Detection, genetic characterization and reassortment analysis of Tribeč virus

This study reports the detection and genetic characterization of Tribeč virus (TRBV), a tick-borne orbivirus of zoonotic relevance. The virus was identified in Ixodes ricinus ticks collected from a goat using a newly developed RT-PCR assay. Goats grazing in the study area showed seroconversion, with low titre neutralization antibody response. The new isolate, designated 16.C/2016/Dubrava/SVK, was successfully cultured in Vero E6 cells and detected by immunofluorescence. Replication efficiency varied across rodent, human, and bovine cell lines. Whole-genome sequencing revealed that most genome segments shared high sequence identity with other TRBV strains and with Muko virus (MUKV) strains from Japan. In contrast, phylogenetic analysis based on the VP4 outer capsid protein indicated that the TRBV clade is more closely related to Kemerovo viruses from Russia. Segment 10 showed the highest variability, where strains from Slovakia and Romania formed sister clades with MUKV, and the Ukrainian strains with KEMV. Indication of a reassortment was observed in segment 4, encoding the NS1 tubule-forming protein, which appears to originate from the prototype TRBV isolated in Slovakia during the 1960s. Collectively, these findings highlight the genetic variability of TRBV and support the need for further studies on the ecology, evolution, and diversity of tick-borne orbiviruses.