian hepatitis E virus (HEV) has been widely spread in China, causing huge economic losses. Several studies have carried out detailed epidemiologic investigations of the avian HEV, but no data were from Jiangxi province. Since early April 2020, diseases similar to hepatic rupture hemorrhage syndrome caused by the avian HEV occurred in a Roman Brown layer farm in Jiangxi province, indicating this virus may also be epidemic there. To make this assumption clear, 20 liver samples were collected from the sick flock and then analyzed by detailed viral detection, which confirmed that the avian HEV should be responsible for the aforementioned disease (6 of 20). Then, the capsid gene of the virus was sequenced to show the molecular characteristics of the strain circulating in the aforementioned flock. Sequence comparison showed that it shared 80.7 to 94.7% identities with 12 published strains, while phylogenetic analysis confirmed that it belongs to a new subtype of genotype 3. Moreover, basing on a 242 bp fragment, the novel also shared high similarities to reference strains identified as genotypes before, revealing the genotype 3 maybe very popular in China and even can be divided into several subgroups. In conclusion, a novel avian HEV strain was identified in this study, which belongs to a new subtype of genotype 3. The analysis makes up for the molecular epidemiologic data of avian HEV and provides a basis for further understanding the spread of avian HEV in China.

Genetic diversity of avian hepatitis E virus in China, 2018-2019.

Avian hepatitis E virus (HEV) is highly variable and has multiple genotypes. Previous studies showed that the current epidemic strain in China is genotype 5, but the relevant detection was only carried out in flocks with hepatic rupture haemorrhage syndrome, which does not mean that other genotypes do not exist. In this study, a broader analysis involving different chicken flocks was performed to understand the epidemic status of avian HEV in China. The results showed that the HEV-positive rate of all samples was 7.92% (78/985), and four different genotypes have been identified by analysing a truncate capsid gene fragment, while the homology between them is about 80%. Two of them are separately known as genotype 3 and genotype 5, while the other two are completely unidentified, indicating that there are multiple genotypes of avian HEV prevalent in China. At the same time, the distribution of these genotypes has no obvious geographical clustering pattern, only slightly different in commercial layers, broilers and some indigenous species. This study shows the genetic diversity of avian HEV in China and reminds us to pay more attention to its variation and evolution.

Identification and pathogenicity of a novel genotype avian hepatitis E virus from silkie fowl (gallus gallus)

Hepatitis E virus (HEV) is a public health concern because of its zoonotic potential; however, the host species spectrum and the genetic diversity of HEV in many birds are unknown. In the present study, a novel genotype avian HEV was isolated from a bird, silkie fowl, and designated CHN-GS-aHEV (GenBank No. MN562265). The genome of CHN-GS-aHEV was analyzed in comparison with other avian HEVs’ and the pathogenicity in silkie fowl was characterized. The results show that the CHN-GS-aHEV shares about 81 % identity with known avian HEV in chickens, ORF3 shares the highest identity (85.1 %–88.0 %) at the nucleotide level, while ORF2 shares the highest identity (96.5 %–98.0 %) at the amino acid level, indicating that the CHN-GS-aHEV belongs to a new genotype avian HEV. The pathogenicity study showed that silkie fowl experimentally infected with the CHN-GS-aHEV demonstrated seroconversion, viremia, fecal virus shedding, liver lesions, and increased ALT level. Furthermore, ultrastructural changes in hepatocyte cells by transmission electron microscopy were characterized by the loss of mitochondrial cristae and swollen mitochondria and endoplasmic reticulum in the infected birds, suggesting that these two organelles may play a significant role in HEV replication. Overall, this study reports the complete genome characterization of a novel avian HEV and successful experimental infection in silkie fowl, and may be serving as a prominent indicator for additional avian HEV detection in other species.

Complete genome analysis of avian hepatitis E virus from chicken with hepatic rupture hemorrhage syndrome

Since 2016, severe outbreaks of hepatic rupture hemorrhage syndrome (HRHS) associated with infections of tentative novel avian hepatitis E virus (HEV) have emerged in chickens in China, causing increased mortality and decreased laying rate in adult hens and disturbing the hatching and breeding of chicks. To further identify the genotype and gain a better understanding of the genetic properties of the avian HEV responsible for that, a strain from Hebei province was isolated, purified and sequenced in this study. Results identified a novel avian HEV genotype, sharing 79.5–86.9% identities with other published avian HEV strains, and having higher identities with Orthohepevirus A HEV strains. More importantly, the new isolate contains various amino-acid substitutions in its functional proteins, including methyltransferase, helicase, RNA-dependent RNA polymerase. The data presented in this report will enhance the current understanding of the genetic diversity of the avian HEV and provide additional insight into the critical factors that determine the pathogenicity.

Co-infection with avian hepatitis E virus and avian leukosis virus subgroup J as the cause of an outbreak of hepatitis and liver hemorrhagic syndromes in a brown layer chicken flock in China

Hens of a commercial Hy-line brown layer flock in China exhibited increased mortality and decreased egg production at 47 wk of age. From 47 to 57 wk, average weekly mortality increased from 0.11 to 3.0%, and egg production decreased from 10 to 30%, with a peak mortality rate (3.0%) observed at 54 wk of age. Necropsy of 11 birds demonstrated tissue damage that included hepatitis, liver hemorrhage, rupture, and/or enlarged livers. Microscopic liver lesions exhibited hepatocytic necrosis, lymphocytic periphlebitis, and myeloid leukosis. While no bacteria were recovered from liver and spleen samples, avian hepatitis E virus (HEV) RNA was detected in all 11 tested hens by nested reverse transcription-polymerase chain reaction. Of these, subgroup J avian leukosis virus (ALV-J) proviral DNA was detected in 5 hens by PCR. Alignments of partial ORF2 gene sequences obtained here demonstrated shared identity (76 to 97%) with corresponding sequences of other known avian HEV isolates. Env sequences of ALV-J isolates obtained here shared 50.1 to 55% identity with other ALV subgroups and 91.8 to 95.5% identity with other known ALV-J isolates. Phylogenetic tree analysis of selected sequences obtained here grouped an avian HEV sequence with genotype 3 HEV and assigned an ALV-J sequence to a branch separate from known ALV-J subgroups. Immunohistochemical results confirmed the presence of avian HEV and ALV-J in livers. Therefore, these results suggest that avian HEV and ALV-J co-infection caused the outbreak of hepatitis and liver hemorrhagic syndrome observed in the layer hen flock analyzed in this study.

CD8+ lymphocytes but not B lymphocytes are required for protection against chronic hepatitis E virus infection in chickens.

Hepatitis E is an important global disease, causing outbreaks of acute hepatitis in many developing countries and sporadic cases in industrialized countries. Hepatitis E virus (HEV) infection typically causes self-limiting acute hepatitis but can also progress to chronic disease in immunocompromised individuals. The immune response necessary for the prevention of chronic infection is T cell-dependent; however, the arm of cellular immunity responsible for this protection is not currently known. To investigate the contribution of humoral immunity in control of HEV infection and prevention of chronicity, we experimentally infected 20 wild-type (WT) and 18 immunoglobulin knockout (JH-KO) chickens with a chicken strain of HEV (avian HEV). Four weeks postinfection (wpi) with avian HEV, JH-KO chickens were unable to elicit anti-HEV antibody but had statistically significantly lower liver lesion scores than the WT chickens. At 16 wpi, viral RNA in fecal material and liver, and severe liver lesions were undetectable in both groups. To determine the role of cytotoxic lymphocytes in the prevention of chronicity, we infected 20 WT and 20 cyclosporine and CD8+ antibody-treated chickens with the same strain of avian HEV. The CD8 + lymphocyte-depleted, HEV-infected chickens had higher incidences of prolonged fecal viral shedding and statistically significantly higher liver lesion scores than the untreated, HEV-infected birds at 16 wpi. The results indicate that CD8 + lymphocytes are required for viral clearance and reduction of liver lesions in HEV infection while antibodies are not necessary for viral clearance but may contribute to the development of liver lesions in acute HEV infection.

Avian Hepatitis E Virus: With the Trend of Genotypes and Host Expansion

Avian hepatitis E virus (HEV) is a single-stranded, positive-sense RNA virus with a complete genome of approximately 6.6 kb in size. To date, four major genotypes of avian HEV have been identified and classified into the Orthohepevirus B genus of the family Hepeviridae. The avian HEV associated with hepatitis-splenomegaly syndrome, big liver and spleen disease or hepatic rupture hemorrhage syndrome in chickens is genetically and antigenically related to mammalian HEV. With the increased genotypes of avian HEV identified, a broader host tropism is also notable in the epidemiological studies. Due to the lack of an efficient cell culture system, the mechanisms of avian HEV replication and pathogenesis are still poorly understood. The recent identification and characterization of animal strains of avian HEV has demonstrated the virus’ ability of cross-species infection. Although it has not yet been detected in humans, the potential threat of a zoonotic HEV capable of transmission to humans needs to be taken into consideration. This review article focuses on the current knowledge regarding avian HEV in virology, epidemiology, pathogenesis, clinical presentation, transmission, diagnosis and prevention.HIGHLIGHTS- The mechanisms of avian HEV replication and pathogenesis are still poorly understood due to the lack of an efficient cell culture system.- A broader host tropism is also notable in the epidemiological studies with the increased genotypes of avian HEV identified.- The recent identification and characterization of animal strains of avian HEV has demonstrated the virus’ ability of cross-species infection.- The potential threat of a zoonotic HEV capable of transmission to humans needs to be taken into consideration.

Chicken Organic Anion-Transporting Polypeptide 1A2, a Novel Avian Hepatitis E Virus (HEV) ORF2-Interacting Protein, Is Involved in Avian HEV Infection.

Avian hepatitis E virus (HEV) is the main causative agent of big liver and spleen disease in chickens. Due to the absence of a highly effective cell culture system, there are few reports about the interaction between avian HEV and host cells. In this study, organic anion-transporting polypeptide 1A2 (OATP1A2) from chicken liver cells was identified to interact with ap237, a truncated avian HEV capsid protein spanning amino acids 313 to 549, by a glutathione S-transferase (GST) pulldown assay. GST pulldown and indirect enzyme-linked immunosorbent assays (ELISAs) further confirmed that the extracellular domain of OATP1A2 directly binds with ap237. The expression levels of OATP1A2 in host cells are positively correlated with the amounts of ap237 attachment and virus infection. The distribution of OATP1A2 in different tissues is consistent with avian HEV infection in vivo Finally, when the functions of OATP1A2 in cells are inhibited by its substrates or an inhibitor or blocked by ap237 or anti-OATP1A2 sera, attachment to and infection of host cells by avian HEV are significantly reduced. Collectively, these results displayed for the first time that OATP1A2 interacts with the avian HEV capsid protein and can influence viral infection in host cells. The present study provides new insight to understand the process of avian HEV infection of host cells.IMPORTANCE The process of viral infection is centered around the interaction between the virus and host cells. Due to the lack of a highly effective cell culture system in vitro, there is little understanding about the interaction between avian HEV and its host cells. In this study, a total of seven host proteins were screened in chicken liver cells by a truncated avian HEV capsid protein (ap237) in which the host protein OATP1A2 interacted with ap237. Overexpression of OATP1A2 in the cells can promote ap237 adsorption as well as avian HEV adsorption and infection of the cells. When the function of OATP1A2 in cells was inhibited by substrates or inhibitors, attachment and infection by avian HEV significantly decreased. The distribution of OATP1A2 in different chicken tissues corresponded with that in tissues during avian HEV infection. This is the first finding that OATP1A2 is involved in viral infection of host cells.

Epidemiological investigation of the novel genotype avian hepatitis E virus and co-infected immunosuppressive viruses in farms with hepatic rupture haemorrhage syndrome, recently emerged in China.

Since 2016, hepatic rupture haemorrhage syndrome (HRHS) appeared in chickens of China and caused huge economic loss. To assess the infection status of the avian hepatitis E virus (HEV) and co-infected viruses, including avian leukosis virus (ALV), reticuloendotheliosis virus (REV), fowl adenovirus (FAdV), and chicken infectious anaemia virus (CIAV), in farms with HRHS, 180 liver samples were collected from 24 farms in different provinces and detected by strict molecular virology methods. Results showed that the positive rates of HEV, ALV, REV, FAdV, and CIAV were 74.44%, 20.00%, 27.78%, 31.11%, and 12.22%, respectively, whereas there are also 112 samples with co-infection, for a rate of 58%. Meanwhile, the positive rate of HEV decreased gradually with age; the lowest positive rate of ALV (5.76%) and REV (19.23%) appeared in 25-35weeks age, during which the positive rate of CIAV was the highest (19.23%); the positive rate of HEV in layers (64.00%) was lower than that of broilers (83.33%), but the positive rates of ALV (38.46%) and CIAV (15.38%) in layers were higher than that of broilers (5.88%, 9.80%); the positive rates of HEV (75.88%) and CIAV (15.60%) in parental generation (PG) were higher than that of commodity generation (CG, 64.10%, 0.00%), whereas the positive rate of ALV showed inverse relationship (PG: 14.89%; CG: 38.46%). Additionally, phylogenetic analysis showed that all the avian HEV identified this study belong to a novel genotype, and found the close relationship between the wild strains (REV and CIAV) and corresponding isolates from contaminated vaccine. The data presented in this report will enhance the current understanding of the epidemiology characteristics in farms with HRHS in China.