Molecular detection of Trypanosoma lewisi: association between Xenopsylla cheopis and Rattus spp. in densely populated residential areas.

Trypanosoma lewisi is a flagellated hemoprotozoan that primarily infects rats (Rattus spp.), with Xenopsylla cheopis acting as its cyclical vector. Although typically host-specific, occasional zoonotic transmission to humans (zoonosis) has raised One Health concerns, particularly in densely populated areas with poor sanitation. This study aimed to determine the infection rate of T. lewisi in both X. cheopis and rats using molecular approaches. Rats were trapped in residential areas of Kampung Mandar and Lateng Villages in Banyuwangi, East Java, Indonesia. Fleas were obtained by combing the rats' fur, and blood samples were subsequently collected intracardially. Identification of X. cheopis was performed using stereomicroscopy, scanning electron microscopy, and PCR employing specific primers Xchvgsc F and Xchvgsc R. Detection of T. lewisi was conducted via PCR using LEW1S/LEW1R primers. Sequencing and phylogenetic analyses were performed using GAPDH and SSU gene markers. A total of 152 rats were captured, consisting of Rattus norvegicus (53.29%) and R. tanezumi (46.71%). The prevalence of T. lewisi infection was 23.68% in rats and 71.91% in fleas. Among the 89 flea-infested rats, 46 (51.69%) were negative for T. lewisi in their blood but carried T. lewisi-positive X. cheopis fleas. Phylogenetic analysis revealed that T. lewisi from rats and fleas clustered within the same lineage, indicating genetic similarity between parasites detected in the two hosts. These findings demonstrate the presence of genetically similar T. lewisi parasites in rats and naturally infected X. cheopis fleas, suggesting that the parasite may circulate between these hosts within rodent populations. The present study also underscores the importance of integrated rodent and flea surveillance for understanding the epidemiology and circulation of T. lewisi in rodent populations.