# Genomic Characterization and Molecular Epidemiology of Tusaviruses and Related Novel Protoparvoviruses (Family Parvoviridae) from Ruminant Species (Bovine, Ovine and Caprine) in Hungary

## TL;DR

This study identifies and characterizes three new ruminant protoparvoviruses in Hungary, revealing their genetic relationships and host-specific patterns.

## Contribution

The discovery of three novel protoparvoviruses in bovine, ovine, and caprine species, along with their genomic and epidemiological analysis.

## Key findings

- Three novel protoparvoviruses (misavirus, sisavirus, gisavirus) were identified in ruminants with distinct host associations.
- Phylogenetic analysis suggests a shared origin among these viruses and tusaviruses, forming a separate lineage.
- Epidemiological data show high prevalence in young animals and potential interspecies recombination events.

## Abstract

Tusavirus 1 of species Protoparvovirus incertum 1 (family Parvoviridae) was first identified in humans and later in small ruminants (caprine and ovine). This study reports the full-length coding sequences (~4400–4600 nt) of three novel tusavirus-related protoparvoviruses from ovine (“misavirus”, PV540792), for the first time bovine (“sisavirus”, PV540793) and subsequently from caprine (“gisavirus” PV540850/51) fecal samples, using next-generation sequencing (NGS) and PCR techniques. Their NS1, VP1 and VP2 proteins shared 61–63% amino acid identities with each other and with tusaviruses, suggesting these three viruses belong to three novel species in the genus Protoparvovirus. Phylogenetic analyses placed them with tusaviruses on a separate main branch, implying a shared origin among these most likely ruminant protoparvoviruses. A small-scale epidemiological investigation on 318 ruminant enteric samples using novel generic NS1 primers found misavirus in 14/51 (27.5%) ovine and sisavirus in 19/203 (9.4%) bovine samples from multiple Hungarian farms. Tusavirus was present in 5/51 (9.8%) ovine and 15/62 (24.2%) caprine samples, all from one farm. The highest prevalences for all three viruses were found in animals aged 2–12 months, though sporadic cases were also found in other age groups. Partial NS and VP sequence-based phylogenetic trees showed virus-specific lineages for misa-, sisa-, gisa- and tusaviruses, with various strains forming sub-lineages. These findings suggest the presence of multiple genotypes and/or members of additional species, which was supported by a VP sequence-based hierarchical cluster analysis. The study’s viruses were mostly phylogenetically separated by host; however, two bovine sisavirus strains with diverse phylogenetic localizations in the NS (belonging to bovine sisaviruses) and VP1 trees (distantly related to ovine misaviruses) could indicate previous (interspecies?) recombination events.