Seroepidemiology and molecular characterization of hepatitis E virus infection in swine and occupationally exposed workers in Punjab,India

Hepatitis E virus (HEV) has two discrete epidemiological patterns: waterborne epidemics in developing countries only, caused by HEV genotype I, and sporadic zoonotic outbreaks in developing and developed countries caused by genotypes III and IV. This study was designed to investigate seroprevalence, molecular detection and the characterization of HEV by nested RT‐PCR in swine as well as the occupational risk to exposed human population in Punjab state of north‐western India. The occupational risk‐exposed group comprised of swine farmers (organized – mixed feed feeders and unorganized – swill feeders), slaughterhouse workers, sewage workers and veterinary internes. During the study period, blood and faecal samples were collected from 320 swine and 360 humans with both high and low occupational exposure risks. The overall seroprevalence of swine HEV was 65.00%, with a significantly higher seropositivity in growing pigs (2–8 months of age). The prevalence of HEV RNA in swine faecal samples by nRT‐PCR was 8.75% with a significantly higher detection in swill‐fed pigs. With humans in the high occupational exposure risk population, significantly higher anti‐HEV IgG seropositivity was observed (60.48%) as compared to control population (10.71%). Strong evidence of association between human anti‐HEV IgG seropositivity and certain occupational exposure risk groups was observed (p < 0.05). This indicates that unorganized swine farmers, slaughterhouse workers and sewage workers have higher odds of HEV infection in this study region. Percentage of nucleotide similarity between swine and human HEV isolates was less than that found in countries with zoonotic HEV outbreaks. Molecular characterization revealed the circulation of G IV and G I genotypes among swine and human population in Punjab state, respectively.     

Detection of hepatitis E virus RNA in rats caught in pig farms from Northern Italy

Hepatitis E virus (HEV) strains belonging to the Orthohepevirus genus are divided into four species (A–D). HEV strains included in the Orthohepevirus A species infect humans and several other mammals. Among them, the HEV‐3 and HEV‐4 genotypes are zoonotic and infect both humans and animals, of which, pigs and wild boar are the main reservoirs. Viruses belonging to the Orthohepevirus C species (HEV‐C) have been considered to infect rats of different species and carnivores. Recently, two studies reported the detection of HEV‐C1 (rat HEV) RNA in immunocompromised and immunocompetent patients, suggesting a possible transmission of rat HEV to humans. The role of rats and mice as reservoir of HEV and the potential zoonotic transmission is still poorly known and deserves further investigation. To this purpose, in this study, the presence of HEV RNA was investigated in the intestinal contents and liver samples from 47 Black rats (Rattus rattus) and 21 House mice (Mus musculus) captured in four pig farms in Northern Italy. The presence of both Orthohepevirus A and C was investigated by the real‐rime RT‐PCR specific for HEV‐1 to HEV‐4 genotypes of Orthohepevirus A species and by a broad spectrum hemi‐nested RT‐PCR capable of detecting different HEV species including rat HEV. The intestinal content from two Black rats resulted positive for HEV‐C1 RNA and for HEV‐3 RNA, respectively. None of the House mice was HEV RNA positive. Sequence analyses confirmed the detection of HEV‐C1, genotype G1 and HEV‐3 subtype e. The viral strain HEV‐3e detected in the rat was identical to swine HEV strains detected in the same farm. Liver samples were negative for the detection of either rat HEV or HEV‐3.     

A systematic review and meta‐analysis of predictors of human hepatitis E virus exposure in non‐endemic countries

The reported incidence of clinical hepatitis E cases is rising in some non‐endemic countries, with concurrent concerns regarding potential hepatitis E virus (HEV) contamination of the blood supply. Therefore, the characterization of major potential sources of human HEV exposure is important to inform risk assessment and public health policy. A systematic review was conducted, including a comprehensive search in six electronic bibliographic databases, verified by hand‐searching reference lists of HEV reviews, and a grey literature search, of the broad research question ‘what is the evidence of the association between predictors of human HEV exposure, and HEV IgG seropositivity, in non‐endemic countries?’ Using forms designed a priori, captured studies were appraised at first‐level screening, second‐level characterization, and third‐level data extraction and risk of bias assessment. Meta‐analysis yielded summary estimates of association between potential predictors and odds of HEV seropositivity. Meta‐analysis and meta‐regression of the odds of HEV seroprevalence in specific groups characterized potential sources of HEV exposure. From 4,163 captured citations, 245 relevant studies underwent data extraction, investigating HEV seroprevalence or predictors in both healthy subjects and targeted patient groups. Across these groups, increasing age was a predictor of HEV IgG seropositivity. Both human immunodeficiency virus patients and haemodialysis patients had significantly increased odds of HEV seropositivity relative to the general population. Working with pigs, in forestry, or in hospitals, was significantly associated with increased odds of HEV seropositivity, as were consumption of meat, pork or game meat, or hunting. Chronological time was not associated with HEV seropositivity within our data sets. Further study of the distribution of potential dietary or behavioural predictors between high and lower prevalence areas within non‐endemic countries could improve our understanding of the relative importance of specific HEV transmission pathways.     

屠宰猪肝和血清中乙型肝炎病毒及戊型肝炎病毒的检测

应用1对乙型肝炎病毒（HBV）S基因保守区的引物，采用PCR方法从屠宰猪肝、血清中检测到了HBV，序列分析表明，扩增片段与已发表的HBVS基因的同源性高达98％～100％。电镜负染色样品观察结果表明，在HBV表面抗原ELISA检测强阳性反应的血清样品中存在有形态、大小与人HBV Dane颗粒和小球状颗粒相似的病毒粒子。针对戊型肝炎病毒（HEV）（）RF2／ORF3重叠区设计了简并引物，采用巢式RT-PCR对屠宰猪肝和血清样品进行了检测。结果表明，部分屠宰猪肝中存在HEV。     

Prevalence of hepatitis E virus infection among people and swine in mainland China: A systematic review and meta‐analysis

Hepatitis E virus (HEV) infection remains an important public health problem, and it is endemic primarily in developing countries. This study aimed to evaluate the seroprevalence of HEV among the general population, occupational population and swine in mainland China and its risk factors based on a systematic review and meta‐analysis. Systematic search from EMBASE, PubMed, Web of Science, Cochrane Library and several Chinese databases, such as Wanfang (WF) Data, China National Knowledge Infrastructure (CNKI), Chinese Science and Technology Periodical Database (VIP) and SINOMED, was searched from inception up to 25 April 2018. The overall seroprevalence of HEV and its corresponding 95% confidence interval (CI) as well as the correlation coefficients between different groups were estimated using stata 12.0 and r ‐3.4.1 software. Potential sources of heterogeneity were explored using subgroup and sensitivity analyses. Twenty‐eight studies with 57,274 participants (including human and swine) were included. The seroprevalence of anti‐HEV immunoglobulin G (IgG) among the general population, occupational population and swine was 27.3% (95% CI: 22.4–32.2), 47.4% (95% CI: 40.1–54.8) and 66.4% (95% CI: 61.7–71.1), respectively. The overall prevalence of IgM among the general population was 1.8% (95% CI: 0.7–2.9). The odds ratio for the occupational population, as compared to the general population, was 2.63. The highest anti‐HEV IgG prevalence (59%) was observed in East China, whereas the lowest (34.8%) was noted in Northeast and North China. In the occupational population, the highest prevalence (77.0%) was observed among swine vendors. Seven studies included 30,392 participants (humans and swine); the correlation coefficient for the prevalence of anti‐HEV IgG between the professional population and adult pigs was 0.88. Sensitivity analyses showed that the stability of results was not considered significant. This research found that HEV is common in China, and contact with pork or other pig products may be an important mode of HEV transmission.     

Prevalence and genotyping of hepatitis E virus in swine population in Korea between 1995 and 2004: a retrospective study

Hepatitis E virus (HEV) infections have been reported in pigs throughout the world but have only recently been recorded in Korean pigs. The aim of this study was to investigate whether HEV was present in archived porcine hepatic tissues collected between 1995 and 2004 using RT-PCR and immunohistochemistry and, if so, to determine the genotype of the isolates. Swine HEV was identified in the liver tissue of 42 pigs of 388 submissions (four pigs every year on average). The isolates showed genetic homology with swine and human HEV isolates identified in the United States and Japan (92.5-97%) and phylogenetic tree analysis indicated they belonged to genotype III. The study indicates that HEV is not a newly emerging virus in Korean pigs, but a pathogen that has existed in the country since at least 1995.     

Serological evidence of hepatitis E virus infection in pigs and jaundice among pig handlers in Bangladesh

Hepatitis E virus (HEV) is the most common cause of viral hepatitis in humans. Pigs may act as a reservoir of HEV, and pig handlers were frequently identified with a higher prevalence of antibodies to HEV. The objectives of this study were to identify evidence of HEV infection in pigs and compare the history of jaundice between pig handlers and people not exposed to pigs and pork. Blood and faecal samples were collected from 100 pigs derived from three slaughterhouses in the Gazipur district of Bangladesh from January to June, 2011. We also interviewed 200 pig handlers and 250 non‐exposed people who did not eat pork or handled pigs in the past 2 years. We tested the pig sera for HEV‐specific antibodies using a competitive ELISA and pig faecal samples for HEV RNA using real‐time RT‐PCR. Of 100 pig sera, 82% (n = 82) had detectable antibody against HEV. Of the 200 pig handlers, 28% (56/200) demonstrated jaundice within the past 2 years, whereas only 17% (43/250) of controls had a history of jaundice (p < .05). Compared to non‐exposed people, those who slaughtered pigs (31% versus 15%, p < .001), reared pigs (37% versus 20%, p < .001), butchered pigs (35% versus 19%, p < .001) or involved in pork transportation (28% versus 13%, p < .001) were more likely to be affected with jaundice in the preceding 2 years. In multivariate logistic regression analysis, exposure to pigs (odds ratio [OR]: 2.2, 95% CI: 1.2–3.9) and age (OR: 0.97, 95% CI: 0.95–0.99) was significantly associated with jaundice in the past 2 years. Pigs in Bangladesh demonstrated evidence of HEV infection, and a history of jaundice was significantly more frequent in pig handlers. Identifying and genotyping HEV in pigs and pig handlers may provide further evidence of the pig's role in zoonotic HEV transmission in Bangladesh.     

Comparative pathogenesis in specific-pathogen-free chickens of two strains of avian hepatitis E virus recovered from a chicken with Hepatitis–Splenomegaly syndrome and from a clinically healthy chicken

Avian hepatitis E virus (avian HEV) is the primary causative agent of Hepatitis–Splenomegaly (HS) syndrome in chickens. Recently, a genetically unique strain of avian HEV, designated avian HEV-VA, was recovered from healthy chickens in Virginia. The objective of this study was to experimentally compare the pathogenicity of the prototype strain recovered from a chicken with HS syndrome and the avian HEV-VA strain in specific-pathogen-free chickens. An infectious stock of the avian HEV-VA strain was first generated and its infectivity titer determined in chickens. For the comparative pathogenesis study, 54 chickens of 6-week-old were assigned to 3 groups of 18 chickens each. The group 1 chickens were each intravenously inoculated with 5 × 10^2.5 50% chicken infectious dose of the prototype strain. The group 2 received the same dose of the avian HEV-VA strain, and the group 3 served as negative controls. Six chickens from each group were necropsied at 2, 3 and 4 weeks post-inoculation (wpi). Most chickens in both inoculated groups seroconverted by 3 wpi, and the mean anti-avian HEV antibody titers were higher for the prototype strain group than the avian HEV-VA strain group. There was no significant difference in the patterns of viremia and fecal virus shedding. Blood analyte profiles did not differ between treatment groups except for serum creatine phosphokinase levels which were higher for prototype avian HEV group than avian HEV-VA group. The hepatic lesion score was higher for the prototype strain group than the other two groups. The results indicateded that the avian HEV-VA strain is only slightly attenuated compared to the prototype strain, suggesting that the full spectrum of HS syndrome is likely associated with other co-factors.     

The study of seroprevalence of hepatitis E virus and an investigation into the lifestyle behaviours of the aborigines in Malaysia

Malaysia is a non‐endemic country for hepatitis E virus (HEV) infection. However, seroprevalence as high as 50% among samples of aboriginal people were reported over two decades ago. A total of 207 samples collected from seven aboriginal villages in rural settlements across two states in Malaysia were analysed for anti‐HEV IgG and IgM by an enzyme‐linked immunoassay. Following the detection of anti‐HEV seroprevalence, we organized health outreach to inform and educate the community. Qualitative interviews were conducted with individuals tested positive for anti‐HEV antibodies. Data derived from interviews and observations were used to investigate possible lifestyle behaviours associated with HEV infection. Anti‐HEV IgG was detected in six samples (5.9%) from the village of Dusun Kubur. Qualitative inquiry and observation study revealed poor dietary and household hygiene, contaminated food and water, contact with animal faeces, unsanitary and domestic waste disposal, and wildlife reservoirs could be the contributing factors for transmission and acquisition of HEV infection. Investigation during health outreach is important to provide insights for future empirical research and implementation for improvement of lifestyle behaviours among the aborigines. Managing the risk of HEV infection in the aborigines may reduce the risk of HEV transmission to the local communities.     

东北边境地区野猪及放养杂交野猪戊型肝炎流行病学调查

为了解东北边境地区野猪及放养杂交野猪群体猪戊型肝炎病毒(HEV)感染情况,于2015—2018年在吉林省、黑龙江省的中朝、中俄边境和内蒙古自治区境内加格达奇周边地区采集6月龄以上杂交野猪血清、粪便或肛拭子样品共520份,采集野猪血清和粪便样品共248份。ELISA检测、RT-nPCR检测、全基因组测序、同源性及进化分析结果显示,杂交野猪和野猪感染HEV的血清抗体总阳性率为34.1%(136/399);核酸总阳性率为1.56%(12/771),12份核酸阳性样品均来自杂交野猪,病毒基因组ORF2部分核苷酸序列同源性为85.4%～100.0%,属于基因4型,4a、4b亚型。对4a亚型的1份阳性样品(LJG-18)进行病毒全基因组扩增测序,其核苷酸序列与日本的人源毒株JKO-ChiSai98C同源性最高,为94.9%,与吉林省猪源毒株Ch-S-1同源性为90.2%。结果表明:东北边境地区放养杂交野猪群具有较高的HEV血清抗体阳性率,HEV流行毒株以4a亚型为主。本试验针对我国野猪及放养杂交野猪群体开展猪戊型肝炎流行病学调查,为该病的流行情况提供了新的科学数据,对我国养猪业健康发展和公共卫生安全具有重要意义。     

A Cross‐Sectional Study of Hepatitis E Virus Infection in Healthy People Directly Exposed and Unexposed to Pigs in a Rural Community in Northern Thailand

A cross‐sectional study of the association between occupational pig exposure and hepatitis E virus (HEV) infection in adult pig farmers and the general population who were not directly exposed to pigs was conducted in Nan Province, Thailand, from November 2010 to April 2011. All participants were interviewed to provide information on their job history, eating habits and other potential confounders. The prevalence of anti‐HEV immunoglobulin G antibodies (IgG) among 513 subjects was 23.0%. Hand washing with water and soap was associated with a lower seroprevalence of HEV infection, whereas living in an area with frequent flooding (OR 1.64, 95% CI: 1.00–2.68) and consuming internal pig organs more than twice per week (OR 3.23, 95%CI: 1.15–9.01) were both associated with a higher seroprevalence of anti‐HEV IgG. There was no association between HEV seroprevalence and frequent, direct occupational pig contact.     

High hepatitis E virus antibody positive rates in dogs and humans exposed to dogs in the south‐west of China

Hepatitis E (HE) is a zoonotic viral disease caused by hepatitis E virus (HEV). The objective of this study was to investigate the prevalence of HEV infection among dogs and humans exposed to dogs in the south‐west region of China. A total of 4,490 dog serum samples and 2,206 relative practitioner serum samples were collected from 18 pet hospitals and dog farms in Yunnan, Sichuan and Guizhou province, and the anti‐HEV IgG antibodies were detected by ELISA. The results showed that the total positive rate of anti‐HEV antibodies was 36.55% with the highest rate in city stray dogs, and the differences in distinct species and growth phases were significant. The positive rate of anti‐HEV antibody in veterinarian and farm staff‐related practitioners was significantly higher than the general population. The finding of the present survey suggested that high HEV seroprevalence in dogs and humans exposed to dogs in the south‐west area of China poses a significant public health concern. It is urgent to improve integrated strategies to detect, prevent and control HEV infection in dogs and humans exposed to dogs in this area.