Evaluation of EZplex MTBC/NTM Real-Time PCR kit: diagnostic accuracy and efficacy in vaccination

Abstract

PurposeTuberculosis (TB) is mainly caused by Mycobacterium tuberculosis, which is a pathogenic mycobacterial species grouped under Mycobacterium tuberculosis complex (MTBC) with four other pathogenic mycobacterial species. The mycobacteria not included in MTBC are known as nontuberculous mycobacteria (NTM), and cause several pulmonary diseases including pneumonia. Currently, NTM occurrences in TB-suspected respiratory specimens have increased, due to which, precise detection of MTBC and NTM is considered critical for the diagnosis and vaccination of TB. Among the various methods available, real-time PCR is frequently adopted for MTBC/NTM detection due to its rapidness, accuracy, and ease of handling. In this study, we evaluated a new real-time PCR kit for analytical and clinical performance on sputum, bronchial washing, and culture specimens.Materials and MethodsFor assessing its analytical performance, limit of detection (LOD), reactivity, and repeatability test were performed using DNA samples. To evaluate clinical performance, 612 samples were collected and clinically tested at a tertiary hospital.ResultsLOD was confirmed as 0.584 copies/µL for MTBC and 47.836 copies/µL for NTM by probit analysis (95% positive). For the reactivity test, all intended strains were detected and, in the repeatability test, stable and steady results were confirmed with coefficient of variation ranging from 0.36 to 1.59. For the clinical test, sensitivity and specificity were 98.6%–100% and 98.8%–100% for MTBC and NTM, respectively.ConclusionThe results proved the usefulness of the kit in TB diagnosis. Furthermore, it could be adopted for the assessment of vaccine efficacy.

Which Current and Novel Diagnostic Avenues for Bacterial Respiratory Diseases?

“…Non-tuberculous mycobacteria (NTM, 140 species reported), that are not related to MTB but are also responsible for pneumonia such as M. avium or M. kansassi , can also be tested through Ezplex ® MTBC/NTM Real-time PCR kit. In 2 h, this kit can test samples for the presence of 100 different NTM species and shows high sensitivity and specificity (96–100%) compared to classical PCR or culture tests ( Lee et al, 2018 ).…”

Of tuberculosis and non-tuberculous mycobacterial infections – a comparative analysis of epidemiology, diagnosis and treatment

“…The EZplex MTBC/NTM is a Real-Time PCR kit used to detect Mtb complex and NTMs. This assay has an overall sensitivity and specificity of 98.6%-100% for the detection of Mtb [42]. The VereMTB Detection Kit is a NAAT based Lab-on-Chip (LoC) assay used for the detection and identification of Mtb complex, NTMs, and the diagnosis of MDR-TB.…”

“…The EZplex MTBC/NTM kit and VereMTB detection kit described earlier were also evaluated and shown to be useful for detecting NTM species directly from a sputum sample. The sensitivity and specificity of this method were 98.8%-100% for NTM [42,43]. The Genedia MTB/ NTM Detection Kit is a multiplex real-time PCR assay that targets the IS6110 region of Mtb, and the internal transcribed spacer region and the rpoB gene of NTM.…”

Development of a multiplex real-time PCR assay for the simultaneous detection of four bacterial pathogens causing pneumonia

“…In the last few decades, conventional PCR assays have been performed for rapid identification of a wide range of pathogens; however, it requires the tube to be opened for post-detection analysis, leading to putative cross-contamination of the amplicon and consequent false-positive results [ 13 ]. Recently, the TaqMan method (Applied Biosystems, Inc., CA, USA) has been developed in the multiplex real-time PCR (mRT-PCR) platform, enabling quantitative applications [ 14 ]. The mRT-PCR assay is highly sensitive, specific, rapid, and less prone to false-positives [ 13 ].…”