Novel approach based on one-tube nested PCR and a lateral flow strip for highly sensitive diagnosis of tuberculous meningitis

Rapid and sensitive detection of Mycobacterium tuberculosis (M. Tb) in cerebrospinal fluid is crucial in the diagnosis of tuberculous meningitis (TBM), but conventional diagnostic technologies have limited sensitivity and specificity or are time-consuming. In this work, a novel, highly sensitive mol...

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Bibliographic Details
Published in:PloS one Vol. 12; no. 10; p. e0186985
Main Authors: Sun, Yajuan, Chen, Jiajun, Li, Jia, Xu, Yawei, Jin, Hui, Xu, Na, Yin, Rui, Hu, Guohua
Format: Journal Article
Language:English
Published: United States Public Library of Science 30.10.2017
Public Library of Science (PLoS)
Subjects:
Biology and Life Sciences
Complications and side effects
Diagnosis
Genetic aspects
Humans
Medicine and Health Sciences
Meningitis
Polymerase chain reaction
Polymerase Chain Reaction - methods
Research and Analysis Methods
Risk factors
Sensitivity and Specificity
Tuberculosis
Tuberculosis, Meningeal - diagnosis
China
ISSN:1932-6203, 1932-6203
Online Access:Get full text
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Summary:Rapid and sensitive detection of Mycobacterium tuberculosis (M. Tb) in cerebrospinal fluid is crucial in the diagnosis of tuberculous meningitis (TBM), but conventional diagnostic technologies have limited sensitivity and specificity or are time-consuming. In this work, a novel, highly sensitive molecular diagnostic method, one-tube nested PCR-lateral flow strip test (OTNPCR-LFST), was developed for detecting M. tuberculosis. This one-tube nested PCR maintains the sensitivity of conventional two-step nested PCR and reduces both the chance of cross-contamination and the time required for analysis. The PCR product was detected by a lateral flow strip assay, which provided a basis for migration of the test to a point-of-care (POC) microfluidic format. The developed assay had an improved sensitivity compared with traditional PCR, and the limit of detection was up to 1 fg DNA isolated from M. tuberculosis. The assay was also specific for M. tuberculosis, and no cross-reactions were found in other non-target bacteria. The application of this technique to clinical samples was successfully evaluated, and OTNPCR-LFST showed 89% overall sensitivity and 100% specificity for TBM patients. This one-tube nested PCR-lateral flow strip assay is useful for detecting M. tuberculosis in TBM due to its rapidity, high sensitivity and simple manipulation.
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Competing Interests: The authors have declared that no competing interests exist.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0186985