High Prevalence and Genetic Variability of Hepatozoon canis in Grey Wolf (Canis lupus L. 1758) Population in Serbia

SIMPLE SUMMARY: There are a number of different animal species that can be infected by Hepatozoon canis, but the most common are domestic and wild carnivores. This study presents the first results of the occurrence, distribution, and diversity of H. canis in Serbia and contributes to the limited knowledge regarding the role of grey wolves in the enzootic cycles of this pathogen. Based on a comparison with previously published data, we found a higher prevalence and higher genetic diversity of H. canis. The obtained results showed an overall high prevalence of the pathogen, with 57.94% of tested animals positive for H. canis, while genetic analysis of a sequenced fragment of the 18S ssrRNA gene showed variability at five positions leading to five sequence types present in grey wolves, where only two were previously known of. In addition to the known patterns of transmission of this pathogen, through tick ingestion during grooming or the transplacental route, the high diversity of H. canis in Serbia could be explained by the diet of grey wolves in this area. Further studies are needed to determine the mechanism of transmission, the potential source of infection, and the impact of this pathogen on wild carnivores. ABSTRACT: Wild canids are globally recognised as hosts and reservoirs of a large number of ecto- and endoparasites. Data that reveal the importance of the grey wolf (Canis lupus L.1758) in the spread of hepatozoonosis are very scarce. There are a large number of different potential host species that can be infected by Hepatozoon canis, but the most common are domestic and wild carnivores, such as dogs, jackals, foxes, and wolves. In this study, the epidemiological significance of the grey wolf as a host for the pathogen was analysed for the first time in Serbia, as well as the genetic variability of H. canis. The presence of H. canis in wolf spleens has been demonstrated using molecular methods. A total of 107 wolf spleen samples from 30 localities in Serbia were analysed. The presence of H. canis was confirmed in 62 (57.94%) individuals from 26 out of 30 localities. According to the analysis, the sampled H. canis sequences were found to be characterised by a certain heterogeneity. Based on five mutated nucleotide sites in the sequences, H. canis could be divided into five sequence types, S1 to S5. The five sequence types can potentially circulate in grey wolf populations as well as among other domestic and wild canids. This study is the first confirmation of the presence of H. canis in grey wolf populations in Serbia. Considering that the role of this vector-borne disease is poorly researched in wild carnivores, it is very important to indicate the role of this species in the circulation of this pathogen in natural ecosystems.