The hepatitis E virus (HEV) hypervariable region (HVR) presents the highest divergence of the entire HEV genome. It is characteristically rich in proline, and so is also known as the “polyproline region” (PPR). HEV genotype 3 (HEV-3) exhibits different PPR lengths due to insertions, PPR and/or RNA-dependent RNA polymerase (RdRp) duplications and deletions. A total of 723 PPR-HEV sequences were analyzed, of which 137 HEV-3 sequences were obtained from clinical specimens (from acute and chronic infection) by Sanger sequencing. Eight swine stool/liver samples were also analyzed. N- and C-terminal fragments were confirmed as being conserved, but they harbored differences between genotypes and were not proline-plentiful regions. The genuine PPR is the intermediate region between them. HEV-3 PPR contains a higher percentage (30.4%) of prolines than other genotypes. We describe for the first time: (1) the specific placement of HEV-3 PPR rearrangements in sites 1 to 14 of the PPR, noting that duplications are more frequently attached to sites 11 and 12 (AAs 74–79 and 113–118, respectively); (2) the cadence of repetitions follows a circular-like pattern of blocks A to J, with F, G, H, and I being the most frequent; (3) a previously unreported insertion homologous to apolipoprotein C1; and (4) the increase in frequency of potential N-glycosylation sites and differences in AAs composition related to duplications.

Emerging infectious diseases are one of the most important global health challenges because of their impact on human and animal health. The vector-borne West Nile virus (WNV) is transmitted between birds by mosquitos, but it can also infect humans and horses causing disease. The local circulation of WNV in Spain has been known for decades, and since 2010, there have been regular outbreaks in horses, although only six cases were reported in humans until 2019. In 2020, Spain experienced a major outbreak with 77 human cases, which was followed by 6 additional cases in 2021, most of them in the Andalusian region (southern Spain). This study aimed to characterize the genomes of the WNV circulating in wild-trapped mosquitoes during 2020 and 2021 in Andalusia. We sequenced the WNV consensus genome from two mosquito pools and carried out the phylogenetic analyses. We also compared the obtained genomes with those sequenced from human samples obtained during the outbreak and the genomes obtained previously in Spain from birds (2007 and 2017), mosquitoes (2008) and horses (2010) to better understand the eco-epidemiology of WNV in Spain. As expected, the WNV genomes recovered from mosquito pools in 2020 were closely related to those recovered from humans of the same outbreak. In addition, the strain of WNV circulating in 2021 was highly related to the WNV strain that caused the 2020 outbreak, suggesting that WNV is overwintering in the area. Consequently, future outbreaks of the same strain may occur in in the future.

HEV antibody detection constitutes the main screening test for HEV infection. The aim of this study is to compare the sensitivity and specificity of four techniques: LIAISON^® MUREX DiaSorin anti-HEV IgG and anti-HEV IgM assays, Hepatitis E VIRCLIA^® IgM and IgG monotests, WANTAI HEV-IgM and IgG ELISA and VIDAS^® anti-HEV IgM and IgG tests in five panels of samples configurated according to the immunoblot (RecomLine, Mikrogen, Neuss, Germany). Anti-HEV IgM sensitivity in the acute phase was 100% in all techniques, while sensitivity, including the immediate convalescence phase, was 96.74% for LIAISON^®, 83.14% for VIRCLIA^®, 84.78% for WANTAI and 88.04% for VIDAS^®. Anti-HEV IgM specificity was 100% for both LIAISON^® and VIRCLIA^®. Anti-HEV IgM WANTAI agreed with VIRCLIA^® with a good Kappa coefficient (κ = 0.71). Anti-HEV IgG post-infection sensitivity was 100% for LIAISON^®, VIDAS^® and VIRCLIA^® and 99% for WANTAI. Anti-HEV IgG specificity reached 97.17% for LIAISON and 88.68% for VIRCLIA^®. Our results demonstrated a better capacity of LIAISON^® MUREX anti-HEV IgM than that of competitors for detecting acute infections as well as accurate anti-HEV IgG results and in how to resolve them.