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. 2021 Apr 16;10(4):485.
doi: 10.3390/pathogens10040485.

A Remarkable Genetic Diversity of Rotavirus A Circulating in Red Fox Population in Croatia

Daniel Čolić  1   2 , Nina Krešić  1 , Željko Mihaljević  1 , Tibor Andreanszky  3 , Davor Balić  4 , Marica Lolić  4 , Dragan Brnić  1
Affiliations

A Remarkable Genetic Diversity of Rotavirus A Circulating in Red Fox Population in Croatia

Daniel Čolić et al. Pathogens. .

Abstract

Rotaviruses (RV), especially Rotavirus A (RVA), are globally recognized as pathogens causing neonatal diarrhea, but they also affect intensive animal farming. However, the knowledge on their significance in wildlife is rather limited. The aim of the study was to unveil the prevalence, molecular epidemiology, and genetic diversity of RVA strains circulating in the red fox (Vulpes vulpes) population in Croatia. From 2018 to 2019, 370 fecal samples from fox carcasses hunted for rabies monitoring were collected. All samples were first tested using a VP2 real-time RT-PCR; in the subsequent course, positives were subjected to VP7 and VP4 genotyping. The results revealed an RVA prevalence of 14.9%, while the circulating RVA strains showed a remarkable genetic diversity in terms of 11 G and nine P genotypes, among which one G and three P were tentatively identified as novel. In total, eight genotype combinations were detected: G8P[14], G9P[3], G9P[23], G10P[11], G10P[3], G11P[13], G15P[21], and G?P[?]. The results suggest a complex background of previous interspecies transmission events, shedding new light on the potential influence of foxes in RVA epidemiology. Their role as potential reservoirs of broad range of RVA genotypes, usually considered typical solely of domestic animals and humans, cannot be dismissed.

Keywords: Croatia; Rotavirus A; genetic diversity; genotype; interspecies transmission; molecular epidemiology; phylogenetic analysis; red fox.

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Conflict of interest statement

The authors declare no conflict of interest. The funding body had no role in the study design; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the study.

Figures

Figure 1
Figure 1
Phylogenetic relatedness of fox Rotavirus A (RVA) strains detected in Croatia with the reference RVA strains, determined on the basis of the VP7 genome segment (G1 and G2 genotypes). According to the alignment of shorter VP7 nucleotide sequences (~300 nt), the phylogenetic tree was generated using the MEGAX Software and the maximum-likelihood (ML) method by virtue of applying the T92 + G substitution model. The stability of the proposed branching order was assessed by bootstrapping (1000 replicates; indicated adjacent to the nodes when >70%). Fox RVA strains detected in Croatia are given in bold. The GenBank accession numbers of the selected RVA reference strains are designated within taxa. The scale bar represents the number of nucleotide substitutions per site. For the sake of simplicity in displaying the RVA strain nomenclature, P genotype numbers were omitted from the brackets.
Figure 2
Figure 2
Phylogenetic relatedness of fox RVA strains detected in Croatia with the reference RVA strains, determined on the basis of the VP7 genome segment (G8, G9, G10, G11, and G15 genotypes and a tentatively novel one). According to the alignment of longer VP7 nucleotide sequences (~800 nt), the phylogenetic tree was generated using the MEGAX Software and the maximum-likelihood method by virtue of applying the T92 + G substitution model. The stability of the proposed branching order was assessed by bootstrapping (1000 replicates; indicated adjacent to the nodes when >70%). Fox RVA strains detected in Croatia are given in bold. The tentatively novel G genotype is represented by two RVA strains tagged with black dots. The GenBank accession numbers of the selected RVA reference strains are designated within taxa. The scale bar represents the number of nucleotide substitutions per site. For the sake of simplicity in displaying the RVA strain nomenclature, P genotype numbers were omitted from the brackets.
Figure 3
Figure 3
Phylogenetic relatedness of fox RVA strains detected in Croatia with the reference RVA strains, determined on the basis of the VP4 genome segment. According to the alignment of VP4 nucleotide sequences (~680 nt), the phylogenetic tree was generated using the MEGAX Software and the maximum-likelihood method by virtue of applying the TN93 + G substitution model. The stability of the proposed branching order was assessed by bootstrapping (1000 replicates; indicated adjacent to the nodes when >70%). Fox RVA strains detected in Croatia are given in bold. The RVA strains of tentatively novel P genotypes are tagged with black dots. The GenBank accession numbers of the selected RVA reference strains are designated within taxa. The scale bar represents the number of nucleotide substitutions per site. For the sake of simplicity in displaying the RVA strain nomenclature, P genotype numbers were omitted from the brackets.
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