Identification of novel rodent and shrew orthohepeviruses sheds light on hepatitis E virus evolution

The family Hepeviridae has seen an explosive expansion in its host range in recent years, yet the evolutionary trajectory of this zoonotic pathogen remains largely elusive. The emergence of rat hepatitis E virus (HEV) poses a new public health threat due to its potential for zoonotic transmission. In this study, we investigated 2,464 wild small mammals spanning four animal orders, eight families, 21 genera, and 37 species in Yunnan Province, China. Employing conventional broadly reactive screening RT-PCR, we systematically explored the presence and prevalence of orthohepevirus. Our findings revealed that 192 specimens from 10 animal species tested positive for orthohepevirus, yielding an overall detection rate of 7.79%. Through genomic amplification via next-generation sequencing method, we obtained a total of 24 full-length genomic sequences of orthohepevirus strains from eight animal species, including Bandicota bengalensis, Eothenomys eleusis, and Episoriculus caudatus, marking their first reported host species for orthohepevirus. Our comprehensive sequence and phylogenetic analyses unveiled significant genetic diversity among these rodent and shrew orthohepeviruses. Notably, 20 strains were classified as Rocahepevirus ratti C1, two as C3, and one as Rocahepevirus eothenomi, while the remaining strain awaits classification due to its marked divergence. Additionally, our evolutionary analysis illuminated both the intimate relationship among orthohepeviruses within their host taxon and distinct clustering patterns across different host orders. Our results underscore the significance and occurrence of co-speciation and cross-species transmission during the evolutionary history of genera Paslahepevirus and Rocahepevirus.