Argas persicus and Carios vespertilionis Ticks Infesting Ducks, Domestic Fowls and Bats in Pakistan: First Report on Molecular Survey and Phylogenetic Position of Borrelia anserina

Argasid ticks have the vectorial potential for transmitting disease-causing pathogens to avian hosts, resulting in economic losses that may not be fully estimated. Borrelia species are the responsible agents of borreliosis in poultry, animals and humans. Our previous studies have reported a high prevalence of Argas persicus infesting domestic fowls in Khyber Pakhtunkhwa (KP), Pakistan. However, molecular screening and genetic characterization of Borrelia spp. in A. persicus have been neglected in Pakistan. In this study, we focused on the molecular epidemiology and genetic characterization of Borrelia spp. associated with A. persicus ticks infesting domestic fowls and ducks, and Carios vespertilionis infesting bats in selected districts of KP. Overall, 1818 ticks, including females (415: 23%), males (345: 19%), nymphs (475: 26%) and larvae (583: 32%), were collected from 27 locations in nine districts (Peshawar, Mardan, Swabi, Charsadda, Chitral, Lakki Marwat, Bannu, Bajaur and Hangu) from domestic fowls, ducks and their shelters, and bats. A subset of 197 ticks was selected for DNA extraction and PCR to amplify fragments of the cytochrome c oxidase (cox) gene for ticks and flagellin B (flaB) for the detection and genetic characterization of associated Borrelia spp. Among these, only Borrelia anserina DNA was detected in 40 ticks (27.2%) of different life stages, where highest prevalence was found in female ticks (18: 45%), followed by nymphs (12: 30%), larvae (7: 17.5%) and males (3: 7.5%). Tick infestation in shelters (1081: 77%) was higher than on hosts (323: 23%). The resultant cox amplicons of A. persicus showed 100% identity with the same species reported from Pakistan, China, Iran, Kenya, Kazakhstan, Algeria and Egypt and C. vespertilionis show 100% identity with the species reported from Pakistan, China, Japan, Kenya, Vietnam, Spain, Netherlands, the United Kingdom and Hungry, and clustered with the aforementioned species in the phylogenetic tree. The obtained Borrelia sequences showed 100% identity with B. anserina and revealed a close resemblance to the relapsing fever group and clustered in a monophyletic clade with B. anserina from India, Iran and Brazil in a phylogenetic tree. These results establish the first molecular characterization of B. anserina in A. persicus infesting domestic fowls and ducks in the region, as well as their shelters. To effectively control zoonotic consequences, country-wide surveillance research should be encouraged to screen soft ticks infesting various birds for associated pathogens.