High-resolution melting (HRM) of the cytochrome B gene: a powerful approach to identify blood-meal sources in Chagas disease Vectors

Abstract

Methods to determine blood-meal sources of hematophagous Triatominae bugs (Chagas disease vectors) are serological or based on PCR employing species-specific primers or heteroduplex analysis, but these are expensive, inaccurate, or problematic when the insect has fed on more than one species. To solve those problems, we developed a technique based on HRM analysis of the mitochondrial gene cytochrome B (Cyt b). This technique recognized 14 species involved in several ecoepidemiological cycles of the transmission of Trypanosoma cruzi and it was suitable with DNA extracted from intestinal content and feces 30 days after feeding, revealing a resolution power that can display mixed feedings. Field samples were analyzed showing blood meal sources corresponding to domestic, peridomiciliary and sylvatic cycles. The technique only requires a single pair of primers that amplify the Cyt b gene in vertebrates and no other standardization, making it quick, easy, relatively inexpensive, and highly accurate.

Figure 1. Tm distribution of 14 genotypes included in the study.

Tm: melting temperature, Sp: species.

Figure 2. Dissociation curve of 13 of the 14 genotypes included in the study.

Figure 3. Field sample showing mixed feeding including opossum and human with their respective controls.