Zhibiao Yang

Hepatitis E Virus Infection among Domestic Animals in Eastern China

Hepatitis E virus (HEV) is a zoonotic pathogen of which several species of animal were reported as reservoirs. Antibodies to HEV and HEV RNA have been detected in some Chinese population and swine groups but few other domestic animals. In this study, to investigate the HEV prevalence, we tested sera from 788 pigs, 100 cows, 50 goats, 49 horses, 101 pet dogs, 105 chickens, 47 duck and 45 pigeons in eastern China for anti‐HEV immunoglobulin G (IgG). We also tested 50% of the swine sera, all of sera from the other domestic animals and 13 Shanghai human sera which were positive for anti‐HEV immunoglobulin M (IgM) for HEV RNA using reverse transcriptase‐polymerase chain reaction. Our results indicated that 82.5% (222/269) of the sows, 53.9% (104/193) of the 4‐ to 6‐month‐old swine, 63.4% (168/265) of the 1‐ to 3‐month‐old swine, 55.7% (34/61) of the slaughterhouse swine, 24% (12/50) of the goats, 16.3% (8/49) of the horses, 17.8% (21/101) of the pet dogs, 6% (6/100) of the cows, 12.8% (6/47) of the ducks, 4.4% (2/45) of the pigeons and 1.9% (2/105) of the chickens exhibited positive for anti‐HEV IgG. Inhibition assay confirmed the infection with HEV or HEV‐like viruses in these domestic animals except pigeons and chickens. From the sera, we isolated 18 swine HEV strains, one horse HEV strain and two human HEV strains. Sequence analysis showed that the horse HEV isolate and one swine isolate belonged to genotype 3. The other isolates belonged to genotype 4. The two human isolates were phylogenetically closely related to eight of the swine isolates. In short, the presence of anti‐HEV antibody had been confirmed in several species of domestic animals in eastern China and HEV RNA has been identified in swine, human and horse. This suggested that the authorities should pay more attention to the prevalence of HEV in eastern China.

Experimental infection of Balb/c nude mice with Hepatitis E virus.

BackgroundSeveral animal species can reportedly act as reservoirs for Hepatitis E virus (HEV), a zoonotic pathogen. HEV and antibody to the virus have been detected in a variety of animals including rodents. Pig and rat models for HEV have been established for HEV, but a nude mouse has not yet been developed.MethodsBalb/c nude mice were inoculated with swine HEV, both orally and via intravenous injection to insure infection. Negative control and experimental contact-exposed groups of mice were also included in the study. The liver, spleen, kidney, jejunum, ileum, cecum and colon of each mouse from all three groups were collected for reverse transcription nested polymerase chain reaction (RT-nPCR) detection, indirect immunofluorescence observation and histopathologic examination. The sera from nude mice were tested for anti-HEV IgG by enzyme linked immunosorbent assay (ELISA). Activities of liver enzymes, including alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), as well as total bilirubin (TBIL) were also measured in the sera of the nude mice.ResultsHEV antigens and HEV RNA were detected in liver, spleen, kidney, jejunum, ileum and colon both by indirect immunofluorescence and by RT-nPCR in all of the inoculated and in one of the contact-exposed nude mice. Histopathological changes were observed in the liver and spleen of these mice. Infected mice showed increased levels of AST, ALP, and anti-HEV IgG in sera. The livers of contact-exposed mice showed obvious histopathological damage.ConclusionNude mice could be readily infected by HEV isolated from pigs. The nude mouse may therefore be a useful animal model for studying the pathogenesis of HEV.

Molecular characterization of a porcine astrovirus strain in China

Pigs are increasingly recognized to harbor a wide range of viruses that apparently establish long-term persistence in these animals. They serve as reservoirs for a number of human zoonotic diseases. In this study, a porcine astrovirus (PAstV) strain, designated as PAstV JWH-1, is identified from a diarrheal pig in China, and it is partially characterized genetically. Sequence analysis shows that the PAstV JWH-1 strain contains divergent nucleotide sequences in both the open reading frame (ORF)1b/ORF2 consensus and the 3′-UTR regions (s2m motif), which are usually highly conserved among members of the family Astroviridae. Phylogenetic analysis indicates that the JWH-1 strain clusters closely with newly identified strains PAstV 12-4 and 14-4 and forms a group of mamastroviruses with the proposed novel deer astrovirus. Further recombination analysis shows that two possible interspecies recombination events between porcine and deer astroviruses occurred in the genome of the JWH-1 strain. This study further confirms that multiple lineages are present among PAstVs, and each lineage likely represents an independent origin. Additionally, the possibility of interspecies transmission among PAstVs is also suggested.

Hepatitis E Virus Infection in Central China Reveals No Evidence of Cross-Species Transmission between Human and Swine in This Area

Hepatitis E virus (HEV) is a zoonotic pathogen of which several species of animal were reported as reservoirs. Swine stands out as the major reservoir for HEV infection in humans, as suggested by the close genetic relationship of swine and human virus. Since 2000, Genotype 4 HEV has become the dominant cause of hepatitis E disease in China. Recent reports showed that genotype 4 HEV is freely transmitted between humans and swine in eastern and southern China. However, the infection status of HEV in human and swine populations in central China is still unclear. This study was conducted in a rural area of central China, where there are many commercial swine farms. A total of 1476 serum and 554 fecal specimens were collected from the general human and swine populations in this area, respectively. The seroepidemiological study was conducted by enzyme-linked immunosorbent assay. Conserved genomic sequences of open reading frame 2 were detected using reverse transcription-PCR. The results indicated that the overall viral burden of the general human subjects was 0.95% (14/1476), while 7.0% (39/554) of the swine excreted HEV in stool. The positive rate of anti-HEV IgG and IgM in the serum samples was 7.9% (117/1476) and 1.6% (24/1476), respectively. Phylogenetic analysis based on the 150 nt partial sequence of the capsid protein gene showed that the 53 swine and human HEV isolates in the current study all belonged to genotype 4, clustering into three major groups. However, the HEV isolates prevalent in the human and swine populations were classified into known distinct subgenotypes, which suggested that no cross-species transmission between swine and humans had taken place in this area. This result was confirmed by cloning and phylogenetic analysis of the complete capsid protein gene sequence of three representative HEV strains in the three major groups. The cross reactivity between anti-HEV IgG from human sera and the two representative strains from swine in central China was confirmed by Dot-blot assay. In conclusion, although all the HEV strains prevalent in central China belonged to genotype 4, there is no evidence of cross-species transmission between human and swine in this area.

Isolation and characterization of the first Chinese porcine sapelovirus strain

Investigations were carried out to identify the causative agent of acute diarrhea, respiratory distress, and polioencephalomyelitis of pigs on a swine farm in Shanghai, China. Samples from the affected animals were tested for viruses and bacteria that are known to cause similar symptoms in swine, and only porcine sapelovirus (PSV; designated as csh strain) was isolated. The presence of PSV was further confirmed by the specific cytopathic effects observed in susceptible cells and by the results of PCR and electron microscopy. Nucleotide sequencing and phylogenetic analysis showed that this isolate is PSV. When inoculated into healthy pigs, PSV.csh caused the same symptoms as observed in the affected herd. Therefore, PSV.csh is the causative agent of this disease. To the best of our knowledge, this is the first report of PSV infecting piglets in China.

Prevalence of antibody to hepatitis E virus among pet dogs in the Jiang-Zhe area of China

In order to investigate the prevalence of HEV infection and immunity in pet dogs in the Jiang-Zhe area of China, we examined 192 dog serum samples collected from 11 animal hospitals in this area for anti-HEV IgG and HEV RNA, using ELISA and RT-PCR assays, respectively. Two dogs were injected intravenously with swine HEV to obtain anti-HEV antibody positive serum for ELISA assay. After the ELISA assay, 26 samples showing OD values higher than 0.20 were selected and examined by Dot-blot assay, and 88.5% (23/26) were confirmed to be positive for anti-HEV IgG. Although our results indicated swine HEV could experimentally induce anti-HEV IgG in the serum, we failed to detect HEV RNA in the serum samples from either the 2 intravenously injected dogs or 192 serum samples collected from animal hospitals. This implied that dog may not be a reservoir of HEV in this area.

Inhibition of porcine transmissible gastroenteritis virus (TGEV) replication in mini-pigs by shRNA

Abstract Transmissible gastroenteritis virus (TGEV) is the causative agent of porcine transmissible gastroenteritis (TGE), characterized by high mortality and severely retarded growth in piglets that dramatically affects the porcine industry. Previously, we have identified two shRNA-expressing plasmids pEGFP-U6/P1 and pEGFP-U6/P2 that target RNA-dependent RNA polymerase (RdRP) gene of TGEV with more than 95% of virus inhibition in vitro. In this study, inhibition of the TGEV replication by pEGFP-U6/P1 and pEGFP-U6/P2 was tested in mini-pigs. SPF mini-pigs at 25 days old were injected with the shRNA-expressing plasmids and then infected with TGEV. The results from the analyses of clinical signs, histopathology, indirect immunofluorescence (IIF) and RT-PCR show that the two shRNA-expressing plasmids could significantly decrease the quantity of TGEV in different organs and protect mini-pigs from TGEV infection. These findings illustrate the prospect for TGEV-specific shRNAs to be new anti-TGEV agents.

Haemotrophic mycoplasma: review of aetiology and prevalence

Haemoplasma, composed of Eperythrozoon and Haemobartonella, is a widespread agent infecting different vertebrate animals as well as humans. It is a small, pleomorphic bacterium that is an obligative parasite of erythrocytes in which it causes deformity and haemolysis. Clinical symptoms include anaemia, icterus, acidosis and fever. The prevalence of haemoplasma infection has been demonstrated in several countries. Some possible routes of transmission have been suggested, but all require further validation. Host-specific cross-transmission has been shown by sequence alignment analysis.

Serological evidence of West Nile virus in dogs and cats in China

We evaluated West Nile virus (WNV) seroprevalence in dogs and cats in Shanghai, China. Seventeen of the 367 dogs (4.6%) and 46 of the 309 cats (14.9%) tested positive for WNV antibodies. A higher WNV seroprevalence was found with outdoor and rural pets than with indoor and urban pets. However, WNV seroprevalence between the sexes were not significantly different. The results indicate that WNV-positive serum antibodies are present in dogs and cats in China, and pets, especially strays, could be served as effective sentinels for WNV surveillance.

Isolation and characterization of a genotype 4 Hepatitis E virus strain from an infant in China

In the present study, a genotype 4 HEV strain was identified in the fecal specimen from a seven months old infant with no symptom of hepatitis in Shanghai Childrens hospital. The full capsid protein gene (ORF2) sequence of this strain was determined by RT-PCR method. Sequence analysis based on the full ORF2 sequence indicated that this HEV strain shared the highest sequence identity (97.6%) with another human HEV strain isolated from a Japanese patient who was infected by genotype 4 HEV during traveling in Shanghai. Phylogenetic analysis showed that this genotype 4 HEV was phylogenetically far from the genotype 4 HEV strain that was commonly prevalent in Shanghai swine group, suggesting that this strain may not come from swine group and not involved in zoonotic transmission in this area.