Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus fam...

Autores: Muñoz Calderon, Arturo Alejandro; Lucero, Raul Horacio; Brusés, Bettina Laura; Formichelli, Laura Belén; Schijman, Alejandro Gabriel
Año de publicación: 2019
Idioma: inglés
Tipo de recurso: documento de conferencia
Estado: versión publicada
Descripción: The Trypanosomiasis caused by Trypanosoma evansi affects a wide diversity of mammals being zoonotic potential in man, with a case reported in 2005 in India. This haemoflagellate protozoan can parasitize most domestic mammals, being horses, dogs, and cattle the most affected species. Diagnostic tools for this parasitic infection are scarce, even though this trypanosomiasis can be very lethal if the animals are not treated. This work reports the development of a multi-diagnosis assay based on qPCR coupled to HRM that differentiates infections with diverse species of trypanosomatids and Leishmanias with zoonotic potential in peripheral blood samples from canines. The molecular marker selected was the Internal Transcribed Spacer (ITS1) present in the ribosomal RNA locus. This marker is highly conserved and present size variability among trypanosomes species. The results using as a template gDNA of different trypanosomatid species showed specific amplification with distinctive patterns in Melting Curves for T. evansi, T. cruzi, T. brucei, T. rangeli and different species of Leishmanias. This was confirmed in agarose gels, resulting in single or multiple bands with a size range from 250 to 480 bp. Its clinical validation was carried out on 14 peripheral blood samples from domestic canines from northeastern Argentina. The results showed positivity for infection with T. evansi in 36 % of the samples. Additionally, through this standardized technique, in one sample it could be detected infection with Leishmania infantum with low parasitemia, confirmed by sequencing and subsequent alignment of the ITS1 region with reference sequences. Therefore, molecular diagnosis of animal trypanosomiasis by HRM-qPCR represents a viable tool for wide-scale epidemiological studies, which may be used to report the true prevalence of the infection and allow implementation strategies to control these zoonotic diseases in Argentina, as well as the rest of South America, Africa, and Asia.
Fil: Muñoz Calderon, Arturo Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina
Fil: Lucero, Raul Horacio. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina
Fil: Brusés, Bettina Laura. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina
Fil: Formichelli, Laura Belén. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina
Fil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina
Reunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio
Mar del Plata
Argentina
Sociedad Argentina de Investigación Clínica
Asociación Argentina de Farmacología Experimenta
Sociedad Argentina de Biología
Sociedad Argentina de Protozoología
Asociación Argentina de Nanomedicinas
Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio
Materia: Trypanosoma evansi
Real Time PCR
Molecular diagnostic
High-resolution melting
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:11336/201596

Acceder

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network_name_str	CONICET Digital (CONICET)
spelling	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiarisMuñoz Calderon, Arturo AlejandroLucero, Raul HoracioBrusés, Bettina LauraFormichelli, Laura BelénSchijman, Alejandro GabrielTrypanosoma evansiReal Time PCRMolecular diagnosticHigh-resolution meltinghttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1The Trypanosomiasis caused by Trypanosoma evansi affects a wide diversity of mammals being zoonotic potential in man, with a case reported in 2005 in India. This haemoflagellate protozoan can parasitize most domestic mammals, being horses, dogs, and cattle the most affected species. Diagnostic tools for this parasitic infection are scarce, even though this trypanosomiasis can be very lethal if the animals are not treated. This work reports the development of a multi-diagnosis assay based on qPCR coupled to HRM that differentiates infections with diverse species of trypanosomatids and Leishmanias with zoonotic potential in peripheral blood samples from canines. The molecular marker selected was the Internal Transcribed Spacer (ITS1) present in the ribosomal RNA locus. This marker is highly conserved and present size variability among trypanosomes species. The results using as a template gDNA of different trypanosomatid species showed specific amplification with distinctive patterns in Melting Curves for T. evansi, T. cruzi, T. brucei, T. rangeli and different species of Leishmanias. This was confirmed in agarose gels, resulting in single or multiple bands with a size range from 250 to 480 bp. Its clinical validation was carried out on 14 peripheral blood samples from domestic canines from northeastern Argentina. The results showed positivity for infection with T. evansi in 36 % of the samples. Additionally, through this standardized technique, in one sample it could be detected infection with Leishmania infantum with low parasitemia, confirmed by sequencing and subsequent alignment of the ITS1 region with reference sequences. Therefore, molecular diagnosis of animal trypanosomiasis by HRM-qPCR represents a viable tool for wide-scale epidemiological studies, which may be used to report the true prevalence of the infection and allow implementation strategies to control these zoonotic diseases in Argentina, as well as the rest of South America, Africa, and Asia.Fil: Muñoz Calderon, Arturo Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Lucero, Raul Horacio. Universidad Nacional del Nordeste. Instituto de Medicina Regional; ArgentinaFil: Brusés, Bettina Laura. Universidad Nacional del Nordeste. Instituto de Medicina Regional; ArgentinaFil: Formichelli, Laura Belén. Universidad Nacional del Nordeste. Instituto de Medicina Regional; ArgentinaFil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaReunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de LaboratorioMar del PlataArgentinaSociedad Argentina de Investigación ClínicaAsociación Argentina de Farmacología ExperimentaSociedad Argentina de BiologíaSociedad Argentina de ProtozoologíaAsociación Argentina de NanomedicinasAsociación Argentina de Ciencia y Tecnología de Animales de LaboratorioMedicina (Buenos Aires)2019info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectReuniónJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/201596Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris; Reunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio; Mar del Plata; Argentina; 2019; 229-2290025-76801669-9106CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://medicinabuenosaires.com/revistas/vol79-19/s4/vol79_s4.pdfInternacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:45:15Zoai:ri.conicet.gov.ar:11336/201596instacron: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 09:45:15.735CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
title	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
spellingShingle	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris Muñoz Calderon, Arturo Alejandro Trypanosoma evansi Real Time PCR Molecular diagnostic High-resolution melting
title_short	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
title_full	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
title_fullStr	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
title_full_unstemmed	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
title_sort	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris
dc.creator.none.fl_str_mv	Muñoz Calderon, Arturo Alejandro Lucero, Raul Horacio Brusés, Bettina Laura Formichelli, Laura Belén Schijman, Alejandro Gabriel
author	Muñoz Calderon, Arturo Alejandro
author_facet	Muñoz Calderon, Arturo Alejandro Lucero, Raul Horacio Brusés, Bettina Laura Formichelli, Laura Belén Schijman, Alejandro Gabriel
author_role	author
author2	Lucero, Raul Horacio Brusés, Bettina Laura Formichelli, Laura Belén Schijman, Alejandro Gabriel
author2_role	author author author author
dc.subject.none.fl_str_mv	Trypanosoma evansi Real Time PCR Molecular diagnostic High-resolution melting
topic	Trypanosoma evansi Real Time PCR Molecular diagnostic High-resolution melting
purl_subject.fl_str_mv	https://purl.org/becyt/ford/1.6 https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv	The Trypanosomiasis caused by Trypanosoma evansi affects a wide diversity of mammals being zoonotic potential in man, with a case reported in 2005 in India. This haemoflagellate protozoan can parasitize most domestic mammals, being horses, dogs, and cattle the most affected species. Diagnostic tools for this parasitic infection are scarce, even though this trypanosomiasis can be very lethal if the animals are not treated. This work reports the development of a multi-diagnosis assay based on qPCR coupled to HRM that differentiates infections with diverse species of trypanosomatids and Leishmanias with zoonotic potential in peripheral blood samples from canines. The molecular marker selected was the Internal Transcribed Spacer (ITS1) present in the ribosomal RNA locus. This marker is highly conserved and present size variability among trypanosomes species. The results using as a template gDNA of different trypanosomatid species showed specific amplification with distinctive patterns in Melting Curves for T. evansi, T. cruzi, T. brucei, T. rangeli and different species of Leishmanias. This was confirmed in agarose gels, resulting in single or multiple bands with a size range from 250 to 480 bp. Its clinical validation was carried out on 14 peripheral blood samples from domestic canines from northeastern Argentina. The results showed positivity for infection with T. evansi in 36 % of the samples. Additionally, through this standardized technique, in one sample it could be detected infection with Leishmania infantum with low parasitemia, confirmed by sequencing and subsequent alignment of the ITS1 region with reference sequences. Therefore, molecular diagnosis of animal trypanosomiasis by HRM-qPCR represents a viable tool for wide-scale epidemiological studies, which may be used to report the true prevalence of the infection and allow implementation strategies to control these zoonotic diseases in Argentina, as well as the rest of South America, Africa, and Asia. Fil: Muñoz Calderon, Arturo Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina Fil: Lucero, Raul Horacio. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina Fil: Brusés, Bettina Laura. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina Fil: Formichelli, Laura Belén. Universidad Nacional del Nordeste. Instituto de Medicina Regional; Argentina Fil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina Reunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio Mar del Plata Argentina Sociedad Argentina de Investigación Clínica Asociación Argentina de Farmacología Experimenta Sociedad Argentina de Biología Sociedad Argentina de Protozoología Asociación Argentina de Nanomedicinas Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio
description	The Trypanosomiasis caused by Trypanosoma evansi affects a wide diversity of mammals being zoonotic potential in man, with a case reported in 2005 in India. This haemoflagellate protozoan can parasitize most domestic mammals, being horses, dogs, and cattle the most affected species. Diagnostic tools for this parasitic infection are scarce, even though this trypanosomiasis can be very lethal if the animals are not treated. This work reports the development of a multi-diagnosis assay based on qPCR coupled to HRM that differentiates infections with diverse species of trypanosomatids and Leishmanias with zoonotic potential in peripheral blood samples from canines. The molecular marker selected was the Internal Transcribed Spacer (ITS1) present in the ribosomal RNA locus. This marker is highly conserved and present size variability among trypanosomes species. The results using as a template gDNA of different trypanosomatid species showed specific amplification with distinctive patterns in Melting Curves for T. evansi, T. cruzi, T. brucei, T. rangeli and different species of Leishmanias. This was confirmed in agarose gels, resulting in single or multiple bands with a size range from 250 to 480 bp. Its clinical validation was carried out on 14 peripheral blood samples from domestic canines from northeastern Argentina. The results showed positivity for infection with T. evansi in 36 % of the samples. Additionally, through this standardized technique, in one sample it could be detected infection with Leishmania infantum with low parasitemia, confirmed by sequencing and subsequent alignment of the ITS1 region with reference sequences. Therefore, molecular diagnosis of animal trypanosomiasis by HRM-qPCR represents a viable tool for wide-scale epidemiological studies, which may be used to report the true prevalence of the infection and allow implementation strategies to control these zoonotic diseases in Argentina, as well as the rest of South America, Africa, and Asia.
publishDate	2019
dc.date.none.fl_str_mv	2019
dc.type.none.fl_str_mv	info:eu-repo/semantics/publishedVersion info:eu-repo/semantics/conferenceObject Reunión Journal http://purl.org/coar/resource_type/c_5794 info:ar-repo/semantics/documentoDeConferencia
status_str	publishedVersion
format	conferenceObject
dc.identifier.none.fl_str_mv	http://hdl.handle.net/11336/201596 Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris; Reunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio; Mar del Plata; Argentina; 2019; 229-229 0025-7680 1669-9106 CONICET Digital CONICET
url	http://hdl.handle.net/11336/201596
identifier_str_mv	Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus familiaris; Reunión Anual de Sociedades de Biociencias 2019; LXIV Reunión Anual de la Sociedad Argentina de Investigación Clínica; LI Reunión Anual de la Asociación Argentina de Farmacología Experimenta; XXI Reunión Anual de la Sociedad Argentina de Biología; XXXI Reunión Anual de la Sociedad Argentina de Protozoología; IX Reunión Anual de la Asociación Argentina de Nanomedicinas; VI Reunión Científica Regional de la Asociación Argentina de Ciencia y Tecnología de Animales de Laboratorio; Mar del Plata; Argentina; 2019; 229-229 0025-7680 1669-9106 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://medicinabuenosaires.com/revistas/vol79-19/s4/vol79_s4.pdf
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dc.coverage.none.fl_str_mv	Internacional
dc.publisher.none.fl_str_mv	Medicina (Buenos Aires)
publisher.none.fl_str_mv	Medicina (Buenos Aires)
dc.source.none.fl_str_mv	reponame:CONICET Digital (CONICET) instname:Consejo Nacional de Investigaciones Científicas y Técnicas
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repository.mail.fl_str_mv	dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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Development of quantitative real-time polymerase chain reaction coupled with high-resolution melting (HRM- qPCR) analysis for the diagnosis of Trypanosoma evansi in Canis lupus fam...

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