Metabarcoding reveals low prevalence of microsporidian infections in castor bean tick (Ixodes ricinus)

BACKGROUND: Microsporidia is a large group of eukaryotic obligate intracellular spore-forming parasites, of which 17 species can cause microsporidiosis in humans. Most human-infecting microsporidians belong to the genera Enterocytozoon and Encephalitozoon. To date, only five microsporidian species, including Encephalitozoon-like, have been found in hard ticks (Ixodidae) using microscopic methods, but no sequence data are available for them. Furthermore, no widespread screening for microsporidian-infected ticks based on DNA analysis has been carried out to date. Thus, in this study, we applied a recently developed DNA metabarcoding method for efficient microsporidian DNA identification to assess the role of ticks as potential vectors of microsporidian species causing diseases in humans. METHODS: In total, 1070 (493 juvenile and 577 adult) unfed host-seeking Ixodes ricinus ticks collected at urban parks in the city of Poznan, Poland, and 94 engorged tick females fed on dogs and cats were screened for microsporidian DNA. Microsporidians were detected by PCR amplification and sequencing of the hypervariable V5 region of 18S rRNA gene (18S profiling) using the microsporidian-specific primer set. Tick species were identified morphologically and confirmed by amplification and sequencing of the shortened fragment of cytochrome c oxidase subunit I gene (mini-COI). RESULTS: All collected ticks were unambiguously assigned to I. ricinus. Potentially zoonotic Encephalitozoon intestinalis was identified in three fed ticks (3.2%) collected from three different dogs. In eight unfed host-seeking ticks (0.8%), including three males (1.1%), two females (0.7%) and three nymphs (0.7%), the new microsporidian sequence representing a species belonging to the genus Endoreticulatus was identified. CONCLUSIONS: The lack of zoonotic microsporidians in host-seeking ticks suggests that I. ricinus is not involved in transmission of human-infecting microsporidians. Moreover, a very low occurrence of the other microsporidian species in both fed and host-seeking ticks implies that mechanisms exist to defend ticks against infection with these parasites.