Hepatitis E Virus RNA Detection in Liver and Muscle Tissues Sampled from Home Slaughtered Domestic Pigs in Central Italy

Abstract

Hepatitis E virus is a worldwide emerging foodborne pathogen; raw or undercooked meats and liver pork products can cause infection through the orofecal route. In Central-Southern Italy, small traditional farming method, associated with the possibility of environmental sharing with wild species, can facilitate HEV diffusion and persistence. The aim of this study was to determine HEV genotype and subtype in Marche region from home slaughtered domestic pigs involved in small and traditional food chains. A total of 236 liver and muscle tissues and 6 pooled salami samples were screened. Laboratory workflow started with homogenization, followed by RNA extraction. Nested reverse transcription PCR and qRT-PCR were used to amplify specific parts of overlapping open reading frames belonging to the HEV genome. A total of 42/236 (17.79%) liver and 8/236 (3.39%) diaphragm specimens were positive; none of the pooled salami specimens showed positive HEV signal. The discovered HEV3c presented high nucleotide similarities with ones amplified from wild boar populations hunted in the same province. Extensive farming methods and environmental sharing with wild animal species support cross-infection infections, as observed in the present study. Although salami resulted negative for HEV RNA detection, the effects of food technologies on viral loads remain unclear. Therefore, further scientific investigations coupled with efficacious standardized laboratory procedures will be the next challenge.

Keywords: Environmental sharing; Food safety; Hepatitis E virus; Liver and muscle; Molecular biology; One-health.

Fig. 1

Geographical distribution of positive domestic pigs and their respective farming areas in the Ascoli Piceno province, Marche region, Italy

Fig. 2

Neighboring-joining phylogenetic tree constructed basing on the nucleotide fragment of 333 bp, referring to HEV ORF1 from liver and muscle samples. The p-distance model with bootstrapping of 1000 replicates was performed. Sequences OR933697 and OR933698 (red marked) were discovered from liver and muscle tissue