Prevalence and Molecular Characterization of the Hepatitis E Virus in Retail Pork Products Marketed in Canada

Abstract

Infection with the hepatitis E virus (HEV) is very common worldwide. HEV causes acute viral hepatitis with approximately 20 million cases per year. While HEV genotypes 1 and 2 cause large waterborne and foodborne outbreaks with a significant mortality in developing countries, genotypes 3 and 4 are more prevalent in developed countries with transmission being mostly zoonotic. In North America and Europe, HEV has been increasingly detected in swine, and exposure to pigs and pork products is considered to be the primary source of infection. Therefore we set out to investigate the prevalence of HEV in retail pork products available in Canada, by screening meal-size portions of pork pâtés, raw pork sausages, and raw pork livers. The presence of the HEV genomes was determined by RT-PCR and viral RNA was quantified by digital droplet PCR. Overall, HEV was detected in 47% of the sampled pork pâtés and 10.5% of the sampled raw pork livers, but not in the sampled pork sausages, and sequencing confirmed that all HEV strains belonged to genotype 3. Further phylogenetic analysis revealed that except for one isolate that clusters with subtype 3d, all isolates belong to subtype 3a. Amino acid variations between the isolates were also observed in the sequenced capsid region. In conclusion, the prevalence of HEV in pâtés and raw pork livers observed in this study is in agreement with the current HEV distribution in pork products reported in other developed countries.

Keywords: Droplet digital PCR; Hepatitis E virus; Molecular detection; Phylogenetic analysis; Pork products.

Fig. 1

Phylogenetic tree constructed by the neighbor-joining method, based on the 300 nucleotides at the 5′ end of ORF2 gene of the identified isolates in this study as well as closely related strains isolated from swine and human patients. The subtypes of the reference genomes (Smith et al. 2014) are shown in parenthesis. The robustness of the phylogenetic analysis was assessed through bootstrap analysis of 1000 pseudo-replicates. The scale bar represents 2% sequence divergence. Each entry is identified with its GenBank Accession Number or the isolate name, as well as the region from which it was isolated. QC province of Quebec, Canada

Fig. 2

Non-synonymous differences within 100 amino acid sequence of the N domain of the capsid protein (ORF2). Residues are numbered according to a Quebec isolate (Accession No: DQ832264)