The Borrelia burgdorferi s.l. complex is known for its complex genome and significant genetic variability. We used multilocus sequence typing (MLST) to study the genetic variability of Borrelia in five European countries: Slovakia, Croatia, Ukraine, Finland and Bulgaria. We compared the prevalence, genospecies spectrum and genetic variability of Borrelia in 4013 questing Ixodes ricinus ticks. The highest prevalence of Borrelia was found in Slovakia (26.7 %) and Ukraine (22 %), while in Finland and Bulgaria the prevalence was found to be 12.4 % and 7.2 %, respectively. Genospecies diversity was the highest in Croatia with seven identified genospecies. This was in contrast to Finland and Bulgaria where only three species were detected. In phylogenetic trees Borrelia species clustered according to species designation. Borrelia afzelii and Borrelia garinii were the most prevalent amongst the Borrelia species and were found in all collection regions. Borrelia garinii but also B. afzelii isolates did not show a phylogeographic pattern in minimum spanning trees. Based on MLST, Borrelia lusitaniae isolates from Slovakia, Croatia, Ukraine and Bulgaria belonged to the “northern European” clade, which differs genetically from B. lusitaniae isolates found in southern Portugal and northern Africa. Borrelia bavariensis from the Slovak Republic and Ukraine firmly clustered with Western European isolates. Besides questing ticks, Edible dormice (Glis glis) earlobe biopsies were analyzed as B. afzelii clustering with other European isolates of this species. In this study we report the first evidence of Borrelia genospecies in the following countries: Borrelia spielmanii, B. bavariensis and B. lusitaniae in Ukraine; B. spielmanii in Bulgaria; Borrelia burgdorferi s.s., B. spielmanii, B. lusitaniae and B. bavariensis in Croatia.
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