Structural insights into NS5B protein of novel equine hepaciviruses and pegiviruses complexed with polymerase inhibitors.

Infections produced by hepaciviruses have been associated with liver disease in horses. Currently, at least three viruses belonging to the Flaviviridae family are capable of producing a chronic infection in equines: non-primate hepacivirus (NPHV), Theiler's disease-associated virus (TDAV), and equine pegivirus (EPgV). The RNA-dependent RNA polymerases of viruses (RdRp) (NS5 protein), from the flavivirus family, use de novo RNA synthesis to initiate synthesis. The two antiviral drugs currently used to treat hepatitis C (HCV), sofosbuvir and dasabuvir, act on the viral NS5B polymerase as nucleoside and non-nucleoside inhibitors, respectively. Both drugs have shown significant clinical inhibition of viral response. In this work, we aimed to model the NS5B polymerase of the equine hepacivirus (EHCV) subtypes 1 and 2, TDAV and EPgV, to assess whether current direct-acting antiviral drugs against HCV interact with these proteins. Crystal structures of HCV-NS5B were used as templates for modeling target sequences in both conformations (open and closed). Also, molecular docking of sofosbuvir and dasabuvir were performed to predict their possible binding modes at the modeled NS5B polymerase binding sites. We observed that the NS5B models of the EHCV and EPgV shared well-conserved 3D structures to HCV-NS5B and other RdRps, suggesting functional conservation. Interactions of EHCV subtypes 1, 2 and TDAV polymerases with sofosbuvir showed a similar molecular interaction pattern compared to HCV-NS5B, while interactions with dasabuvir were less conserved. In silico studies of molecular interactions between these modeled structures and sofosbuvir suggest that this compound could be efficient in combating equine pathogens, thus contributing to animal welfare.

Epidemiological investigation and analysis of the NS5B gene and protein variability of non-primate hepacivirus in several horse cohorts in Rio de Janeiro state, Brazil.

Among the hepacivirus species recently described, the non-primate hepacivirus/hepacivirus A found in horses and donkeys is closely related to the human hepatitis C virus (HCV). Therefore, the equine is an attractive surrogate large animal model for the study of HCV therapy, pathogenesis and prophylaxis. Despite global efforts, epidemiological and genetic studies have not elucidated the risk factors, virus distribution or genetic variability of the hepacivirus A, which are also important issues for the equine welfare. Little information about this background scenery is available in Brazil. The aims of this study were to investigate potential risk factors associated with hepacivirus A infection among different horse cohorts throughout the state of Rio de Janeiro and to evaluate the diversity of the viral NS5B gene and protein. Hepacivirus A RNA was detected in horse cohorts from all geographical mesoregions, independent of horse activity or breed investigated. Statewide prevalence ranged from 4.0% to 27.5%. Potential risk factors such as geographical location and age of female horses were significantly associated with the presence of virus RNA. Phylogenetic analysis revealed the circulation of subtype 2 in all mesoregions. NS5B gene sequences clustered according to geographical origin, while the NS5B fragments did not allow discriminant analysis. The predicted NS5B protein showed marked conservation, especially in the thumb domain. In conclusion, the higher frequency of hepacivirus A RNA detection in horses bred for reproduction purposes as well as in young females suggests a direct link between reproduction practices and the virus's spread. Additional studies are necessary to understand the distribution of this genetically conserved hepacivirus.