Triatoma venosa and Panstrongylus geniculatus challenge the certification of interruption of vectorial Trypanosoma cruzi transmission by Rhodnius prolixus in eastern Colombia

Reactivation of Trypanosoma cruzi transmission by native vectors with different domiciliation capabilities is a major concern for Chagas disease control programs. T. cruzi transmission via intra-domestic Rhodnius prolixus was certified as interrupted by the Pan American Health Organization in Miraflores municipality (Boyacá, Colombia) in 2019. However, Triatoma venosa, a native vector infected with T. cruzi has been increasingly found inside human dwellings across rural areas. In this study, the aim was to describe the eco-epidemiological aspects of T. cruzi transmission in the rural area of Miraflores. For this, we designed a comprehensive, multi-faceted study in 6 rural villages and performed: (i) A cross-sectional serological and molecular study enrolling 155 people and 58 domestic dogs living within 80 households, (ii) a domestic entomological survey, (iii) a determination of the natural infection and blood meal source in collected triatomine bugs, and (iv) an evaluation of synanthropic mammal infection by parasitological and molecular tools. The T. cruzi seroprevalence rates in humans and dogs were 9.03% (14/155) and 22.4% (13/58), respectively. Most infected humans were adults between the ages of 55 and 85 years old. No evidence of T. cruzi DNA was found using qPCR in human blood samples, but we found high parasitemia levels in the infected dogs. In total, 38 triatomine bugs were collected inside dwellings and peridomestic areas: 68.4% (26/38) Triatoma venosa, 29% (11/38) Panstrongylus geniculatus, and 2.6% (1/38) P. rufotuberculatus. Natural infection prevalence was 88% (22/25) for T. venosa, 100% (12/12) for P. geniculatus, and 100% (1/1) P. rufotuberculatus: only TcI was found. No evidence of R. prolixus was found in the area. Two feeding sources were identified in T. venosa (humans and cats), while P. geniculatus fed on cows and bats. Lastly, seven D. marsupialis were captured in peridomestic areas, three were infected with T. cruzi (TcI). The results suggest the existence of T. cruzi transmission cycle between triatomines, dogs, and opossums representing a risk of infection for the human population in rural areas of Miraflores. Despite PAHO declaring Miraflores municipality, Colombia an area of T. cruzi transmission interruption in 2019, this study documents evidence of a secondary vector establishing in domestic settings. T. venosa entomological surveillance is warranted to evaluate prospective human transmission risk in an otherwise 'no-risk' perceived Chagas disease region.