DNA barcoding identifies hard tick (Acari: Ixodidae) species infesting domesticated animals in Tamil Nadu, South...

The diverse and vast ecological landscapes of India support a rich diversity of ticks, many of which are known vectors of a wide range of pathogens. Accurate identification of tick species is critical for incriminating specific vectors involved in pathogen transmission. The present study aims to generate DNA barcodes using molecular markers for the identification of tick fauna from Tamil Nadu, southern India, where molecular taxonomic studies remain limited. A total of 57 specimens representing 12 different species were subjected to DNA barcoding using cytochrome c oxidase subunit I (COI) and internal transcribed spacer 2 (ITS2). Polymerase chain reaction (PCR) assays targeting the COI gene were successful in 7 species from four genera: Haemaphysalis, Hyalomma, Rhipicephalus, and Amblyomma. Similarly, 6 species from two genera, Rhipicephalus and Haemaphysalis, were successfully amplified using the ITS2 gene marker. Further analysis of inter-species diversity based on COI markers across eight species revealed better resolution compared to ITS2 markers. Inter-species distances of 16%, 15%, 14%, and 13% were recorded among four Rhipicephalus species using both markers, with the highest genetic divergence (16%) observed between R. microplus and R. sanguineus. The lowest inter-species divergence was 6% (COI) and 1% (ITS2), observed between R. microplus and R. annulatus. To our knowledge, this study provides the first DNA barcode records based on COI for Hyalomma anatolicum and Amblyomma integrum, and based on ITS2 for Rhipicephalus annulatus and Haemaphysalis intermedia from India. In conclusion, for four Rhipicephalus and two Haemaphysalis species, the dual-marker barcoding approach effectively complements conventional identification methods by resolving ambiguities arising from morphological similarities among tick species in this region.