Trypanosoma cruzi: Evaluation of PCR as a Laboratory Tool to Follow up the Evolution of Parasite Load

Abstract

Background: The study evaluated qualitative PCR, primers 121-122 as a tool to follow up evolution parasite load of Trypanosoma cruzi. Methods: The study was conducted at the State University of Maringa, in 2015. Step 1, dilutions 1/10 were performed from T. cruzi-Y strain to obtain preparations of 50,000-0.05 parasites/mL from which DNA were extracted, quantified, and amplified. Step 2, the extracted DNA in the dilutions 5-0.05 parasites/mL was re-diluted 1/10, 1/100, 1/1000, quantified, and amplified. Polyacrylamide gels were photographed and thicknesses of the 330 bp kDNA fragments were measured. Results: Step 1, in the dilutions 50,000-50 parasites/mL kDNA fragments had same thickness and, dilutions 5-0.05 parasites/mL showed progressive decrease in thicknesses and staining intensity of the 330 bp fragments. Step 2, demonstrated that dilutions of five (re-dilutions 1/10 and 1/100) and 0.5 (1/10) parasites/mL produced similar thicknesses of the 330 bp fragments obtained in Step 1. However, very dilute DNA samples make difficult to reproduce the fragments thicknesses. Conclusion: PCR, despite its limitations, was able to detect progressive decrease in thicknesses/staining intensity of kDNA fragments in the dilutions 5-0.05 parasites/mL. Hence, has the potential to be used to follow-up evolution of parasite load, not by quantifying the number of parasites, but by dynamic evolution of the fragments thicknesses during etiological treatment.

Keywords

• Trypanosoma cruzi
• Parasite load
• PCR
• kDNA fragments