Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples
- Autores
- Bontempi, Iván; Bizai, María Laura; Ortiz, Sylvia; Manattini, Silvia; Fabbro, Diana Lucrecia; Solari, Aldo; Diez, Cristina Noemí
- Año de publicación
- 2016
- Idioma
- inglés
- Tipo de recurso
- artículo
- Estado
- versión publicada
- Descripción
- Different DNA markers to genotype Trypanosoma cruzi are now available. However, due to the low quantity of parasites present in biological samples, DNA markers with high copy number like kinetoplast minicircles are needed. The aim of this study was to complete a DNA assay called minicircle lineage specific-PCR (MLS-PCR) previously developed to genotype the T. cruzi DTUs TcV and TcVI, in order to genotype DTUs TcI and TcII and to improve TcVI detection. We screened kinetoplast minicircle hypervariable sequences from cloned PCR products from reference strains belonging to the mentioned DTUs using specific kDNA probes. With the four highly specific sequences selected, we designed primers to be used in the MLS-PCR to directly genotype T. cruzi from biological samples. High specificity and sensitivity were obtained when we evaluated the new approach for TcI, TcII, TcV and TcVI genotyping in twenty two T. cruzi reference strains. Afterward, we compared it with hybridization tests using specific kDNA probes in 32 blood samples from chronic chagasic patients from North Eastern Argentina. With both tests we were able to genotype 94% of the samples and the concordance between them was very good (kappa = 0.855). The most frequent T. cruzi DTUs detected were TcV and TcVI, followed by TcII and much lower TcI. A unique T. cruzi DTU was detected in 18 samples meantime more than one in the remaining; being TcV and TcVI the most frequent association. A high percentage of mixed detections were obtained with both assays and its impact was discussed.
Fil: Bontempi, Iván. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Tecnología Inmunológica; Argentina
Fil: Bizai, María Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; Argentina
Fil: Ortiz, Sylvia. Universidad de Chile; Chile
Fil: Manattini, Silvia. Provincia de Santa Fe. Hospital Central de Reconquista; Argentina
Fil: Fabbro, Diana Lucrecia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; Argentina
Fil: Solari, Aldo. Universidad de Chile; Chile
Fil: Diez, Cristina Noemí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas; Argentina - Materia
-
Dtu
Genotype
Hybridization
Mixed Infections
Mls-Pcr
Trypanosoma Cruzi - Nivel de accesibilidad
- acceso abierto
- Condiciones de uso
- https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
- Repositorio
- Institución
- Consejo Nacional de Investigaciones Científicas y Técnicas
- OAI Identificador
- oai:ri.conicet.gov.ar:11336/78249
Ver los metadatos del registro completo
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Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samplesBontempi, IvánBizai, María LauraOrtiz, SylviaManattini, SilviaFabbro, Diana LucreciaSolari, AldoDiez, Cristina NoemíDtuGenotypeHybridizationMixed InfectionsMls-PcrTrypanosoma Cruzihttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Different DNA markers to genotype Trypanosoma cruzi are now available. However, due to the low quantity of parasites present in biological samples, DNA markers with high copy number like kinetoplast minicircles are needed. The aim of this study was to complete a DNA assay called minicircle lineage specific-PCR (MLS-PCR) previously developed to genotype the T. cruzi DTUs TcV and TcVI, in order to genotype DTUs TcI and TcII and to improve TcVI detection. We screened kinetoplast minicircle hypervariable sequences from cloned PCR products from reference strains belonging to the mentioned DTUs using specific kDNA probes. With the four highly specific sequences selected, we designed primers to be used in the MLS-PCR to directly genotype T. cruzi from biological samples. High specificity and sensitivity were obtained when we evaluated the new approach for TcI, TcII, TcV and TcVI genotyping in twenty two T. cruzi reference strains. Afterward, we compared it with hybridization tests using specific kDNA probes in 32 blood samples from chronic chagasic patients from North Eastern Argentina. With both tests we were able to genotype 94% of the samples and the concordance between them was very good (kappa = 0.855). The most frequent T. cruzi DTUs detected were TcV and TcVI, followed by TcII and much lower TcI. A unique T. cruzi DTU was detected in 18 samples meantime more than one in the remaining; being TcV and TcVI the most frequent association. A high percentage of mixed detections were obtained with both assays and its impact was discussed.Fil: Bontempi, Iván. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Tecnología Inmunológica; ArgentinaFil: Bizai, María Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; ArgentinaFil: Ortiz, Sylvia. Universidad de Chile; ChileFil: Manattini, Silvia. Provincia de Santa Fe. Hospital Central de Reconquista; ArgentinaFil: Fabbro, Diana Lucrecia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; ArgentinaFil: Solari, Aldo. Universidad de Chile; ChileFil: Diez, Cristina Noemí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas; ArgentinaElsevier Science2016-09info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/78249Bontempi, Iván; Bizai, María Laura; Ortiz, Sylvia; Manattini, Silvia; Fabbro, Diana Lucrecia; et al.; Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples; Elsevier Science; Infection, Genetics and Evolution; 43; 9-2016; 123-1291567-1348CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S1567134816301988info:eu-repo/semantics/altIdentifier/doi/10.1016/j.meegid.2016.05.026info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:50:54Zoai:ri.conicet.gov.ar:11336/78249instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 09:50:54.654CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse |
dc.title.none.fl_str_mv |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
title |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
spellingShingle |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples Bontempi, Iván Dtu Genotype Hybridization Mixed Infections Mls-Pcr Trypanosoma Cruzi |
title_short |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
title_full |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
title_fullStr |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
title_full_unstemmed |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
title_sort |
Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples |
dc.creator.none.fl_str_mv |
Bontempi, Iván Bizai, María Laura Ortiz, Sylvia Manattini, Silvia Fabbro, Diana Lucrecia Solari, Aldo Diez, Cristina Noemí |
author |
Bontempi, Iván |
author_facet |
Bontempi, Iván Bizai, María Laura Ortiz, Sylvia Manattini, Silvia Fabbro, Diana Lucrecia Solari, Aldo Diez, Cristina Noemí |
author_role |
author |
author2 |
Bizai, María Laura Ortiz, Sylvia Manattini, Silvia Fabbro, Diana Lucrecia Solari, Aldo Diez, Cristina Noemí |
author2_role |
author author author author author author |
dc.subject.none.fl_str_mv |
Dtu Genotype Hybridization Mixed Infections Mls-Pcr Trypanosoma Cruzi |
topic |
Dtu Genotype Hybridization Mixed Infections Mls-Pcr Trypanosoma Cruzi |
purl_subject.fl_str_mv |
https://purl.org/becyt/ford/1.6 https://purl.org/becyt/ford/1 |
dc.description.none.fl_txt_mv |
Different DNA markers to genotype Trypanosoma cruzi are now available. However, due to the low quantity of parasites present in biological samples, DNA markers with high copy number like kinetoplast minicircles are needed. The aim of this study was to complete a DNA assay called minicircle lineage specific-PCR (MLS-PCR) previously developed to genotype the T. cruzi DTUs TcV and TcVI, in order to genotype DTUs TcI and TcII and to improve TcVI detection. We screened kinetoplast minicircle hypervariable sequences from cloned PCR products from reference strains belonging to the mentioned DTUs using specific kDNA probes. With the four highly specific sequences selected, we designed primers to be used in the MLS-PCR to directly genotype T. cruzi from biological samples. High specificity and sensitivity were obtained when we evaluated the new approach for TcI, TcII, TcV and TcVI genotyping in twenty two T. cruzi reference strains. Afterward, we compared it with hybridization tests using specific kDNA probes in 32 blood samples from chronic chagasic patients from North Eastern Argentina. With both tests we were able to genotype 94% of the samples and the concordance between them was very good (kappa = 0.855). The most frequent T. cruzi DTUs detected were TcV and TcVI, followed by TcII and much lower TcI. A unique T. cruzi DTU was detected in 18 samples meantime more than one in the remaining; being TcV and TcVI the most frequent association. A high percentage of mixed detections were obtained with both assays and its impact was discussed. Fil: Bontempi, Iván. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Laboratorio de Tecnología Inmunológica; Argentina Fil: Bizai, María Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; Argentina Fil: Ortiz, Sylvia. Universidad de Chile; Chile Fil: Manattini, Silvia. Provincia de Santa Fe. Hospital Central de Reconquista; Argentina Fil: Fabbro, Diana Lucrecia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Centro de Investigaciones sobre Endemias Nacionales; Argentina Fil: Solari, Aldo. Universidad de Chile; Chile Fil: Diez, Cristina Noemí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; Argentina. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas; Argentina |
description |
Different DNA markers to genotype Trypanosoma cruzi are now available. However, due to the low quantity of parasites present in biological samples, DNA markers with high copy number like kinetoplast minicircles are needed. The aim of this study was to complete a DNA assay called minicircle lineage specific-PCR (MLS-PCR) previously developed to genotype the T. cruzi DTUs TcV and TcVI, in order to genotype DTUs TcI and TcII and to improve TcVI detection. We screened kinetoplast minicircle hypervariable sequences from cloned PCR products from reference strains belonging to the mentioned DTUs using specific kDNA probes. With the four highly specific sequences selected, we designed primers to be used in the MLS-PCR to directly genotype T. cruzi from biological samples. High specificity and sensitivity were obtained when we evaluated the new approach for TcI, TcII, TcV and TcVI genotyping in twenty two T. cruzi reference strains. Afterward, we compared it with hybridization tests using specific kDNA probes in 32 blood samples from chronic chagasic patients from North Eastern Argentina. With both tests we were able to genotype 94% of the samples and the concordance between them was very good (kappa = 0.855). The most frequent T. cruzi DTUs detected were TcV and TcVI, followed by TcII and much lower TcI. A unique T. cruzi DTU was detected in 18 samples meantime more than one in the remaining; being TcV and TcVI the most frequent association. A high percentage of mixed detections were obtained with both assays and its impact was discussed. |
publishDate |
2016 |
dc.date.none.fl_str_mv |
2016-09 |
dc.type.none.fl_str_mv |
info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://purl.org/coar/resource_type/c_6501 info:ar-repo/semantics/articulo |
format |
article |
status_str |
publishedVersion |
dc.identifier.none.fl_str_mv |
http://hdl.handle.net/11336/78249 Bontempi, Iván; Bizai, María Laura; Ortiz, Sylvia; Manattini, Silvia; Fabbro, Diana Lucrecia; et al.; Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples; Elsevier Science; Infection, Genetics and Evolution; 43; 9-2016; 123-129 1567-1348 CONICET Digital CONICET |
url |
http://hdl.handle.net/11336/78249 |
identifier_str_mv |
Bontempi, Iván; Bizai, María Laura; Ortiz, Sylvia; Manattini, Silvia; Fabbro, Diana Lucrecia; et al.; Simple methodology to directly genotype Trypanosoma cruzi discrete typing units in single and mixed infections from human blood samples; Elsevier Science; Infection, Genetics and Evolution; 43; 9-2016; 123-129 1567-1348 CONICET Digital CONICET |
dc.language.none.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
info:eu-repo/semantics/altIdentifier/url/https://www.sciencedirect.com/science/article/pii/S1567134816301988 info:eu-repo/semantics/altIdentifier/doi/10.1016/j.meegid.2016.05.026 |
dc.rights.none.fl_str_mv |
info:eu-repo/semantics/openAccess https://creativecommons.org/licenses/by-nc-sa/2.5/ar/ |
eu_rights_str_mv |
openAccess |
rights_invalid_str_mv |
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/ |
dc.format.none.fl_str_mv |
application/pdf application/pdf application/pdf application/pdf application/pdf |
dc.publisher.none.fl_str_mv |
Elsevier Science |
publisher.none.fl_str_mv |
Elsevier Science |
dc.source.none.fl_str_mv |
reponame:CONICET Digital (CONICET) instname:Consejo Nacional de Investigaciones Científicas y Técnicas |
reponame_str |
CONICET Digital (CONICET) |
collection |
CONICET Digital (CONICET) |
instname_str |
Consejo Nacional de Investigaciones Científicas y Técnicas |
repository.name.fl_str_mv |
CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas |
repository.mail.fl_str_mv |
dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar |
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score |
13.070432 |