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

Abstract INTRODUCTION In order to detect Trypanosoma cruzi and determine the genetic profiles of the parasite during the chronic phase of Chagas disease (ChD), parasitological and molecular diagnostic methods were used to assess the blood of 91 patients without specific prior treatment. METHODS Blood samples were collected from 68 patients with cardiac ChD and 23 patients with an indeterminate form of ChD, followed by evaluation using blood culture and polymerase chain reaction. T . cruzi isolates were genotyped using three different genetic markers. RESULTS: Blood culture was positive in 54.9% of all patients, among which 60.3% had the cardiac form of ChD, and 39.1% the indeterminate form of ChD. There were no significant differences in blood culture positivity among patients with cardiac and indeterminate forms. Additionally, patient age and clinical forms did not influence blood culture results. Polymerase chain reaction (PCR) was positive in 98.9% of patients, although comparisons between blood culture and PCR results showed that the two techniques did not agree. Forty-two T . cruzi stocks were isolated, and TcII was detected in 95.2% of isolates. Additionally, one isolate corresponded to TcIII or TcIV, and another corresponded to TcV or TcVI. CONCLUSIONS Blood culture and PCR were both effective for identifying T. cruzi using a single blood sample, and their association did not improve parasite detection. However, we were not able to establish an association between the clinical form of ChD and the genetic profile of the parasite

Whole genome sequencing of Trypanosoma cruzi field isolates reveals extensive genomic variability and complex aneuploidy patterns within TcII DTU

Abstract Background Trypanosoma cruzi, the etiologic agent of Chagas disease, is currently divided into six discrete typing units (DTUs), named TcI-TcVI. TcII is among the major DTUs enrolled in human infections in South America southern cone, where it is associated with severe cardiac and digestive symptoms. Despite the importance of TcII in Chagas disease epidemiology and pathology, so far, no genome-wide comparisons of the mitochondrial and nuclear genomes of TcII field isolates have been performed to track the variability and evolution of this DTU in endemic regions. Results In the present work, we have sequenced and compared the whole nuclear and mitochondrial genomes of seven TcII strains isolated from chagasic patients from the central and northeastern regions of Minas Gerais, Brazil, revealing an extensive genetic variability within this DTU. A comparison of the phylogeny based on the nuclear or mitochondrial genomes revealed that the majority of branches were shared by both sequences. The subtle divergences in the branches are probably consequence of mitochondrial introgression events between TcII strains. Two T. cruzi strains isolated from patients living in the central region of Minas Gerais, S15 and S162a, were clustered in the nuclear and mitochondrial phylogeny analysis. These two strains were isolated from the other five by the Espinhaço Mountains, a geographic barrier that could have restricted the traffic of insect vectors during T. cruzi evolution in the Minas Gerais state. Finally, the presence of aneuploidies was evaluated, revealing that all seven TcII strains have a different pattern of chromosomal duplication/loss. Conclusions Analysis of genomic variability and aneuploidies suggests that there is significant genomic variability within Minas Gerais TcII strains, which could be exploited by the parasite to allow rapid selection of favorable phenotypes. Also, the aneuploidy patterns vary among T. cruzi strains and does not correlate with the nuclear phylogeny, suggesting that chromosomal duplication/loss are recent and frequent events in the parasite evolution