Ticks are important vectors of rickettsial pathogens, particularly in regions where livestock, wildlife, and humans overlap. Despite increasing reports of rickettsioses in India, molecular evidence of Rickettsia species in questing ticks from the Western Ghats of Kerala remains scarce. Questing ticks were collected from forest-edge and fringe habitats in Wayanad, Idukki, and Thrissur districts using standardized dragging. Species were identified morphologically, pooled, and screened for Rickettsia DNA by PCR targeting gltA and ompA genes. Amplicons were sequenced, and phylogenetic analysis was performed using reference sequences. Among 237 ticks belonging to six species, 24 % (12/50 pools) were PCR-positive for Rickettsia. The highest prevalence was observed in Haemaphysalis spinigera (50 %), followed by Haemaphysalis bispinosa (30 %) and Haemaphysalis turturis (9.5 %). Sequence analysis revealed the presence of Rickettsia massiliae and Rickettsia vini like species. Importantly, sequences closely related to “Candidatus Rickettsia jingxinensis” were detected for the first time in India, in H. turturis. Several isolates showed genetic divergence from known strains, suggesting the circulation of uncharacterized Rickettsia species or strains in tick population. This study provides the first molecular evidence of “Ca. R. jingxinensis” in H. turturis from India and demonstrates a high prevalence of Rickettsia spp. in questing ticks of Kerala. Given the pathogenic potential of some of these species, enhanced surveillance and incorporation of rickettsioses into differential diagnoses of febrile illnesses are needed in South India.
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