Background Lyme borreliosis (LB) is the most common vector-borne disease in Europe. Monitoring changes in the prevalence of different Borrelia species in ticks may be an important indicator of risk assessment and of differences in pathogenicity in humans. The objective of our study was to assess the prevalence, co-infection and distribution of Borrelia and Babesia species in ticks removed from humans in a large sample collected during a study period of 4 years. Methods The ticks were collected throughout Poland from March to November over 4-year period from 2016 to 2019. All ticks (n = 1953) were morphologically identified in terms of species and developmental stage. Molecular screening for Borrelia and Babesia by amplification of the flagellin gene (flaB) or 18S rRNA marker was performed. Pathogen identity was confirmed by Sanger sequencing or PCR–restriction fragment length polymorphism analysis. Results The ticks removed from humans in Poland during this study belonged to two species: Ixodes ricinus (97%) and Dermacentor reticulatus (3%). High Borrelia prevalence (25.3%), including B. miyamotoi (8.4%), was confirmed in Ixodes ricinus ticks removed from humans, as was the change in frequency of occurrence of Borrelia species during the 4-year study. Despite Babesia prevalence being relatively low (1.3%), the majority of tested isolates are considered to be pathogenic to humans. Babesia infection was observed more frequently among Borrelia-positive ticks (2.7%) than among ticks uninfected with Borrelia (0.8%). The most frequent dual co-infections were between Borrelia afzelii and Babesia microti. The presence of Borrelia was also confirmed in D. reticulatus (12.7%); however the role of these ticks in spirochete transmission to susceptible hosts is still unclear. Conclusions Although the overall risk of developing LB after a tick bite is low in Europe, knowledge of the prevalence and distribution of Borrelia and Babesia species in ticks might be an important indicator of the risk of both these tick-borne diseases. Graphical abstract

Monitoring changes in the prevalence of different Borrelia genospecies/ species in ticks might be an important indicator of risk assessment and of differences in pathogenicity in humans. Furthermore, the evaluation of pathogens in feeding ticks represents the risk of human exposure better than studies on questing ticks. The objective of our study was to assess the prevalence and distribution of Borrelia and Babesia species in ticks removed from humans, in a larger sample collected for several months during four years of studies. We confirmed high Borrelia prevalence, including B. miyamotoi, in ticks removed from humans as well as the shift in Borrelia genospecies/ species frequency of occurrence during the four-year study. Despite the fact that Babesia prevalence was relatively low, the majority of tested isolates are considered to be pathogenic for humans. The results of our study have also shown that Borrelia and Babesia coinfections in ticks are more common in Borrelia-infected ticks. Even if the overall risk of developing Lyme borreliosis after a tick bite in Europe is rather low, the knowledge of prevalence and distribution of Borrelia and Babesia species in ticks might be an important indicator of both tick-borne disease risk and pathogenicity assessment.