Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay

Background Chagas disease, caused by Trypanosoma cruzi, is a geographically widespread anthropozoonosis that is considered a major public health problem in Latin America. Because this parasite presents high genetic variability, a nomenclature has been adopted to classify the parasite into six discre...

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Autores Principales: Higuera, Sonia L, Guhl, Felipe, Ramírez, Juan David
Formato: Artículo (Article)
Lenguaje:Inglés (English)
Publicado: BioMed Central 2013
Materias:
DTU
Acceso en línea:https://repository.urosario.edu.co/handle/10336/27838
https://doi.org/10.1186/1756-3305-6-112
id ir-10336-27838
recordtype dspace
spelling ir-10336-278382021-01-21T08:15:12Z Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay Identificación de unidades de tipificación discreta (DTU) de Trypanosoma cruzi mediante la implementación de un ensayo de genotipado de fusión de alta resolución (HRM) Higuera, Sonia L Guhl, Felipe Ramírez, Juan David Chagas disease Trypanosoma cruzi DTU High-resolution melting Genotyping Background Chagas disease, caused by Trypanosoma cruzi, is a geographically widespread anthropozoonosis that is considered a major public health problem in Latin America. Because this parasite presents high genetic variability, a nomenclature has been adopted to classify the parasite into six discrete typing units (DTUs): TcI, TcII, TcIII, TcIV, TcV, and TcVI, which present different eco-epidemiological, clinical, and geographic associations. Currently, the available genotyping methods present a series of drawbacks that implies the need for developing new methods for characterizing T. cruzi DTU’s. The aim of this work was to genotype reference populations from T. cruzi by means of a High-Resolution Melting (HRM) genotyping assay. To genotype the DTUs of 38 strains and 14 reference clones of T. cruzi from diverse sources, real-time PCR (qPCR) was coupled to high-resolution melting (HRM) based on the amplification of two molecular markers—the divergent domain of the 24 s? rRNA gene and the intergenic region of the mini-exon gene. Findings Amplification of the mini-exon gene allowed the genotyping of three distinct groups: TcI, TcII- TcIV- TcV, and TcIII-TcVI, while amplification of the 24s? gene generated non-overlapping melting temperature ranges for each DTU that were used to identify the groups in the six existing DTUs of Trypanosoma cruzi. Conclusions The proposed genotyping assay allowed discrimination of the six genetic groups by obtaining specific melting curves for each DTU. The application of this technique is proposed because of its specificity, sensitivity, high performance, and low cost compared with other previously described characterization methods. 2013-04-20 2020-08-19T14:44:10Z info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion EISSN: 1756-3305 https://repository.urosario.edu.co/handle/10336/27838 https://doi.org/10.1186/1756-3305-6-112 eng info:eu-repo/semantics/openAccess application/pdf BioMed Central Parasites and Vectors
institution EdocUR - Universidad del Rosario
collection DSpace
language Inglés (English)
topic Chagas disease
Trypanosoma cruzi
DTU
High-resolution melting
Genotyping
spellingShingle Chagas disease
Trypanosoma cruzi
DTU
High-resolution melting
Genotyping
Higuera, Sonia L
Guhl, Felipe
Ramírez, Juan David
Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
description Background Chagas disease, caused by Trypanosoma cruzi, is a geographically widespread anthropozoonosis that is considered a major public health problem in Latin America. Because this parasite presents high genetic variability, a nomenclature has been adopted to classify the parasite into six discrete typing units (DTUs): TcI, TcII, TcIII, TcIV, TcV, and TcVI, which present different eco-epidemiological, clinical, and geographic associations. Currently, the available genotyping methods present a series of drawbacks that implies the need for developing new methods for characterizing T. cruzi DTU’s. The aim of this work was to genotype reference populations from T. cruzi by means of a High-Resolution Melting (HRM) genotyping assay. To genotype the DTUs of 38 strains and 14 reference clones of T. cruzi from diverse sources, real-time PCR (qPCR) was coupled to high-resolution melting (HRM) based on the amplification of two molecular markers—the divergent domain of the 24 s? rRNA gene and the intergenic region of the mini-exon gene. Findings Amplification of the mini-exon gene allowed the genotyping of three distinct groups: TcI, TcII- TcIV- TcV, and TcIII-TcVI, while amplification of the 24s? gene generated non-overlapping melting temperature ranges for each DTU that were used to identify the groups in the six existing DTUs of Trypanosoma cruzi. Conclusions The proposed genotyping assay allowed discrimination of the six genetic groups by obtaining specific melting curves for each DTU. The application of this technique is proposed because of its specificity, sensitivity, high performance, and low cost compared with other previously described characterization methods.
format Artículo (Article)
author Higuera, Sonia L
Guhl, Felipe
Ramírez, Juan David
author_facet Higuera, Sonia L
Guhl, Felipe
Ramírez, Juan David
author_sort Higuera, Sonia L
title Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
title_short Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
title_full Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
title_fullStr Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
title_full_unstemmed Identification of Trypanosoma cruzi Discrete Typing Units (DTUs) through the implementation of a High-Resolution Melting (HRM) genotyping assay
title_sort identification of trypanosoma cruzi discrete typing units (dtus) through the implementation of a high-resolution melting (hrm) genotyping assay
publisher BioMed Central
publishDate 2013
url https://repository.urosario.edu.co/handle/10336/27838
https://doi.org/10.1186/1756-3305-6-112
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