Recently, mammalian orthoreoviruses (MRVs) were detected for the first time in European bats, and the closely related strain SI-MRV01 was isolated from a child with severe diarrhoea in Slovenia. Genetically similar strains have also been reported from other mammals, which reveals their wide host distribution. The aim of this study was to retrospectively investigate the occurrence and genetic diversity of MRVs in bats in Slovenia, from samples obtained throughout the country in 2008 to 2010, and in 2012 and to investigate the occurrence of the novel SI-MRV01 MRV variant in Slovenian bats. The detection of MRVs in bat guano was based on broad-range RT-PCR and specific bat MRV real-time RT-PCR. Subsequently, MRV isolates were obtained from cell culture propagation, with detailed molecular characterisation through whole-genome sequencing. Overall, bat MRVs were detected in 1.9% to 3.8% of bats in 2008, 2009 and 2012. However, in 2010 the prevalence was 33.0%, which defined an outbreak of the single SI-MRV01 strain. Here, we report on the identification of five MRV isolates of different serotypes that are designated as SI-MRV02, SI-MRV03, SI-MRV04, SI-MRV05 and SI-MRV06. There is high genetic variability between these characterised isolates, with evident genome reassortment seen across their genome segments. In conclusion, we have confirmed the presence of the SI-MRV01 strain in a Slovenian bat population. Moreover, according to genetic characterisation of S1 genome segment, all three MRV serotypes were present in the bat population. In this study, five independent MRV isolates were obtained and detailed whole genome analysis revealed high diversity between them. This study generates new information about the epidemiology and molecular characteristics of emerging bat MRV variants, and provides important molecular data for further studies of their pathogenesis and evolution.

Lyssaviruses are the causative agents of rabies, a zoonotic, fatal disease that is thought to be ancestral to bats. In the last decade, the detection of bat associated lyssaviruses is increasing also in Europe. Within a retrospective bat associated lyssavirus surveillance study a total of 225 dead bats of 21 bat species were collected in Slovenia between 2012 and 2019 and tested by specific real-time RT-PCR method. The first lyssavirus positive sample in bats in Slovenia was detected using the real-time RT-PCR, the fluorescent antibody test, and next generation sequencing, while the rabies tissue culture inoculation test was unsuccessful due to sample degradation and storage conditions. The nearly complete genome of Divača bat lyssavirus from Slovenia consists of 11,871 nucleotides and reflects the characteristic gene organization known for lyssaviruses, encoding the five viral proteins. Phylogenetic analysis of Divača bat lyssavirus revealed that it belongs to phylogroup I lyssaviruses and is most closely related to Kotalahti bat lyssavirus (KBLV) with 87.20% nucleotide and 99.22% amino acid identity. Together with KBLV, Khujand virus, European bat lyssavirus 2, Bakeloh bat lyssavirus, and Aravan virus, Divača bat lyssavirus was detected in the genus Myotis suggesting its key role in the transmission and maintenance of certain lyssaviruses.

In the 1950s, infectious bovine rhinotracheitis/infectious pustular vulvovaginitis (IBR/IPV) disease was clinically detected and documented in cattle for the first time in Slovenia. The bovine herpes virus 1 (BoHV-1) was confirmed several times from infected herds by virus isolation on cell cultures. To keep the IC virus-free, high biosecurity measures were introduced. Before entering the IC, all calves are serologically tested and quarantined. Bulls in Slovenian insemination centres (IC) have been negative for IBR /IPV infection since 1979. From 1985 to 1991, few large-scale studies of the prevalence of IBR/IPV were carried out. In 1985, a high percentage (56.9%) of serologically positive animals were found in large state farms with Holstein Friesian cattle. Epidemiological studies in farm with bulls' mother herds were also carried out in the farms with Simmental and Brown cows. Antibodies against BoHV-1 were detected in the serum of 2.3% of Brown cattle and 3.5% of Simmental cattle. In the year 2000, 3.4% of bulk tank milk samples from 13,349 dairy farms were detected BoHV-1 antibodies positive. The highest percentage of positive animals was found in regions with an intensive grazing system (6.2% positive) and the lowest percentage in the east part of Slovenia (0.9% positive) on farms with mostly Simmental cattle. In 2006, a total 204,662 sera of cattle older than 24 months were tested for the presence of BoHV-1 antibodies and positive cattle were detected in 3.6% of tested farms. These farms kept 34,537 animals that were potential carriers of the BoHV-1. Most of the positive farms kept Holstein Friesian cattle, descendants from the state-owned farms, which were privatised or closed after 1990. In 2015, the Administration of the Republic of Slovenia for Food Safety, Veterinary and Plant Protection issued a rule that describes the conditions for granting and maintaining the status of BoHV-1 free holdings. The rule provides a voluntary control programme for breeders who want to obtain BoHV-1 free status and are willing to cover all the cost of acquiring and maintaining that status. There has been very little response from breeders.

Coronaviruses (CoV) are widely distributed pathogens of human and animals and can cause mild or severe respiratory and gastrointestinal disease. Antigenic and genetic similarity of some CoVs within the Betacoronavirus genus is evident. Therefore, for the first time in Slovenia, we investigated the genetic diversity of partial 390-nucleotides of RNA-dependent-RNA polymerase gene (RdRp) for 66 human (HCoV) and 24 bovine CoV (BCoV) positive samples, collected between 2010 and 2016 from human patients and cattle with respiratory disease. The characterized CoV strains belong to four different clusters, in three separate human clusters HCoV-HKU1 (n = 34), HCoV-OC43 (n = 31) and HCoV 229E (n = 1) and bovine grouping only as BCoVs (n = 24). BCoVs from cattle and HCoV-OC43 were genetically the most closely related and share 96.4–97.1% nucleotide and 96.9–98.5% amino acid identity.

Sylvatic rabies was present in Slovenia between 1973 and 2013, with the red fox as the main reservoir of the rabies virus. The first oral rabies vaccination (ORV) control program in foxes started in 1988, using the manual distribution of baits. Significant improvement of fox vaccination was achieved with the aerial distribution of baits, starting in 1995 and successfully finished with the final, fifty-ninth vaccination campaign in 2019. Between 1979 and 2019, a total of 86,471 samples were tested, and 10,975 (12.69%) rabies-positive animals were identified. Within the ORV, two different vaccines were used, containing modified live virus strain Street Alabama Dufferin (SAD) B19 and SAD Bern, while the last ORV campaigns were completed in 2019, with a vaccine containing a genetically modified strain of SPBN GASGAS. Molecular epidemiological studies of 95 rabies-positive samples, originating from red foxes, badgers, cattle, dogs, martens, cats, and horses, revealed a low genetic diversity of circulating strains and high similarity to strains from neighboring countries. During the elimination program, few vaccine-induced rabies cases were detected: three in red foxes and one case in a marten, with no epidemiological relevance. Slovenia has been officially declared a country free of rabies since 2016.

Pseudorabies (PR) is one of the most economically important diseases in domestic pigs. Since 2010, Slovenia has been free of PR in the domestic pig population, but the disease is endemic in the wild boar population, which can pose a real threat to domestic pigs and other animal species, including dogs. Between 2006 and 2020, infections with the PR virus (PRV) were reported in two pets and three hunting dogs from Slovenia that were found to have a direct contact with the wild boar or raw wild boar or pork meat. Typical clinical signs of PRV infection, including characteristic facial itching, cytopathic effect in cell cultures, positive immunocytochemistry, and positive PCR results confirmed the presence of PRV in all five cases investigated. A phylogenetic comparison of the partial glycoprotein C (gC) genomic region revealed that the Slovenian PRV isolates belong to clade A, with 95.78100 nucleotide identity with strains isolated from dogs, domestic pigs, and wild boars from Europe. Within phylogenetic comparison of the partial glycoprotein D (gD) and partial glycoprotein E (gE) genomic regions of Slovenian PRV isolates, 100 and 99.12100 nucleotide identities were detected, respectively, suggesting low diversity between the PRV strains identified in dogs in Slovenia. This study provides the first molecular characterization of PRV in dogs and suggests that similar PRV strains circulate in the wild boar populations in this geographic area.