Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays
| Autor(a) principal: | |
|---|---|
| Data de Publicação: | 2020 |
| Outros Autores: | , , , , , , , , , |
| Tipo de documento: | Artigo |
| Idioma: | por |
| Título da fonte: | Repositório Institucional da FIOCRUZ (ARCA) |
| Texto Completo: | https://www.arca.fiocruz.br/handle/icict/42019 |
Resumo: | Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil. |
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Manta, Fernanda Saloum de NevesLeal-Calvo, ThyagoMoreira, Suelen Justo M.Marques, Brunna L. C.Ribeiro-Alves, MarceloRosa, Patrícia S.Nery, José Augusto C.Rampazzo, Rita de Cássia PontelloCosta, Alexandre Dias TavaresKrieger, Marco AurelioMoraes, Milton Ozório2020-06-30T20:54:22Z2020-06-30T20:54:22Z2020MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020.1935-2735https://www.arca.fiocruz.br/handle/icict/4201910.1371/journal.pntd.00083251935-2735porPublic Library of SciencePCRLeprosyPolymerase Chain ReactionReagent Kits, DiagnosticLepraReacción en Cadena de la PolimerasaJuego de Reactivos para DiagnósticoHanseníaseReação em Cadeia da PolimeraseKit de Reagentes para DiagnósticoUltra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assaysinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Nacional de Infectologia Evandro Chagas. Laboratório de Pesquisa Clínica em DST-AIDS. Rio de Janeiro, RJ, Brasil.Instituto Lauro de Souza Lima. Bauru, SP, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil. / Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto de Biologia Molecular do Paraná. Curitiba, PR, Brasil. / Fundação Oswaldo Cruz. Instituto Carlos Chagas. Curitiba, PR, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil.Leprosy urgently needs a precise and early diagnostic tool. The sensitivity of the direct (bacilli staining, Mycobacterium leprae DNA) and indirect (antibody levels, T cell assays) diagnostics methods vary based on the clinical form. Recently, PCR-based M. leprae DNA detection has been shown to differentially diagnose leprosy from other dermatological conditions. However, accuracy can still be improved, especially for use with less invasive clinical samples. We tested different commercial DNA extraction kits: DNeasy Blood & Tissue, QIAamp DNA Microbiome, Maxwell 16 DNA Purification, PowerSoil DNA Isolation; as well as in-house phenol-chloroform and Trizol/FastPrep methods. Extraction was performed on M. leprae-infected mouse footpads and different clinical samples of leprosy patients (skin biopsies and scrapings, lesion, oral and nasal swabs, body hair, blood on FTA cards, peripheral whole blood). We observed that the Microbiome kit was able to enrich for mycobacterial DNA, most likely due the enzymatic digestion cocktail along with mechanical disruption involved in this method. Consequently, we had a significant increase in sensitivity in skin biopsies from paucibacillary leprosy patients using a duplex qPCR targeting 16S rRNA (M. leprae) and 18S rRNA (mammal) in the StepOnePlus system. Our data showed that the presence of M. leprae DNA was best detected in skin biopsies and skin scrapings, independent of the extraction method or the clinical form. For multibacillary patients, detection of M. leprae DNA in nasal swabs indicates the possibility of having a much less invasive sample that can be used for the purposes of DNA sequencing for relapse analysis and drug resistance monitoring. Overall, DNA extracted with the Microbiome kit presented the best bacilli detection rate for paucibacillary cases, indicating that investments in extraction methods with mechanical and DNA digestion should be made.info:eu-repo/semantics/openAccessreponame:Repositório Institucional da FIOCRUZ (ARCA)instname:Fundação Oswaldo Cruz (FIOCRUZ)instacron:FIOCRUZLICENSElicense.txtlicense.txttext/plain; charset=utf-83084https://www.arca.fiocruz.br/bitstream/icict/42019/1/license.txt783568c2893d2e25a99990b126be1772MD51ORIGINALjournal.pntd.000832.pdfjournal.pntd.000832.pdfapplication/pdf1434923https://www.arca.fiocruz.br/bitstream/icict/42019/2/journal.pntd.000832.pdf8cf1cbd5c7b0c3c228cb3c2ed1cb4611MD52TEXTjournal.pntd.000832.pdf.txtjournal.pntd.000832.pdf.txtExtracted texttext/plain49711https://www.arca.fiocruz.br/bitstream/icict/42019/3/journal.pntd.000832.pdf.txt790c185fc532fb6c98dcf0ec020e1680MD53icict/420192023-08-02 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| dc.title.pt_BR.fl_str_mv |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| title |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| spellingShingle |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays Manta, Fernanda Saloum de Neves PCR Leprosy Polymerase Chain Reaction Reagent Kits, Diagnostic Lepra Reacción en Cadena de la Polimerasa Juego de Reactivos para Diagnóstico Hanseníase Reação em Cadeia da Polimerase Kit de Reagentes para Diagnóstico |
| title_short |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| title_full |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| title_fullStr |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| title_full_unstemmed |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| title_sort |
Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays |
| author |
Manta, Fernanda Saloum de Neves |
| author_facet |
Manta, Fernanda Saloum de Neves Leal-Calvo, Thyago Moreira, Suelen Justo M. Marques, Brunna L. C. Ribeiro-Alves, Marcelo Rosa, Patrícia S. Nery, José Augusto C. Rampazzo, Rita de Cássia Pontello Costa, Alexandre Dias Tavares Krieger, Marco Aurelio Moraes, Milton Ozório |
| author_role |
author |
| author2 |
Leal-Calvo, Thyago Moreira, Suelen Justo M. Marques, Brunna L. C. Ribeiro-Alves, Marcelo Rosa, Patrícia S. Nery, José Augusto C. Rampazzo, Rita de Cássia Pontello Costa, Alexandre Dias Tavares Krieger, Marco Aurelio Moraes, Milton Ozório |
| author2_role |
author author author author author author author author author author |
| dc.contributor.author.fl_str_mv |
Manta, Fernanda Saloum de Neves Leal-Calvo, Thyago Moreira, Suelen Justo M. Marques, Brunna L. C. Ribeiro-Alves, Marcelo Rosa, Patrícia S. Nery, José Augusto C. Rampazzo, Rita de Cássia Pontello Costa, Alexandre Dias Tavares Krieger, Marco Aurelio Moraes, Milton Ozório |
| dc.subject.other.pt_BR.fl_str_mv |
PCR |
| topic |
PCR Leprosy Polymerase Chain Reaction Reagent Kits, Diagnostic Lepra Reacción en Cadena de la Polimerasa Juego de Reactivos para Diagnóstico Hanseníase Reação em Cadeia da Polimerase Kit de Reagentes para Diagnóstico |
| dc.subject.en.pt_BR.fl_str_mv |
Leprosy Polymerase Chain Reaction Reagent Kits, Diagnostic |
| dc.subject.es.pt_BR.fl_str_mv |
Lepra Reacción en Cadena de la Polimerasa Juego de Reactivos para Diagnóstico |
| dc.subject.decs.pt_BR.fl_str_mv |
Hanseníase Reação em Cadeia da Polimerase Kit de Reagentes para Diagnóstico |
| description |
Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Hanseníase. Rio de Janeiro, RJ, Brasil. |
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2020 |
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2020-06-30T20:54:22Z |
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2020-06-30T20:54:22Z |
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2020 |
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info:eu-repo/semantics/publishedVersion |
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info:eu-repo/semantics/article |
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article |
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publishedVersion |
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MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020. |
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https://www.arca.fiocruz.br/handle/icict/42019 |
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1935-2735 |
| dc.identifier.doi.none.fl_str_mv |
10.1371/journal.pntd.0008325 |
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1935-2735 |
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MANTA, Fernanda Saloum de Neves et al. Ultra-sensitive detection of Mycobacterium leprae: DNA extraction and PCR assays. PLoS Neglected Tropical Diseases, v. 14, n. 5, p. 1-15, 2020. 1935-2735 10.1371/journal.pntd.0008325 |
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Public Library of Science |
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