Detection of Mycobacterium tuberculosis in Respiratory Specimens Using IS6110-PCR Based Method with New Designed Primers

Name: Unchalee Tansuphasiri

Name: Kanda Vathanophas

Name: Anong Pariyanonda

Name: Charnchudhi Chanyasanha

Organization : Mahidol University. Faculty of Public Health. Department of Microbiology

MeSH: Mycobacterium tuberculosis

keyword: Detection

; IS6110-PCR based method

Abstract: We have investigated the potential use of the IS6110-PCR based method using a new designed primer pair that amplifies a 377-bp target located at the end region of the insertion sequence IS6110 for the rapid detection of Mycobacterium tuberculosis in clinical sputum specimens. When tested on purified genomic DNA the method was highly specific for TB complex and was sensitive enough to detect as little as 10 fg of DNA, which is equivalent to approximately three bacilli. The test was applied to 328 sputum specimens collected from patients and results were compared with those of acid-fast staining and culture, where the culture is considered as the "gold standard." Of these 328 specimens, 134 (41 % prevalence rate) were positive for M. tuberculosis by culture. For all specimens tested, the sensitivity, specificity, positive predictive value, negative predictive value, and efficiency of the test were 96.27, 99.48, 99.23, 97.47, and 98.17%, respectively. There is no significant difference in the performances of IS6110- PCR based method and culture (P>0.05). Sensitivities of 100 and 88.89% were obtained for smear-positive and smear-negative specimens, respectively. The corresponding specificities were 100 and 99.48%. Thus the IS6110-PCR based method using this primer set with the optimized condition and specific PCR product detection by gel electrophoresis and confirmation by Southern hybridization, was able to detect M. tuberculosis directly from sputa. For routine clinical practice, however further improvements in the DNA extraction step to remove inhibitors, and in the PCR detection step for solving the problems of its cumbersome and high expense are needed

Mahidol University. Mahidol University Library and Knowledge Center

Address: NAKHONPATHOM

Email: liwww@mahidol.ac.th

Created: 2013

Modified: 2013-03-18

Issued: 1999-07/12

บทความ/Article

application/pdf

Mahidol Journal. 6, 2 (July-Dec. 1999) 71-76

eng

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