Fig 1.
Flowchart used for classification of Trypanosoma cruzi DTUs.
Algorithmic showing the PCRs targets and PCR products size (bp). The intergenic region of Spliced Leader (SL-IRac) [UTCC and TCac primers] was used to distinguish between TcI (150 bp), TcII, V or VI (157 bp) and TcIII or TcIV (200 bp); SL-IR I and II [TCC, TC1 and TC2 primers], to distinguish between TcI (350 bp), TcII, TcV and TcVI (300 bp) and TcIII and TcIV (not amplified)]; D7 domain of the 24Sα ribosomal RNA gene [Heminested PCR: D75 and D76 (first round) and D76 and D71 (second round) were used to distinguish between TcII and TcVI (140 bp), TcIII (125 bp), TcIV (140/145 bp) and TcV (125 or 125 + 140 bp)]; A10 nuclear fragment [Heminested PCR: Pr1 and P6 (first round) and Pr1 and Pr3 (second round), to differentiate TcII (690/580 bp) from TcVI (630/525 bp)].
Table 1.
Demographic, epidemiological, and clinical data of the group of non-etiologically treated Chagas disease patients in Pernambuco State, Brazil.
Fig 2.
Maps showing the geographical location of the different mesoregions of Pernambuco/Brazil.
Map showing the location of the state of Pernambuco on maps of Brazil and South America, and the location of the different mesoregions. This map was created using QGIS 3.34 ‘Prizren’ version software and cartographic bases obtained from the Brazilian Institute of Geography and Statistics (https://ibge.gov.br/). Source: Brazilian Institute of Geography and Statistics (IBGE). Digital Municipal Grid (MMD). Licensed under CC BY 4.0. Municipality border shape available from https://www.ibge.gov.br/geociencias/organizacao-do-territorio/malhas-territoriais/15774-malhas.html. Terms of use available from https://biblioteca.ibge.gov.br/visualizacao/livros/liv102169.pdf.
Fig 3.
Representative 2% agarose gels for conventional PCR for Chagas disease diagnosis and β-globin gene. A. Representative amplification of Trypanosoma cruzi kDNA. Arrow indicates a band at approximately 330 bp (T. cruzi kDNA). MW, Molecular weight; Lane 1, NTC, no template control; Lanes 2-18 correspond to clinical samples of CD patients; PC +, positive control (TcII strain). B. Representative amplification of β-globin gene fragment of 110 bp in samples of serology-positive CD patients. Arrow indicates a band at approximately 110 bp (β-globin gene). MW, Molecular weight; Lanes 1 to 5 correspond to kDNA-negative clinical samples of CD patients.
Table 2.
Samples positive for T. cruzi kDNA and genotyped for DTU in clinically classified Chagas disease patients in the State of Pernambuco, Brazil.
Fig 4.
Distribution of patients with Chagas disease diagnosed by conventional PCR for Trypanosoma cruzi kDNA.
A. Percentages of total patients diagnosed for CD (Total) and positive for T. cruzi kDNA (kDNA+) in the mesoregions of Pernambuco/Brazil. Map of Pernambuco shows: Brown, Sertão (S); Yellow, Agreste (A); Green, Zona da Mata (ZM); Red, Metropolitan Region (MR). Different shades of a color indicate geographically considered sub-areas within a mesoregion. B. Percentage of CD patients diagnosed by PCR for T. cruzi kDNA according to the different clinical forms of CD. C. Percentage of patients positive for T. cruzi kDNA in the different clinical forms of CD. D. Percentage of PCR positivity for T. cruzi kDNA in each clinical group of CD patients. The map was obtained from https://www.ibge.gov.br/geociencias/organizacao-do-territorio/malhas-territoriais/15774-malhas.html.
Table 3.
Samples of DC patients genotyped for DTU distributed by attributes local of birth in geographical mesoregions of the State of Pernambuco, Brazil.
Fig 5.
Agarose gels for different targets of Trypanosoma cruzi in blood samples of Chagas disease patients.
Agarose gels with products obtained after conventional PCR for the different targets of T. cruzi for the classification of DTUs in blood samples of CD patients. Abbreviations: MW, molecular weight marker (100 bp); Lanes 1-16, clinical samples of CD patients; S, Strain; S1-Dm28c (TcI); S2-CL Brener (TcVI); S3-INPA 4167 (TcIV); S4-Y (TcII); S5-INPA 3663 (TcIII); S6-LL014 (TcV); bp, base pairs.
Fig 6.
Maps showing the spatial distribution of clinically classified CD patients and DTUs of Trypanosoma cruzi in the different mesoregions of Pernambuco/Brazil. A. Map, geographic distribution of samples from patients classified into T. cruzi DTUs TcI-TcVI. B. Map, geographic distribution of patients classified according to clinical forms of CD and submitted to genotypic characterization of T. cruzi. This map was created using QGIS 3.34 ‘Prizren’ version software and cartographic bases obtained from the Brazilian Institute of Geography and Statistics https://www.ibge.gov.br/geociencias/organizacao-do-territorio/malhas-territoriais/15774-malhas.html).
Table 4.
Samples genotyped for DTU in clinically classified Chagas disease patients in the State of Pernambuco, Brazil.
Table 5.
Distribution of samples of patient classified in clinical forms of Chagas disease according to genotyped DTU in the mesoregions of Pernambuco (PE), Brazil.