Development of multiplex PCR assays based on the 16S-23S rRNA internal transcribed spacer for the detection of clinically relevant nontuberculous mycobacteria

Aims: To accelerate the identification and differentiation of clinically relevant nontuberculous mycobacteria (NTM) with two sets of multiplex PCR (mPCR) targeting the 16S-23S rRNA internal transcribed spacer (ITS) region for timely patient management. Methods and Results: Two mPCR assays were devel...

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Veröffentlicht in:Letters in applied microbiology 2011-05, Vol.52 (5), p.546-554
Hauptverfasser: Ngan, G.J.Y, Ng, L.M, Jureen, R, Lin, R.T.P, Teo, J.W.P
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Ng, L.M
Jureen, R
Lin, R.T.P
Teo, J.W.P
description Aims: To accelerate the identification and differentiation of clinically relevant nontuberculous mycobacteria (NTM) with two sets of multiplex PCR (mPCR) targeting the 16S-23S rRNA internal transcribed spacer (ITS) region for timely patient management. Methods and Results: Two mPCR assays were developed: Slow-Growers (SG) mPCR was used for the detection of slow-growing mycobacteria, which included Mycobacterium avium complex, Mycobacterium kansasii, Mycobacterium gordonae and Mycobacterium xenopi whilst the other mPCR assay labelled as Fast-Growers (FG) mPCR was used for the detection of Mycobacterium fortuitum complex, Mycobacterium abscessus and Mycobacterium chelonae. In these assays, a common forward primer based on a conserved section of the 16S rRNA region was used in conjunction with species-specific reverse primers. The mPCRs were tested against 247 clinical mycobacterial isolates and demonstrated 100% specificity and sensitivity. Identification of the mycobacterial species was also validated by DNA sequencing of the 16S-23S ITS region and when further confirmation was needed, hsp65 sequencing was performed. Conclusions: The mPCR assays could be a potentially useful diagnostic tool for the rapid and accurate identification of clinically relevant NTM. Significance and Impact of the Study: In this study, we looked at the frequency of hospital isolated NTM over the last 5 years (2005-2010), and an mPCR targeting the ITS region was developed for NTM species that appeared to be more prevalent in the context of Singapore.
doi_str_mv 10.1111/j.1472-765X.2011.03045.x
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Psychology</topic><topic>Humans</topic><topic>internal transcribed spacers</topic><topic>Microbiology</topic><topic>mycobacteria</topic><topic>Mycobacterium - classification</topic><topic>Mycobacterium - genetics</topic><topic>Mycobacterium - isolation &amp; purification</topic><topic>Mycobacterium abscessus</topic><topic>Mycobacterium avium complex</topic><topic>Mycobacterium chelonae</topic><topic>Mycobacterium fortuitum</topic><topic>Mycobacterium gordonae</topic><topic>Mycobacterium Infections - diagnosis</topic><topic>Mycobacterium Infections - microbiology</topic><topic>Mycobacterium kansasii</topic><topic>Mycobacterium xenopi</topic><topic>Polymerase Chain Reaction</topic><topic>Reproducibility of Results</topic><topic>ribosomal RNA</topic><topic>RNA, Ribosomal, 16S - genetics</topic><topic>RNA, Ribosomal, 23S - genetics</topic><topic>Sensitivity and Specificity</topic><topic>Singapore</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Ngan, G.J.Y</creatorcontrib><creatorcontrib>Ng, L.M</creatorcontrib><creatorcontrib>Jureen, R</creatorcontrib><creatorcontrib>Lin, R.T.P</creatorcontrib><creatorcontrib>Teo, J.W.P</creatorcontrib><collection>AGRIS</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>MEDLINE - Academic</collection><jtitle>Letters in applied microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ngan, G.J.Y</au><au>Ng, L.M</au><au>Jureen, R</au><au>Lin, R.T.P</au><au>Teo, J.W.P</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Development of multiplex PCR assays based on the 16S-23S rRNA internal transcribed spacer for the detection of clinically relevant nontuberculous mycobacteria</atitle><jtitle>Letters in applied microbiology</jtitle><addtitle>Lett Appl Microbiol</addtitle><date>2011-05</date><risdate>2011</risdate><volume>52</volume><issue>5</issue><spage>546</spage><epage>554</epage><pages>546-554</pages><issn>0266-8254</issn><eissn>1472-765X</eissn><coden>LAMIE7</coden><abstract>Aims: To accelerate the identification and differentiation of clinically relevant nontuberculous mycobacteria (NTM) with two sets of multiplex PCR (mPCR) targeting the 16S-23S rRNA internal transcribed spacer (ITS) region for timely patient management. 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Conclusions: The mPCR assays could be a potentially useful diagnostic tool for the rapid and accurate identification of clinically relevant NTM. Significance and Impact of the Study: In this study, we looked at the frequency of hospital isolated NTM over the last 5 years (2005-2010), and an mPCR targeting the ITS region was developed for NTM species that appeared to be more prevalent in the context of Singapore.</abstract><cop>Oxford, UK</cop><pub>Blackwell Publishing Ltd</pub><pmid>21395629</pmid><doi>10.1111/j.1472-765X.2011.03045.x</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record>
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source MEDLINE; Wiley Online Library Journals Frontfile Complete; Oxford University Press Journals All Titles (1996-Current); EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection
subjects Bacteriological Techniques - methods
Biological and medical sciences
diagnosis
DNA, Ribosomal Spacer - genetics
Fundamental and applied biological sciences. Psychology
Humans
internal transcribed spacers
Microbiology
mycobacteria
Mycobacterium - classification
Mycobacterium - genetics
Mycobacterium - isolation & purification
Mycobacterium abscessus
Mycobacterium avium complex
Mycobacterium chelonae
Mycobacterium fortuitum
Mycobacterium gordonae
Mycobacterium Infections - diagnosis
Mycobacterium Infections - microbiology
Mycobacterium kansasii
Mycobacterium xenopi
Polymerase Chain Reaction
Reproducibility of Results
ribosomal RNA
RNA, Ribosomal, 16S - genetics
RNA, Ribosomal, 23S - genetics
Sensitivity and Specificity
Singapore
title Development of multiplex PCR assays based on the 16S-23S rRNA internal transcribed spacer for the detection of clinically relevant nontuberculous mycobacteria
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