Rapid detection of carbapenemase genes by multiplex real-time PCR
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 UNIFESP |
Texto Completo: | http://repositorio.unifesp.br/handle/11600/34725 http://dx.doi.org/10.1093/jac/dkr563 |
Resumo: | To develop a single multiplex real-time PCR assay to detect six different genetic types of carbapenemases already identified in Enterobacteriaceae (KPC, GES, NDM, IMP, VIM and OXA-48).A total of 58 bacterial isolates were tested. Thirty were previously characterized as resistant to carbapenems and documented by PCR and sequencing analysis to carry the following genes: bla(KPC) type, bla(GES) type, bla(IMP) type, bla(VIM) type, bla(OXA-48) and bla(NDM-1). These positive strains included 21 Enterobacteriaceae, 1 Acinetobacter baumannii and 8 Pseudomonas aeruginosa isolates. the remaining 28 isolates previously tested susceptible to carbapenems and were negative for these genes. Bacterial DNA was extracted using the easyMag extractor (bioMrieux, France). the real-time PCR was performed using the Rotor-Gene 6000 instrument (Corbett Life Science, Australia) and specific primers for each carbapenemase target were designed using the DNAStar software (Madison, WI, USA).Each one of the six carbapenemase genes tested presented a different melting curve after PCR amplification. the melting temperature (T-m) analysis of the amplicons identified was as follows: bla(IMP) type (T-m 80.1C), bla(OXA-48) (T-m 81.6C), bla(NDM-1) (T-m 84C), bla(GES) type (T-m 88.6C), bla(VIM) type (T-m 90.3C) and bla(KPC) type (T-m 91.6C). No amplification was detected among the negative samples. the results showed 100 concordance with the genotypes previously identified.The new assay was able to detect the presence of six different carbapenemase gene types in a single 3 h PCR. |
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Monteiro, Jussimara [UNIFESP]Widen, Raymond H.Pignatari, Antonio C. C. [UNIFESP]Kubasek, CarlySilbert, SuzaneTampa Gen HospUniversidade Federal de São Paulo (UNIFESP)2016-01-24T14:27:00Z2016-01-24T14:27:00Z2012-04-01Journal of Antimicrobial Chemotherapy. Oxford: Oxford Univ Press, v. 67, n. 4, p. 906-909, 2012.0305-7453http://repositorio.unifesp.br/handle/11600/34725http://dx.doi.org/10.1093/jac/dkr56310.1093/jac/dkr563WOS:000301684900017To develop a single multiplex real-time PCR assay to detect six different genetic types of carbapenemases already identified in Enterobacteriaceae (KPC, GES, NDM, IMP, VIM and OXA-48).A total of 58 bacterial isolates were tested. Thirty were previously characterized as resistant to carbapenems and documented by PCR and sequencing analysis to carry the following genes: bla(KPC) type, bla(GES) type, bla(IMP) type, bla(VIM) type, bla(OXA-48) and bla(NDM-1). These positive strains included 21 Enterobacteriaceae, 1 Acinetobacter baumannii and 8 Pseudomonas aeruginosa isolates. the remaining 28 isolates previously tested susceptible to carbapenems and were negative for these genes. Bacterial DNA was extracted using the easyMag extractor (bioMrieux, France). the real-time PCR was performed using the Rotor-Gene 6000 instrument (Corbett Life Science, Australia) and specific primers for each carbapenemase target were designed using the DNAStar software (Madison, WI, USA).Each one of the six carbapenemase genes tested presented a different melting curve after PCR amplification. the melting temperature (T-m) analysis of the amplicons identified was as follows: bla(IMP) type (T-m 80.1C), bla(OXA-48) (T-m 81.6C), bla(NDM-1) (T-m 84C), bla(GES) type (T-m 88.6C), bla(VIM) type (T-m 90.3C) and bla(KPC) type (T-m 91.6C). No amplification was detected among the negative samples. the results showed 100 concordance with the genotypes previously identified.The new assay was able to detect the presence of six different carbapenemase gene types in a single 3 h PCR.Tampa General Hospital-Esoteric Laboratory TestingCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Tampa Gen Hosp, Esoter Testing Lab, Dept Pathol, Tampa, FL 33606 USAUniversidade Federal de São Paulo, LEMC, São Paulo, BrazilUniversidade Federal de São Paulo, LEMC, São Paulo, BrazilCAPES: 1152/10-6Web of Science906-909engOxford Univ PressJournal of Antimicrobial Chemotherapyhttp://www.oxfordjournals.org/access_purchase/self-archiving_policyb.htmlinfo:eu-repo/semantics/openAccessresistance-lactamasesEnterobacteriaceaeRapid detection of carbapenemase genes by multiplex real-time PCRinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlereponame:Repositório Institucional da UNIFESPinstname:Universidade Federal de São Paulo (UNIFESP)instacron:UNIFESP11600/347252022-11-04 15:20:28.12metadata only accessoai:repositorio.unifesp.br:11600/34725Repositório InstitucionalPUBhttp://www.repositorio.unifesp.br/oai/requestopendoar:34652022-11-04T18:20:28Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP)false |
dc.title.en.fl_str_mv |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
title |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
spellingShingle |
Rapid detection of carbapenemase genes by multiplex real-time PCR Monteiro, Jussimara [UNIFESP] resistance -lactamases Enterobacteriaceae |
title_short |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
title_full |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
title_fullStr |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
title_full_unstemmed |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
title_sort |
Rapid detection of carbapenemase genes by multiplex real-time PCR |
author |
Monteiro, Jussimara [UNIFESP] |
author_facet |
Monteiro, Jussimara [UNIFESP] Widen, Raymond H. Pignatari, Antonio C. C. [UNIFESP] Kubasek, Carly Silbert, Suzane |
author_role |
author |
author2 |
Widen, Raymond H. Pignatari, Antonio C. C. [UNIFESP] Kubasek, Carly Silbert, Suzane |
author2_role |
author author author author |
dc.contributor.institution.none.fl_str_mv |
Tampa Gen Hosp Universidade Federal de São Paulo (UNIFESP) |
dc.contributor.author.fl_str_mv |
Monteiro, Jussimara [UNIFESP] Widen, Raymond H. Pignatari, Antonio C. C. [UNIFESP] Kubasek, Carly Silbert, Suzane |
dc.subject.eng.fl_str_mv |
resistance -lactamases Enterobacteriaceae |
topic |
resistance -lactamases Enterobacteriaceae |
description |
To develop a single multiplex real-time PCR assay to detect six different genetic types of carbapenemases already identified in Enterobacteriaceae (KPC, GES, NDM, IMP, VIM and OXA-48).A total of 58 bacterial isolates were tested. Thirty were previously characterized as resistant to carbapenems and documented by PCR and sequencing analysis to carry the following genes: bla(KPC) type, bla(GES) type, bla(IMP) type, bla(VIM) type, bla(OXA-48) and bla(NDM-1). These positive strains included 21 Enterobacteriaceae, 1 Acinetobacter baumannii and 8 Pseudomonas aeruginosa isolates. the remaining 28 isolates previously tested susceptible to carbapenems and were negative for these genes. Bacterial DNA was extracted using the easyMag extractor (bioMrieux, France). the real-time PCR was performed using the Rotor-Gene 6000 instrument (Corbett Life Science, Australia) and specific primers for each carbapenemase target were designed using the DNAStar software (Madison, WI, USA).Each one of the six carbapenemase genes tested presented a different melting curve after PCR amplification. the melting temperature (T-m) analysis of the amplicons identified was as follows: bla(IMP) type (T-m 80.1C), bla(OXA-48) (T-m 81.6C), bla(NDM-1) (T-m 84C), bla(GES) type (T-m 88.6C), bla(VIM) type (T-m 90.3C) and bla(KPC) type (T-m 91.6C). No amplification was detected among the negative samples. the results showed 100 concordance with the genotypes previously identified.The new assay was able to detect the presence of six different carbapenemase gene types in a single 3 h PCR. |
publishDate |
2012 |
dc.date.issued.fl_str_mv |
2012-04-01 |
dc.date.accessioned.fl_str_mv |
2016-01-24T14:27:00Z |
dc.date.available.fl_str_mv |
2016-01-24T14:27:00Z |
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.citation.fl_str_mv |
Journal of Antimicrobial Chemotherapy. Oxford: Oxford Univ Press, v. 67, n. 4, p. 906-909, 2012. |
dc.identifier.uri.fl_str_mv |
http://repositorio.unifesp.br/handle/11600/34725 http://dx.doi.org/10.1093/jac/dkr563 |
dc.identifier.issn.none.fl_str_mv |
0305-7453 |
dc.identifier.doi.none.fl_str_mv |
10.1093/jac/dkr563 |
dc.identifier.wos.none.fl_str_mv |
WOS:000301684900017 |
identifier_str_mv |
Journal of Antimicrobial Chemotherapy. Oxford: Oxford Univ Press, v. 67, n. 4, p. 906-909, 2012. 0305-7453 10.1093/jac/dkr563 WOS:000301684900017 |
url |
http://repositorio.unifesp.br/handle/11600/34725 http://dx.doi.org/10.1093/jac/dkr563 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.ispartof.none.fl_str_mv |
Journal of Antimicrobial Chemotherapy |
dc.rights.driver.fl_str_mv |
http://www.oxfordjournals.org/access_purchase/self-archiving_policyb.html info:eu-repo/semantics/openAccess |
rights_invalid_str_mv |
http://www.oxfordjournals.org/access_purchase/self-archiving_policyb.html |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
906-909 |
dc.publisher.none.fl_str_mv |
Oxford Univ Press |
publisher.none.fl_str_mv |
Oxford Univ Press |
dc.source.none.fl_str_mv |
reponame:Repositório Institucional da UNIFESP instname:Universidade Federal de São Paulo (UNIFESP) instacron:UNIFESP |
instname_str |
Universidade Federal de São Paulo (UNIFESP) |
instacron_str |
UNIFESP |
institution |
UNIFESP |
reponame_str |
Repositório Institucional da UNIFESP |
collection |
Repositório Institucional da UNIFESP |
repository.name.fl_str_mv |
Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP) |
repository.mail.fl_str_mv |
|
_version_ |
1802764184688001024 |