308 479

Cited 48 times in

Rapid detection of fluoroquinolone-resistant and heteroresistant Mycobacterium tuberculosis by use of sloppy molecular beacons and dual melting-temperature codes in a real-time PCR assay

DC Field Value Language
dc.contributor.author조상래-
dc.contributor.author이은계-
dc.date.accessioned2014-12-20T17:09:47Z-
dc.date.available2014-12-20T17:09:47Z-
dc.date.issued2011-
dc.identifier.issn0095-1137-
dc.identifier.urihttps://ir.ymlib.yonsei.ac.kr/handle/22282913/94076-
dc.description.abstractFluoroquinolones (FQ) are important second-line drugs to treat tuberculosis; however, FQ resistance is an emerging problem. Resistance has been mainly attributed to mutations in a 21-bp region of the Mycobacterium tuberculosis gyrA gene, often called the quinolone resistance-determining region (QRDR). We have developed a simple, rapid, and specific assay to detect FQ resistance-determining QRDR mutations. The assay amplifies the M. tuberculosis gyrA QRDR in an asymmetrical PCR followed by probing with two sloppy molecular beacons (SMBs) spanning the entire QRDR. Mutations are detected by melting temperature (T(m)) shifts that occur when the SMBs bind to mismatched sequences. By testing DNA targets corresponding to all known QRDR mutations, we found that one or both of the SMBs produced a T(m) shift of at least 3.6°C for each mutation, making mutation detection very robust. The assay was also able to identify mixtures of wild-type and mutant DNA, with QRDR mutants identified in samples containing as little as 5 to 10% mutant DNA. The assay was blindly validated for its ability to identify the QRDR mutations on DNA extracted from clinical M. tuberculosis strains. Fifty QRDR wild-type samples, 34 QRDR mutant samples, and 8 heteroresistant samples containing mixtures of wild-type and mutant DNA were analyzed. The results showed 100% concordance to conventional DNA sequencing, including a complete identification of all of the mixtures. This SMB T(m) shift assay will be a valuable molecular tool to rapidly detect FQ resistance and to detect the emergence of FQ heteroresistance in clinical samples from tuberculosis patients.-
dc.description.statementOfResponsibilityopen-
dc.format.extent932~940-
dc.relation.isPartOfJOURNAL OF CLINICAL MICROBIOLOGY-
dc.rightsCC BY-NC-ND 2.0 KR-
dc.rights.urihttps://creativecommons.org/licenses/by-nc-nd/2.0/kr/-
dc.subject.MESHAnti-Bacterial Agents/pharmacology*-
dc.subject.MESHBacteriological Techniques/methods*-
dc.subject.MESHDNA Gyrase/genetics-
dc.subject.MESHDNA Primers/genetics-
dc.subject.MESHDNA, Bacterial/chemistry-
dc.subject.MESHDNA, Bacterial/genetics-
dc.subject.MESHFluoroquinolones/pharmacology*-
dc.subject.MESHHumans-
dc.subject.MESHMycobacterium tuberculosis/drug effects*-
dc.subject.MESHMycobacterium tuberculosis/isolation & purification-
dc.subject.MESHOligonucleotide Probes/genetics-
dc.subject.MESHPolymerase Chain Reaction/methods*-
dc.subject.MESHTransition Temperature-
dc.subject.MESHTuberculosis, Multidrug-Resistant/diagnosis*-
dc.subject.MESHTuberculosis, Multidrug-Resistant/microbiology*-
dc.titleRapid detection of fluoroquinolone-resistant and heteroresistant Mycobacterium tuberculosis by use of sloppy molecular beacons and dual melting-temperature codes in a real-time PCR assay-
dc.typeArticle-
dc.contributor.collegeCollege of Medicine (의과대학)-
dc.contributor.departmentDept. of Microbiology (미생물학)-
dc.contributor.googleauthorSoumitesh Chakravorty-
dc.contributor.googleauthorBola Aladegbami-
dc.contributor.googleauthorKimberley Thoms-
dc.contributor.googleauthorJong Seok Lee-
dc.contributor.googleauthorEun Gae Lee-
dc.contributor.googleauthorVignesh Rajan-
dc.contributor.googleauthorEun-Jin Cho-
dc.contributor.googleauthorHyunchul Kim-
dc.contributor.googleauthorHyunkyung Kwak-
dc.contributor.googleauthorNatalia Kurepina-
dc.contributor.googleauthorSang-Nae Cho-
dc.contributor.googleauthorBarry Kreiswirth-
dc.contributor.googleauthorLaura E. Via-
dc.contributor.googleauthorClifton E. Barry-
dc.contributor.googleauthorDavid Alland-
dc.identifier.doi10.1128/JCM.02271-10-
dc.admin.authorfalse-
dc.admin.mappingfalse-
dc.contributor.localIdA03824-
dc.contributor.localIdA03034-
dc.relation.journalcodeJ01325-
dc.identifier.eissn1098-660X-
dc.identifier.pmid21191047-
dc.contributor.alternativeNameCho, Sang Nae-
dc.contributor.alternativeNameLee, Eun Gae-
dc.contributor.affiliatedAuthorCho, Sang Nae-
dc.contributor.affiliatedAuthorLee, Eun Gae-
dc.rights.accessRightsfree-
dc.citation.volume49-
dc.citation.number3-
dc.citation.startPage932-
dc.citation.endPage940-
dc.identifier.bibliographicCitationJOURNAL OF CLINICAL MICROBIOLOGY, Vol.49(3) : 932-940, 2011-
dc.identifier.rimsid27219-
dc.type.rimsART-
Appears in Collections:
1. College of Medicine (의과대학) > Dept. of Microbiology (미생물학교실) > 1. Journal Papers

qrcode

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.