SYBR Green qPCR Technique for the Detection of Trypanosoma cruzi in Açaí Pulp

Chagas disease, which is found widely in Latin America and has a great impact on public health, is caused by the parasite Trypanosoma cruzi. It is a neglected parasitic disease that urgently requires rapid diagnostic methods. The objective of this study was to develop a SYBR Green real-time quantitative polymerase chain reaction (qPCR) technique for the direct identification and quantification of T. cruzi from experimentally contaminated açai fruit samples. We used discrete typing units, TcI, containing 3.5 × 10⁴ cells/mL, to infect the pulp of the açai fruit. This was followed by DNA extraction using a standardized procedure. The DNA samples were quantified and amplified at specific time and temperature intervals. The specificity of the oligoinitiators used in the qPCR assays was estimated by calculating the primer dissociation curve (melting curve) along with a detection threshold using different concentrations of DNA. The method used here demonstrated good efficiency and precision for the detection and quantification of T. cruzi DNA, with a detection limit of 2.65 × 10^-14 g/μL DNA. The qPCR technique presented here could serve as an important tool for the diagnosis of T. cruzi parasites in açai.