Cargando…
A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction
We report a novel approach utilising a real-time PCR screening assay targeting a 53 bp tandemly repeated element present at various loci within the Mycobacterium tuberculosis (Mtb) genome. Positive samples were identified within a discriminatory melting curve range of 90–94°C, with results obtained...
Autores principales: | , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Hindawi Publishing Corporation
2015
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4745473/ https://www.ncbi.nlm.nih.gov/pubmed/26904747 http://dx.doi.org/10.1155/2015/593745 |
_version_ | 1782414655502680064 |
---|---|
author | Cross, Lisa Jane Anscombe, Catherine McHugh, Timothy D. Abubakar, Ibrahim Shorten, Robert John Thorne, Nicola Arnold, Cath |
author_facet | Cross, Lisa Jane Anscombe, Catherine McHugh, Timothy D. Abubakar, Ibrahim Shorten, Robert John Thorne, Nicola Arnold, Cath |
author_sort | Cross, Lisa Jane |
collection | PubMed |
description | We report a novel approach utilising a real-time PCR screening assay targeting a 53 bp tandemly repeated element present at various loci within the Mycobacterium tuberculosis (Mtb) genome. Positive samples were identified within a discriminatory melting curve range of 90–94°C, with results obtained in under one hour directly from decontaminated sputum samples without extraction. A panel of 89 smear-positive sputa were used for analytical validation of the assay with 100% concordance, with sensitivity matching that of culture. Cross reactivity was detected within a narrow range of mycobacteria other than tuberculosis (MOTT) (five sputa, three in silico), with the highest sensitivity within M. avium complex (MAC). A year-long head to head evaluation of the test with the GeneXpert platform was carried out with 104 consecutive samples at the Royal Free Hospital, UK. Receiver operating characteristics (ROC) analysis of the data revealed that the two tests are approximately equivalent in sensitivity, with the area under the curve being 0.85 and 0.80 for the GeneXpert and our assay, respectively, indicating that the test would be a cost effective screen prior to GeneXpert testing. |
format | Online Article Text |
id | pubmed-4745473 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | Hindawi Publishing Corporation |
record_format | MEDLINE/PubMed |
spelling | pubmed-47454732016-02-22 A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction Cross, Lisa Jane Anscombe, Catherine McHugh, Timothy D. Abubakar, Ibrahim Shorten, Robert John Thorne, Nicola Arnold, Cath Int J Bacteriol Research Article We report a novel approach utilising a real-time PCR screening assay targeting a 53 bp tandemly repeated element present at various loci within the Mycobacterium tuberculosis (Mtb) genome. Positive samples were identified within a discriminatory melting curve range of 90–94°C, with results obtained in under one hour directly from decontaminated sputum samples without extraction. A panel of 89 smear-positive sputa were used for analytical validation of the assay with 100% concordance, with sensitivity matching that of culture. Cross reactivity was detected within a narrow range of mycobacteria other than tuberculosis (MOTT) (five sputa, three in silico), with the highest sensitivity within M. avium complex (MAC). A year-long head to head evaluation of the test with the GeneXpert platform was carried out with 104 consecutive samples at the Royal Free Hospital, UK. Receiver operating characteristics (ROC) analysis of the data revealed that the two tests are approximately equivalent in sensitivity, with the area under the curve being 0.85 and 0.80 for the GeneXpert and our assay, respectively, indicating that the test would be a cost effective screen prior to GeneXpert testing. Hindawi Publishing Corporation 2015 2015-01-06 /pmc/articles/PMC4745473/ /pubmed/26904747 http://dx.doi.org/10.1155/2015/593745 Text en Copyright © 2015 Lisa Jane Cross et al. https://creativecommons.org/licenses/by/3.0/ This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Cross, Lisa Jane Anscombe, Catherine McHugh, Timothy D. Abubakar, Ibrahim Shorten, Robert John Thorne, Nicola Arnold, Cath A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title | A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title_full | A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title_fullStr | A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title_full_unstemmed | A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title_short | A Rapid and Sensitive Diagnostic Screening Assay for Detection of Mycobacteria Including Mycobacterium tuberculosis Directly from Sputum without Extraction |
title_sort | rapid and sensitive diagnostic screening assay for detection of mycobacteria including mycobacterium tuberculosis directly from sputum without extraction |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4745473/ https://www.ncbi.nlm.nih.gov/pubmed/26904747 http://dx.doi.org/10.1155/2015/593745 |
work_keys_str_mv | AT crosslisajane arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT anscombecatherine arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT mchughtimothyd arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT abubakaribrahim arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT shortenrobertjohn arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT thornenicola arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT arnoldcath arapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT crosslisajane rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT anscombecatherine rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT mchughtimothyd rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT abubakaribrahim rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT shortenrobertjohn rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT thornenicola rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction AT arnoldcath rapidandsensitivediagnosticscreeningassayfordetectionofmycobacteriaincludingmycobacteriumtuberculosisdirectlyfromsputumwithoutextraction |