Caliciviruses including noro- and sapoviruses of family Caliciviridae are important enteric human and swine pathogens, while others, like valoviruses, are less known. In this study, we developed a detection and typing pipeline for the most prevalent swine enteric caliciviruses—sapovirus GIII (Sw-SaV), norovirus GII (Sw-NoV), and valovirus GI (Sw-VaV). The pipeline integrates triplex RT-qPCR, 3′RACE semi-nested PCR, and next-generation sequencing (NovaSeq, Illumina) techniques. A small-scale epidemiological investigation was conducted on archived enteric and, for the first time, on oral fluid/saliva samples of diarrheic and asymptomatic swine of varying ages from Hungary and Slovakia. In enteric samples, Sw-SaV was the most prevalent, detected in 26.26% of samples, primarily in diarrheic pigs with low Cq values, followed by Sw-NoV (2.53%) in nursery pigs. In oral fluid samples, Sw-NoV predominated (7.46%), followed by Sw-SaV (4.39%). Sw-VaVs were sporadically found in both sample types. A natural, asymptomatic Sw-SaV outbreak was retrospectively detected where the transient shedding of the virus was <2 weeks. Complete capsid sequences (n = 59; 43 Sw-SaV, 13 Sw-NoV, and 3 Sw-VaV) including multiple (up to five) co-infecting variants were identified. Sw-SaV sequences belong to seven genotypes, while Sw-NoV and Sw-VaV strains clustered into distinct sub-clades, highlighting the complex diversity of these enteric caliciviruses in swine.

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.

Various microbial agents have been found in the feces of both humans and animals, especially in newborns. While some of these agents are recognized as causing diarrhea, the role of others, specifically bopiviruses of the family Picornaviridae, in diarrhea remains uncertain. In this study, we conducted an analysis of 214 fecal samples from cattle (n = 114), sheep (n = 82), and goats (n = 18) with diarrhea, collected from farms across 17 different provinces in Türkiye. All samples were tested using RT-PCR targeting the 3D^(RdRp) region of bopiviruses, and two samples from sheep (2.4%) tested positive. The 7303 nt-long complete coding sequence of Bopivirus/Sheep/KS-1M/2024/TUR and partial 3D^(RdRp), VP3, and 2A-2C sequences of Bopivirus/Sheep/ANK-K30/2017/TUR were determined by additional RT-PCR, 3′RACE-PCR reactions and Sanger sequencing. Both strains show close sequence and phylogenetic relationship to members of species “Bopivirus B” of genus Bopivirus. Bopivirus/Sheep/KS-1M/2024/TUR is most closely related to a sheep Bopivirus B strain (sheep/14-73/2018/ITA) from Italy, but the phylogenetic separation, the low sequence identities and high p-distance values in VP1 to existing genotypes of “B1” and “B2” suggest that both strains could belong to novel genotypes (“B3” and “B4”) in species “Bopivirus B”, although additional closely related sequences are necessary for proper typing.