Molecular detection of Leishmania donovani, Dirofilaria sp. "hongkongensis," and Wolbachia spp. in the dog population from tribal settlements of Western Ghats, Kerala, India.

Leishmaniasis and dirofilariasis are significant emerging zoonotic diseases. This study aimed to assess the prevalence of single and co-infections involving Leishmania donovani, Dirofilaria sp. "hongkongensis", and Wolbachia spp. among dogs from the Kani tribal settlements of Western Ghats, Kerala. In July 2023, blood samples from 75 dogs were collected and analyzed using microscopy, serology, and molecular diagnostics. Molecular analysis using kDNA-PCR and ITS1 PCR identified leishmaniasis in 39 out of 75 dog samples (52.0% [95% CI, 40.2 to 63.7], with phylogenetic analysis confirming the species as L. donovani. Additionally, 19 out of 75 dog samples (25.3% [95% CI, 16.0 to 36.7]) produced a ~ 720 bp band for nematode-specific COI amplification, which was confirmed as Dirofilaria sp. "hongkongensis". Among the 19 Dirofilaria-positive samples, 14 (73.7% [95% CI, 48.8 to 90.9]) tested positive for Wolbachia using the wsp primer. Among the 75 dogs screened, 12 (16% [95% CI, 08.6 to 26.3]) were co-infected with Leishmania and Dirofilaria, while 9 (12% [95% CI, 05.6 to 21.6]) were co-infected with all three parasites: L. donovani, Dirofilaria sp. "hongkongensis", and Wolbachia spp. The findings suggest that Kerala is gradually becoming more vulnerable to zoonoses like leishmaniasis and dirofilariasis emphasizing the need for prompt intervention and prevention measures. These results underscore the critical need for continued development of multi-pathogen detection methods, not only in regions where multiple diseases are endemic but also in areas where such diseases may emerge.