Identification of blood-feeding sources in Panstrongylus, Psammolestes, Rhodnius, and Triatoma using Amplicon-based next-generation sequencing

Triatomines are hematophagous insects that play an important role as vectors of Trypanosoma cruzi, the causative agent of Chagas disease. These insects have adapted to multiple blood-feeding sources that can affect relevant aspects of their life cycle and interactions, thereby influencing parasitic transmission dynamics. We conducted a characterization of the feeding sources of individuals from the primary circulating triatomine genera in Colombia using amplicon-based next-generation sequencing [NGS].We collected 42 triatomines from different departments of Colombia. DNA was extracted from the gut. The presence of T. cruzi was identified using real-time PCR, and discrete typing units [DTUs] were determined by conventional PCR. For blood-feeding source identification, PCR products of the vertebrate 12S-rRNA gene were obtained through PCR and sequenced by Next Generation Sequencing [NGS]. Blood meal sources were inferred using blastn against a curated reference dataset containing the 12S-rRNA sequences belonging to vertebrates with distribution in Latin America that represent a potential feeding source for triatomine bugs. Mean and median comparison tests were performed to evaluate differences in triatomine blood-feeding sources, infection state, and geographical regions. Lastly, the Inverse Simpson diversity index was calculated.The overall frequency of T. cruzi infection was 83.3%. TcI was found as the most predominant DTU [65.7%]. Sixty-seven feeding sources were detected from the analyses of approximately 7 million reads. The predominant feeding source found was Homo sapiens [76.8%], followed by birds [10.5%], artiodactyls [4.4%], and non-human primates [3.9%]. There were differences among numerous feeding sources of triatomines of different species. The diversity of feeding sources also differed depending on the presence of T. cruzi.This is the first study to employ amplicon-based NGS of the 12S-rRNA gene to depict blood feeding sources of multiple triatomine species collected from different regions of Colombia. Our findings report a striking read diversity that has never been previously reported. This is a powerful approach to unravel transmission dynamics at micro geographical levels.