Probing Population Dynamics of Trypanosoma cruzi during Progression of the Chronic Phase in Chagasic Patients

Abstract

Our research aimed to characterize the genetic profiles of 102 Trypanosoma cruzi isolates recently obtained from 44 chronic chagasic patients from different regions of the states of Minas Gerais and Goiás in Brazil. At least two isolates were obtained from each patient at different times in order to study the parasite population dynamics during disease progression in the chronic phase. The isolates were characterized molecularly by genotyping the 3 region of the 24S␣ rRNA, the mitochondrial cytochrome oxidase subunit 2 (COII) gene, and the intergenic region of the spliced leader intergenic region (SL-IR) gene. Seventy-seven isolates were analyzed for nine microsatellite loci. The data presented here show a strong correlation between the T. cruzi lineage II (T. cruzi II) and human infection in these regions of Brazil. Interestingly, isolates from two patients were initially characterized (by rRNA genotyping) as T. cruzi I and hybrid strains, but subsequent analyses of the COII and SL-IR genes confirmed that those isolates belonged to T. cruzi III and a hybrid group, respectively. Our results confirm the risk of misclassifying T. cruzi isolates on the basis of analysis of a single molecular marker. The microsatellite profiles showed that different isolates obtained from the same patient were genetically identical and monoclonal. Exceptions were observed for T. cruzi isolates from two patients who presented differences for the SCLE11 locus and also from two other patients who showed amplification of three peaks for a microsatellite locus (TcAAAT6), implying that they were multiclonal. On the basis of the findings of the studies described here, we were not able to establish a correlation between the clinical forms of Chagas' disease and the genetic profiles of the T. cruzi isolates.

Combined parasitological and molecular-based diagnostic tools improve the detection of Trypanosoma cruzi in single peripheral blood samples from patients with Chagas disease

“…Importantly, our data were consistent with previous studies showing that DTU II was associated with human infection in the State of Minas Gerais 32,57,58 . The methodologies applied for genetic characterization of isolates of different clinical forms may be a limiting factor for typing T. cruzi populations since the analysis was based on a single marker, leading to potential misinterpretations 59 .…”

“…Our results showed that using two molecular markers was not sufficient for typing of T. cruzi isolates. These limitations were also observed by other authors 32,[60][61][62][63] . However, the use of three genetic markers with different evolutionary rates was able to ensure the correct typing of T. cruzi isolates.…”

“…Findings using lowstringency single specific primer PCR and rDNA 24Sα markers showed TcII in all isolates from patients with indeterminate, cardiac, digestive, and cardiodigestive forms 58 . Although TcII was identified in most T. cruzi isolates, no correlation with clinical forms was observed using rDNA 24Sα, COII, and SL-IR markers and polymorphisms in microsatellite analysis 32 . TcII has been reported as the primary cause of acute ChD and is the typical genotype detected in patients with cardiac and digestive forms of ChD, confirming the association of this DTU with human infection in Brazil and Argentina 57,63 .…”

“…Trypanonoma cruzi isolates were typed using three different parasite genomic sequences according to proposed protocols 32 , as follows: 24Sα ribosomal (rRNA) gene D7 domain 33 , mitochondrial cytochrome oxidase subunit 2 gene (COII) 34 , and the spliced leader genes intergenic region 35 , as markers of six DTUs 18 , TcI (Col1.7G2 Colombiana clone), TcII (JG), TcIII (222), TcIV (CAN III clone), TcV (3253 Lages-Silva et al: unpublished data), and TcVI (CL) were used as reference strains and DTU controls 18,34 .…”

Characteristics of Triatomine infestation and natural Trypanosoma cruzi infection in the State of Rio Grande do Norte, Brazil

“…Trypanosoma cruzi isolates were typed using three different parasite genomic sequences as reported (39) , with the 24Sα ribosomal (rRNA) gene D7 domain (40) , mitochondrial cytochrome oxidase subunit 2 gene (COII) (41) , and the spliced leader genes intergenic region (42) as markers for six discrete typing units (DTUs) (43) , using T. cruzi reference strains (15) (44) (45) and clones (46) (47) as DTU controls.…”