Reliability of nucleic acid amplification for detection of Mycobacterium tuberculosis: an international collaborative quality control study among 30 laboratories

Abstract

Nucleic acid amplification to detect Mycobacterium tuberculosis in clinical specimens is increasingly used as a laboratory tool for the diagnosis of tuberculosis. However, the specificity and sensitivity of these tests may be questioned, and no standardized reagents for quality control assessment are available. To estimate the performance of amplification tests for routine diagnosis, we initiated an interlaboratory study involving 30 laboratories in 18 countries. We prepared blinded panels of 20 sputum samples containing no, 100, or 1,000 mycobacterial cells. Each laboratory was asked to detect M. tuberculosis by their routine method of nucleic acid amplification. Only five laboratories correctly identified the presence or absence of mycobacterial DNA in all 20 samples. Seven laboratories detected mycobacterial DNA in all positive samples, and 13 laboratories correctly reported the absence of DNA in the negative samples. Lack of specificity was more of a problem than lack of sensitivity. Reliability was not found to be associated with the use of any particular method. Reliable detection of M. tuberculosis in clinical samples by nucleic acid amplification techniques is possible, but many laboratories do not use adequate quality controls. This study underlines the need for good laboratory practice and reference reagents to monitor the performance of the whole assay, including pretreatment of clinical samples.

Operational Feasibility of Using Loop-Mediated Isothermal Amplification for Diagnosis of Pulmonary Tuberculosis in Microscopy Centers of Developing Countries

“…The complexity and insufficient robustness of existing commercial NAAT formats and their requirement for a precision instru-ment, a high degree of technical support, and quality assurance make them unsuitable for most developing country settings (8,12). Furthermore, the technical skill required has resulted in variable performance even in experienced molecular laboratories in both developed and developing countries (9,20).…”

Examination of specimens for mycobacteria in clinical laboratories in 21 countries: a 10-year review of the UK National Quality Assessment Scheme for Mycobacteria Culture

“…Increasingly, nucleic acid probes are being used for the identification of mycobacteria from positive cultures [10,11,[13][14][15][16]. The methods survey indicated that only 20% of respondents used molecular techniques to identify isolates of mycobacteria.…”

“…The methods survey indicated that only 20% of respondents used molecular techniques to identify isolates of mycobacteria. Although PCR and other nucleic acid amplification techniques are rapid, specific and relatively sensitive, there is a cost implication, and a certain degree of expertise is required in the performance of these tests [6,16]. In this connection, Moströ m et al [20] have highlighted the need for training and education in molecular biology.…”

Multicentre quality control study for detection of Mycobacterium tuberculosis in clinical samples by nucleic amplification methods

“…Overall, the results of the present study were an improvement compared to previous studies on the performance of NATs for the detection of M. tuberculosis. Methods have improved and nowadays many more laboratories are using commercial tests, i.e., 62.4% vs. 26.7% in 1996 [3]. The commercial tests have quality-controlled reagents and, theoretically, should be less prone to contamination than most in-house methods.…”