Molecular detection of hepatitis E virus in sheep from southern Xinjiang, China

Abstract

Hepatitis E virus (HEV) is a causative agent of infectious hepatitis in animals and humans both in developing and developed countries. Here, we collected 500 sheep sera and 75 raw sheep liver samples from a slaughterhouse in the southern part of the Xinjiang region, China, along with 26 sera of butchers from the same slaughterhouse. All serum samples were tested for anti-HEV antibody by enzyme-linked immunosorbent assay. Both serum and liver samples were evaluated for the presence of HEV RNA by nested polymerase chain reaction targeting partial nucleotide sequences of open reading frame 2 (ORF2). The results indicate that sheep seroprevalence was 35.20 % (176/500) and that four of the 75 (5.3 %) sheep livers showed detectable amounts of HEV RNA. The seroprevalence of the butchers was 57.7 % (15/26). The four amplicons shared 97.8-100 % nucleotide sequence identity and had pairwise sequence identities of 81.6-85.3 %, 84.2-85.3 %, 82.1-85.3 % and 84.7-97.9 % with the corresponding regions of genotypes 1, 2, 3 and 4 of HEV, respectively. A phylogenetic tree was constructed based on alignments of an amplified 186-bp ORF2 sequence and corresponding reference strains. The analysis showed that the four sheep strains detected in our study formed a lineage within a genotype 4 cluster that contains hb-3, bjsw1, T1, swCH189 and swCH25, all of which belong to genotype 4, subtype 4d. The results indicated a high level of seroconversion in sheep and suggested that sheep liver may be a source of foodborne HEV infection in humans.

Fig. 1

Geographical distribution of the three different farms in Xinjiang. On the map of China, different provinces are marked with different colours. The Xinjiang Uighur Autonomous Region is marked in light blue. The three farms were distributed in the southern part of Xinjiang, which is demarcated by a black line. Open star indicates Hetian region. Open circle indicates Hetian county of Hetian region where farms A and B were located and which corresponds to the area where the two large-scale outbreaks during 1986 and 1988 occurred. Open diagonal indicates Aksu region where farm C was located (Color figure online)

Fig. 2

Phylogenetic tree based on a 186-bp nucleotide sequences of HEV-ORF2. The tree was constructed by the neighbour-joining method using 46 animal and human HEV reference strains. Strains characterised in this study were indicated with symbol filled triangle. The internal node numbers indicated the bootstrap values as a percentage obtained from 1000 replications. The Arabic numbers and the English letters outside of the dashed curves indicate the genotype and subtype. The bar on the tree shows the scale of 0.05 % nucleotide substitution per site. GenBank accession numbers, country and species of origin are indicated for each isolate represented on the tree