Specific-pathogen-free pigs were inoculated with one of two hepatitis E viruses (HEV) (one recovered from a pig and the other from a human) to study the relative pathogenesis of the two viruses in swine. Fifty-four pigs were randomly assigned to three groups. Seventeen pigs in group 1 served as uninoculated controls, 18 pigs in group 2 were intravenously inoculated with the swine HEV recovered from a pig in the United States, and 19 pigs in group 3 were intravenously inoculated with the US-2 strain of human HEV recovered from a hepatitis patient in the United States. Two to four pigs from each group were necropsied at 3, 7, 14, 20, 27, or 55 days postinoculation (DPI). Evidence of clinical disease or elevation of liver enzymes or bilirubin was not found in pigs from any of the three groups. Enlarged hepatic and mesenteric lymph nodes were observed in both HEV-inoculated groups. Multifocal lymphoplasmacytic hepatitis was observed in 9 of 17, 15 of 18, and 16 of 19 pigs in groups 1 to 3, respectively. Focal hepatocellular necrosis was observed in 5 of 17, 10 of 18, and 13 of 19 pigs in groups 1 to 3, respectively. Hepatitis lesions were very mild in group 1 pigs, mild to moderate in group 2 pigs, and moderate to severe in group 3 pigs. Hepatic inflammation and hepatocellular necrosis peaked in severity at 20 DPI and were still moderately severe at 55 DPI in the group inoculated with human HEV. Hepatitis lesions were absent or nearly resolved by 55 DPI in the swine-HEV-inoculated pigs. All HEVinoculated pigs seroconverted to anti-HEV immunoglobulin G. HEV RNA was detected by reverse transcriptase PCR in feces, liver tissue, and bile of pigs in both HEV-inoculated groups from 3 to 27 DPI. Based on evaluation of microscopic lesions, the US-2 strain of human HEV induced more severe and persistent hepatic lesions in pigs than did swine HEV. Pig livers or cells from the livers of HEV-infected pigs may represent a risk for transmission of HEV from pigs to human xenograft recipients. Since HEV was shed in the feces of infected pigs, exposure to feces from infected pigs represents a risk for transmission of HEV, and pigs should be considered a reservoir for HEV.Hepatitis E virus (HEV) is the leading cause of enterically transmitted non-A, non-B hepatitis in people in many developing countries (21,28,30). Transmission is thought to be primarily by the fecal-oral route, and waterborne epidemics are characteristic of hepatitis E (1, 28, 30). Clinical disease due to HEV infection is rarely diagnosed in industrialized countries, and most cases of HEV infection in industrialized countries occur in people who have traveled to regions where the disease is endemic (10,21,28,30). Clinical cases occur predominantly in developing countries in Asia, Africa, and Mexico (1, 2, 28, 30). However, sporadic cases of acute hepatitis E in people in the United States and other industrialized countries have recently been reported (7,8,11,18,20,22,31,32,42). Hepatitis E generally affects young adults and usually is not fatal, although mortality r...

Hepatitis E virus (HEV) is the major cause of enterically transmitted non-A, non-B hepatitis in many developing countries and is also endemic in many industrialized countries. Due to the lack of an effective cell culture system and a practical animal model, the mechanisms of HEV pathogenesis and replication are poorly understood. Our recent identification of swine HEV from pigs affords us an opportunity to systematically study HEV replication and pathogenesis in a swine model. In an early study, we experimentally infected specificpathogen-free pigs with two strains of HEV: swine HEV and the US-2 strain of human HEV. Eighteen pigs (group 1) were inoculated intravenously with swine HEV, 19 pigs (group 2) were inoculated with the US-2 strain of human HEV, and 17 pigs (group 3) were used as uninoculated controls. The clinical and pathological findings have been previously reported. In this expanded study, we aim to identify the potential extrahepatic sites of HEV replication using the swine model. Two pigs from each group were necropsied at 3, 7, 14, 20, 27, and 55 days postinoculation (DPI). Thirteen different types of tissues and organs were collected from each necropsied animal. Reverse transcriptase PCR (RT-PCR) was used to detect the presence of positive-strand HEV RNA in each tissue collected during necropsy at different DPI. A negative-strand-specific RT-PCR was standardized and used to detect the replicative, negative strand of HEV RNA from tissues that tested positive for the positive-strand RNA. As expected, positive-strand HEV RNA was detected in almost every type of tissue at some time point during the viremic period between 3 and 27 DPI. Positive-strand HEV RNA was still detectable in some tissues in the absence of serum HEV RNA from both swine HEV-and human HEVinoculated pigs. However, replicative, negative-strand HEV RNA was detected primarily in the small intestines, lymph nodes, colons, and livers. Our results indicate that HEV replicates in tissues other than the liver. The data from this study may have important implications for HEV pathogenesis, xenotransplantation, and the development of an in vitro cell culture system for HEV.Hepatitis E virus (HEV), the causative agent of hepatitis E, has been recognized as a major cause of enterically transmitted non-A, non-B hepatitis in many developing countries (1,30,37,39). Transmission of the virus occurs primarily by the fecal-oral route through contaminated drinking water in areas with poor sanitation. The disease affects mainly young adults, with a reported mortality rate of up to 25% for pregnant women (10, 14, 37, 39). In the United States, sporadic cases of acute hepatitis E without known risk factors have been documented and anti-HEV antibodies have been detected in a significant proportion of healthy individuals (21,28,32,37,39,45). HEV is a positive, single-stranded RNA virus without an envelope. The genome, which is about 7.5 kb in size, contains three open reading frames (ORFs) and a short 5Ј and 3Ј nontranslated region (13,18,31,38). ORF 1 is...

Swine hepatitis E virus (swine HEV), the first animal strain of HEV to be isolated, is a zoonotic agent. We report here the construction and in vitro and in vivo characterizations of infectious cDNA clones of swine HEV. Eight overlapping fragments spanning the entire genome were amplified by reverse transcription-PCR and assembled into a full-length cDNA clone, clone C, which contained 14 mutations compared to the consensus sequence of swine HEV. RNA transcripts from clone C were not infectious, as determined by intrahepatic inoculation into pigs and by in vitro transfection of Huh7 cells. Multiple site-based site-directed mutagenesis was performed to generate three new cDNA clones (pSHEV-1, pSHEV-2, and pSHEV-3) which differed from each other. The transfection of capped RNA transcripts into human liver Huh7 cells resulted in the synthesis of both ORF2 capsid and ORF3 proteins, indicating that the cDNA clones were replication competent. Each of the three clones resulted in active swine HEV infections after the intrahepatic inoculation of pigs with capped RNA transcripts. The patterns of seroconversion, viremia, and fecal virus shedding for pigs inoculated with RNA transcripts from clones pSHEV-2 and pSHEV-3 were similar to each other and to those for pigs inoculated with wild-type swine HEV, suggesting that the nucleotide differences between these two cDNA clones were not critical for replication. Pigs inoculated with RNA transcripts from clone pSHEV-1, which contained three nonsilent mutations in the ORF2 capsid gene, had a delayed appearance of seroconversion and fecal virus shedding and had undetectable viremia. The availability of these infectious cDNA clones affords us an opportunity to understand the mechanisms of cross-species infection by constructing chimeric human and swine HEVs.Hepatitis E virus (HEV), the causative agent of human hepatitis E, is a single positive-sense RNA virus in the new genus Hepevirus (8). HEV is transmitted by the fecal-oral route through contaminated drinking water. The mortality rate among hepatitis E patients is usually Ͻ1%, but it can reach up to 20% for infected pregnant women (12,14). Hepatitis E is rarely diagnosed in industrialized countries, even though a significant proportion of healthy individuals in these countries are positive for antibodies to HEV (19, 31). Antibodies to HEV have also been reported for various animal species (1, 10, 15, 22), suggesting that hepatitis E may be a zoonotic disease (21).In 1997, the first animal strain of HEV, swine HEV, was isolated and characterized from a pig in the United States (25). Experimental infections of specific-pathogen-free (SPF) pigs with swine HEV (23) and cross-species infections of HEV between swine and nonhuman primates (24) have been demonstrated. Swine HEV has since been identified in pigs in many other countries; in each case, it was found to be closely related to genotype 3 or 4 strains of human HEV (5, 16, 22). The prototype strain of swine HEV and two closely related U.S. strains of human HEV (US1 and US2) bel...

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