As a leading viral cause of acute gastroenteritis in both humans and pigs, rotavirus A (RVA) poses a potential public health concern. Although zoonotic spillover of porcine RVA strains to humans is sporadic, it has been detected worldwide. The origin of chimeric human–animal strains of RVA is closely linked to the crucial role of mixed genotypes in driving reassortment and homologous recombination, which play a major role in shaping the genetic diversity of RVA. To better understand how genetically intertwined porcine and zoonotic human-derived G4P[6] RVA strains are, the present study employed a spatiotemporal approach to whole-genome characterization of RVA strains collected during three consecutive RVA seasons in Croatia (2018–2021). Notably, sampled children under 2 years of age and weanling piglets with diarrhea were included in the study. In addition to samples tested by real-time RT-PCR, genotyping of VP7 and VP4 gene segments was conducted. The unusual genotype combinations detected in the initial screening, including three human and three porcine G4P[6] strains, were subjected to next-generation sequencing, followed by phylogenetic analysis of all gene segments, and intragenic recombination analysis. Results showed a porcine or porcine-like origin for each of the eleven gene segments in all six RVA strains. The G4P[6] RVA strains detected in children most likely resulted from porcine-to-human interspecies transmission. Furthermore, the genetic diversity of Croatian porcine and porcine-like human G4P[6] strains was propelled by reassortment events between porcine and porcine-like human G4P[6] RVA strains, along with homologous intragenotype and intergenotype recombinations in VP4, NSP1, and NSP3 segments. Described concurrent spatiotemporal approach in investigating autochthonous human and animal RVA strains is essential in drawing relevant conclusions about their phylogeographical relationship. Therefore, continuous surveillance of RVA, following the One Health principles, may provide relevant data for assessing the impact on the protectiveness of currently available vaccines.

Hepatitis E is becoming a growing health concern in European countries as an increase of sporadic human cases of unknown origin has been recorded lately. Its causative agent, Hepatitis E virus (HEV), is known to have zoonotic potential and thus the role of domestic and wild animals in the chain of viral spread should be considered when investigating risk factors and the epidemiology of the disease. A comprehensive survey based on viral RNA detection was carried out in Croatia including blood, spleen and liver samples originating from 1816 different domestic and wild animals and digestive gland samples from 538 molluscs. A high HEV prevalence was detected in domestic pigs (24.5%) and wild boars (12.3%), whereas cattle, molluscs, ruminant and carnivore wildlife samples tested negative. Molecular characterization of both ORF1 and ORF2 genomic regions confirmed the phylogenetic clustering of the obtained sequences into genotype 3, previously reported in Europe. Furthermore, our results proved the presence of identical sequence variants in different samples, regardless of their origin, age or habitat of the host, suggesting transmission events between domestic swine, as well as between domestic swine and wild boars in the country. Moreover, a close genetic relationship of Croatian animal strains and known human HEV strains from GenBank opens the question of possible cross-species HEV transmission in Croatia, especially in the areas with an intensive swine production.

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During the hunting season in February 1999, a total of 44 blood samples were collected from wild boars shot in the area of Moslavacka gora. These blood samples were examined by enzyme immunoassay for the presence of antibodies to classical swine fever (CSFV), Aujeszky's disease (ADV), bovine viral diarrhoea (BVDV), and porcine reproductive and respiratory syndrome (PRRSV) viruses. Out of 44 serum samples examined, 17 (38.63%) were positive for CSFV, 24 (54.54%) were positive for ADV and two (4.54%) were positive for BVDV. All sera were negative for PRRSV. The results, recorded for the first time in Croatia, supported the hypothesis that wild boar act as a potential reservoir of CSFV, ADV and BVDV, and thus have a role in the epidemiology of these diseases.

This work presents the results of findings for brucellosis in wild boars and domestic swine in two regions of Croatia. In the region of Djakovo the blood samples of 211 wild boars were analysed and in 29.4% of the samples serologically positive reactions were established. In the same region the blood samples of 1080 domestic swine on pastures were also analysed and positive serological reactions were established in 12.3%. In the regions around Lonjsko Polje the blood samples of 53 wild boars were analysed and in 22.6% of them positive serological reactions were established. On several locations around Lonjsko Polje the blood samples of 901 domestic swine were serologically analysed and 13.5% of the swine were found to be seropositive. Bacteriological analyses of submitted materials from 24 wild boars resulted in isolation of Brucella from seven (29.2%) samples, and from 43 samples originating from domestic swine that had aborted and had been serologically positive, Brucella were isolated from 25 (58.1%) swine, as well as from 10 (62.5%) out of 16 aborted piglets. In all the isolates Brucella suis biovar 2 was identified. Wild boars are carriers and reservoirs of Brucella suis biovar 2 in Croatia.

Due to SARS CoV-2 recombination rates, number of infected people and recent reports of environmental contamination, the possibility of SARS CoV-2 transmission to animals can be expected. We tested samples of dominant free-living and captive wildlife species in Croatia for the presence of anti-SARS CoV-2 antibodies and viral RNA. In total, from June 2020 until February 2021, we tested blood, muscle extract and fecal samples of 422 free-living wild boars (Sus scrofa), red foxes (Vulpes vulpes) and jackals (Canis aureus); blood and cloacal swabs of 111 yellow-legged gulls (Larus michahellis) and fecal samples of 32 zoo animals. A commercially available ELISA (ID.Vet, France) and as a confirmatory test, a surrogate virus neutralization test (sVNT; GenScript, Netherlands) were used. Fecal samples were tested for the presence of viral RNA by a real-time RT–PCR protocol. Fifteen out of 533 (2.8%) positive ELISA results were detected; in wild boars (3.9%), red foxes (2.9%) and jackals (4.6%). However, the positive findings were not confirmed by sVNT. No viral RNA was found. In conclusion, no spillover occurred within the investigated period (second COVID-19 wave). However, further investigation is needed, especially regarding wildlife sample features for serological tests.

Hepatitis E has become an emerging infection in many European countries. We analysed the prevalence of hepatitis E virus (HEV) infection in selected population groups in Croatia. Overall HEV IgG seropositivity was 5.6%, while 1.9% participants showed IgM antibodies suggestive of recent infection. No IgM-positive sample was positive for HEV RNA. HEV IgG antibodies were most prevalent in alcohol abusers (8.9%) and war veterans (8.6%), compared with 6.1% among injecting drug users and 2.7% in healthcare professionals. No individual with high-risk sexual behaviour tested HEV seropositive. HEV IgG positivity increased significantly with age from 1.8% to 2.3% in individuals younger than 40 years to 11.3% in individuals older than 50 years (P = 0.023). The mean age of HEV-positive participants was significantly higher than that of HEV-negative participants (50.9 ± 11.8 years versus 41.2 ± 11.8 years, P = 0.008). Seroprevalence rates were significantly higher in residents of suburban and rural areas compared with residents of urban areas (14.5% versus 2.5%, P = 0.003). Additionally, an increasing prevalence of HEV IgG antibodies was observed from 1.8% in participants living in families with two household members to 12.1% in those living with more than four members (P = 0.046). Gender, marital status, educational level, sexual orientation, source of drinking water, history of blood transfusions, surgical procedures, tattooing and travelling were not associated with HEV seroprevalence. Logistic regression showed that living in suburban/rural areas was the main risk factor for HEV seropositivity (OR = 6.67; 95%CI = 1.89-25.0; AOR = 7.14, 95%CI = 1.89-25.0).

Introduction: Solid-organ transplant recipients are at risk of hepatitis E virus (HEV) infection. We analyzed the seroprevalence/ risk factors of HEV in Croatian liver transplant recipients. Methods: Two hundred forty-two serum samples were tested for HEV immunoglobuline IgG/IgM and HEV RNA. Sociodemographic data and risk factors were collected using a questionnaire. Results: HEV IgG seroprevalence rate was 24.4%. Positive/equivocal HEV IgM were found in two patients. HEV RNA was not detected. Logistic regression showed that older age, female gender, rural area/farm, water well, and septic tank were associated with HEV seropositivity. Conclusions: This study revealed a high exposure rate to HEV in Croatian liver recipients.