Serological and molecular survey of rat hepatitis E virus (Rocahepevirus ratti) in drug users

Abstract

ABSTRACTRat hepatitis E virus (ratHEV) is an emerging cause of acute hepatitis of zoonotic origin. Since seroprevalence studies are scarce, at-risk groups are almost unknown. Because blood-borne infections frequently occur in people with drug use, who are particularly vulnerable to infection due to lack of housing and homelessness, this population constitutes a priority in which ratHEV infection should be evaluated. Therefore, the aim of this study was to evaluate the ratHEV seroprevalence and RNA detection rate in drug users as a potential at-risk population. We designed a retrospective study involving individuals that attended drug rehabilitation centres. Exposure to ratHEV was assessed by specific antibody detection using ELISA and dot blot (DB) assay and the presence of active infection by ratHEV RNA detection using RT-qPCR. Three-hundred and forty-one individuals were included, the most of them being men (67.7%) with an average age of 45 years. A total of 17 individuals showed specific IgG antibodies against ratHEV (4.6%; 95% CI; 3.1%-7.9%). One case of active ratHEV infection was identified (0.3%; 95% CI: 0.1%-1.8%). This was a 57-year-old homeless woman with limited financial resources, who had active cocaine and heroin use via parenteral route. In conclusion, we identified a potential exposure to ratHEV among drug users. Targeted studies in drug users with proper control groups are necessary to evaluate high-risk populations and transmission routes more accurately.

Keywords: Rat hepatitis E virus; Zoonoses; drug users; hepatitis E; public health.

Figure 1.

DB results for sera that tested positive for ratHEV-specific IgG antibodies. Serum sample set of patients suffering for acute hepatitis E virus infection (n = 3), patients with acute ratHEV infection (n = 3) and patients with acute hepatitis of unknown origin (n = 5) were analyzed as control of the assay used (Figure 1(A)). Individuals included in the study with detection of ratHEV-specific IgG antibodies according to ELISA evaluation status (Figure 1(B)). The numbers correspond to the sample’s identification. His-tagged and affinity-purified carboxy-terminal segments of Escherichia coli (E. coli)-expressed capsid proteins served as antigens for HEV-3 and ratHEV (HEV and ratHEV). As negative substrate control (SC1 (-)), a his-tagged E. coli-expressed and affinity purified nucleocapsid protein derivative of the Puumala orthohantavirus strain Vranica/Hällnäs was used. As positive substrate control (SC2 (+)), an E. coli M15pREP4 total lysate was used.

Figure 2.

Phylogenetic analysis of the ratHEV sequence identified in the study. Sequence of the patient identified in the present study is marked with a circle (•). The evolutionary history was inferred by using the Maximum Likelihood method based on the Tamura-Nei model. The bootstrap consensus tree inferred from 1000 replicates is taken to represent the evolutionary history of the taxa analyzed. Branches corresponding to partitions reproduced in less than 50% bootstrap replicates are collapsed. Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Joining and BioNJ algorithms to a matrix of pairwise distances estimated using the Maximum Composite Likelihood (MCL) approach, and then selecting the topology with superior log likelihood value. The analysis involved 70 nucleotide sequences. Codon positions included were 1st + 2nd + 3rd + Noncoding. All positions containing gaps and missing data were eliminated. There are a total of 770 positions in the final dataset.