Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species
Autor(a) principal: | |
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Data de Publicação: | 2012 |
Outros Autores: | , , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UnB |
Texto Completo: | http://repositorio.unb.br/handle/10482/28305 https://dx.doi.org/10.1590/S0074-02762012000500014 |
Resumo: | In this study, PCR assays targeting different Leishmania heat-shock protein 70 gene (hsp70) regions, producing fragments ranging in size from 230-390 bp were developed and evaluated to determine their potential as a tool for the specific molecular diagnosis of cutaneous leishmaniasis (CL). A total of 70 Leishmania strains were analysed, including seven reference strains (RS) and 63 previously typed strains. Analysis of the RS indicated a specific region of 234 bp in the hsp70 gene as a valid target that was highly sensitive for detection of Leishmania species DNA with capacity of distinguishing all analyzed species, after polymerase chain reaction-restriction fragment length polymorfism (PCR-RFLP). This PCR assay was compared with other PCR targets used for the molecular diagnosis of leishmaniasis: hsp70 (1400-bp region), internal transcribed spacer (ITS)1 and glucose-6-phosphate dehydrogenase (G6pd). A good agreement among the methods was observed concerning the Leishmania species identification. Moreover, to evaluate the potential for molecular diagnosis, we compared the PCR targets hsp70-234 bp, ITS1, G6pd and mkDNA using a panel of 99 DNA samples from tissue fragments collected from patients with confirmed CL. Both PCR-hsp70-234 bp and PCR-ITS1 detected Leishmania DNA in more than 70% of the samples. However, using hsp70-234 bp PCR-RFLP, identification of all of the Leishmania species associated with CL in Brazil can be achieved employing a simpler and cheaper electrophoresis protocol. |
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Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite speciesLeishmanioseDiagnóstico molecularIdentificação de espéciesReação em cadeia de polimeraseIn this study, PCR assays targeting different Leishmania heat-shock protein 70 gene (hsp70) regions, producing fragments ranging in size from 230-390 bp were developed and evaluated to determine their potential as a tool for the specific molecular diagnosis of cutaneous leishmaniasis (CL). A total of 70 Leishmania strains were analysed, including seven reference strains (RS) and 63 previously typed strains. Analysis of the RS indicated a specific region of 234 bp in the hsp70 gene as a valid target that was highly sensitive for detection of Leishmania species DNA with capacity of distinguishing all analyzed species, after polymerase chain reaction-restriction fragment length polymorfism (PCR-RFLP). This PCR assay was compared with other PCR targets used for the molecular diagnosis of leishmaniasis: hsp70 (1400-bp region), internal transcribed spacer (ITS)1 and glucose-6-phosphate dehydrogenase (G6pd). A good agreement among the methods was observed concerning the Leishmania species identification. Moreover, to evaluate the potential for molecular diagnosis, we compared the PCR targets hsp70-234 bp, ITS1, G6pd and mkDNA using a panel of 99 DNA samples from tissue fragments collected from patients with confirmed CL. Both PCR-hsp70-234 bp and PCR-ITS1 detected Leishmania DNA in more than 70% of the samples. However, using hsp70-234 bp PCR-RFLP, identification of all of the Leishmania species associated with CL in Brazil can be achieved employing a simpler and cheaper electrophoresis protocol.Faculdade de Medicina (FMD)Instituto Oswaldo Cruz, Ministério da Saúde2017-12-07T04:58:14Z2017-12-07T04:58:14Z2012-08info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfGRACA, Grazielle Cardoso da et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species. Memórias do Instituto Oswaldo Cruz, Rio de Janeiro, v. 107, n. 5, p. 664-674, ago. 2012. DOI: https://doi.org/10.1590/S0074-02762012000500014. Disponível em: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000500014&lng=en&nrm=iso. Acesso em: 04 dez. 2020.http://repositorio.unb.br/handle/10482/28305https://dx.doi.org/10.1590/S0074-02762012000500014Memórias do Instituto Oswaldo Cruz - All the contents of this journal, except where otherwise noted, is licensed under a Creative Commons Attribution License (CC BY NC 4.0). Fonte: https://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000500014&lng=en&tlng=en. Acesso em: 04 dez. 2020.info:eu-repo/semantics/openAccessGraça, Grazielle Cardoso daVolpini, Angela CristinaRomero, Gustavo Adolfo SierraOliveira Neto, Manoel Paes deHueb, MarciaPorrozzi, RenatoBoité, Mariana CôrtesCupolillo, Elisaengreponame:Repositório Institucional da UnBinstname:Universidade de Brasília (UnB)instacron:UNB2024-05-24T10:39:53Zoai:repositorio.unb.br:10482/28305Repositório InstitucionalPUBhttps://repositorio.unb.br/oai/requestrepositorio@unb.bropendoar:2024-05-24T10:39:53Repositório Institucional da UnB - Universidade de Brasília (UnB)false |
dc.title.none.fl_str_mv |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
title |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
spellingShingle |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species Graça, Grazielle Cardoso da Leishmaniose Diagnóstico molecular Identificação de espécies Reação em cadeia de polimerase |
title_short |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
title_full |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
title_fullStr |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
title_full_unstemmed |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
title_sort |
Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species |
author |
Graça, Grazielle Cardoso da |
author_facet |
Graça, Grazielle Cardoso da Volpini, Angela Cristina Romero, Gustavo Adolfo Sierra Oliveira Neto, Manoel Paes de Hueb, Marcia Porrozzi, Renato Boité, Mariana Côrtes Cupolillo, Elisa |
author_role |
author |
author2 |
Volpini, Angela Cristina Romero, Gustavo Adolfo Sierra Oliveira Neto, Manoel Paes de Hueb, Marcia Porrozzi, Renato Boité, Mariana Côrtes Cupolillo, Elisa |
author2_role |
author author author author author author author |
dc.contributor.author.fl_str_mv |
Graça, Grazielle Cardoso da Volpini, Angela Cristina Romero, Gustavo Adolfo Sierra Oliveira Neto, Manoel Paes de Hueb, Marcia Porrozzi, Renato Boité, Mariana Côrtes Cupolillo, Elisa |
dc.subject.por.fl_str_mv |
Leishmaniose Diagnóstico molecular Identificação de espécies Reação em cadeia de polimerase |
topic |
Leishmaniose Diagnóstico molecular Identificação de espécies Reação em cadeia de polimerase |
description |
In this study, PCR assays targeting different Leishmania heat-shock protein 70 gene (hsp70) regions, producing fragments ranging in size from 230-390 bp were developed and evaluated to determine their potential as a tool for the specific molecular diagnosis of cutaneous leishmaniasis (CL). A total of 70 Leishmania strains were analysed, including seven reference strains (RS) and 63 previously typed strains. Analysis of the RS indicated a specific region of 234 bp in the hsp70 gene as a valid target that was highly sensitive for detection of Leishmania species DNA with capacity of distinguishing all analyzed species, after polymerase chain reaction-restriction fragment length polymorfism (PCR-RFLP). This PCR assay was compared with other PCR targets used for the molecular diagnosis of leishmaniasis: hsp70 (1400-bp region), internal transcribed spacer (ITS)1 and glucose-6-phosphate dehydrogenase (G6pd). A good agreement among the methods was observed concerning the Leishmania species identification. Moreover, to evaluate the potential for molecular diagnosis, we compared the PCR targets hsp70-234 bp, ITS1, G6pd and mkDNA using a panel of 99 DNA samples from tissue fragments collected from patients with confirmed CL. Both PCR-hsp70-234 bp and PCR-ITS1 detected Leishmania DNA in more than 70% of the samples. However, using hsp70-234 bp PCR-RFLP, identification of all of the Leishmania species associated with CL in Brazil can be achieved employing a simpler and cheaper electrophoresis protocol. |
publishDate |
2012 |
dc.date.none.fl_str_mv |
2012-08 2017-12-07T04:58:14Z 2017-12-07T04:58:14Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
GRACA, Grazielle Cardoso da et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species. Memórias do Instituto Oswaldo Cruz, Rio de Janeiro, v. 107, n. 5, p. 664-674, ago. 2012. DOI: https://doi.org/10.1590/S0074-02762012000500014. Disponível em: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000500014&lng=en&nrm=iso. Acesso em: 04 dez. 2020. http://repositorio.unb.br/handle/10482/28305 https://dx.doi.org/10.1590/S0074-02762012000500014 |
identifier_str_mv |
GRACA, Grazielle Cardoso da et al. Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species. Memórias do Instituto Oswaldo Cruz, Rio de Janeiro, v. 107, n. 5, p. 664-674, ago. 2012. DOI: https://doi.org/10.1590/S0074-02762012000500014. Disponível em: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0074-02762012000500014&lng=en&nrm=iso. Acesso em: 04 dez. 2020. |
url |
http://repositorio.unb.br/handle/10482/28305 https://dx.doi.org/10.1590/S0074-02762012000500014 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Instituto Oswaldo Cruz, Ministério da Saúde |
publisher.none.fl_str_mv |
Instituto Oswaldo Cruz, Ministério da Saúde |
dc.source.none.fl_str_mv |
reponame:Repositório Institucional da UnB instname:Universidade de Brasília (UnB) instacron:UNB |
instname_str |
Universidade de Brasília (UnB) |
instacron_str |
UNB |
institution |
UNB |
reponame_str |
Repositório Institucional da UnB |
collection |
Repositório Institucional da UnB |
repository.name.fl_str_mv |
Repositório Institucional da UnB - Universidade de Brasília (UnB) |
repository.mail.fl_str_mv |
repositorio@unb.br |
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1810580855043653632 |