Abstract
The GeneLEAD VIII (Diagenode, Belgium) is a new, fully automated, sample-to-result precision instrument for the extraction of DNA and PCR detection of Mycobacterium tuberculosis complex (MTBC) directly from clinical samples. The Deeplex Myc-TB® assay (Genoscreen, France) is a diagnostic kit based on the deep sequencing of a 24-plexed amplicon mix allowing simultaneously the detection of resistance to 13 antituberculous (antiTB) drugs and the determination of spoligotype. We evaluated the performance of a strategy combining the both mentioned tools to detect directly from clinical samples, in 8 days, MTBC and its resistance to 13 antiTB drugs, and identify potential transmission of strains from patient-to-patient. Using this approach, we screened 112 clinical samples (65 smear-negative) and 94 MTBC cultured strains. The sensitivity and the specificity of the GeneLEAD/Deeplex Myc-TB approach for MTBC detection were 79.3% and 100%, respectively. One hundred forty successful Deeplex Myc-TB results were obtained for 46 clinical samples and 94 strains, a total of 85.4% of which had a Deeplex Myc-TB susceptibility and resistance prediction consistent with phenotypic drug susceptibility testing (DST). Importantly, the Deeplex Myc-TB assay was able to detect 100% of the multidrug-resistant (MDR) MTBC tested. The lowest concordance rates were for pyrazinamide, ethambutol, streptomycin, and ethionamide (84.5%, 81.5%, 73%, and 55%, respectively) for which the determination of susceptibility or resistance is generally difficult with current tools. One of the main difficulties of Deeplex Myc-TB is to interpret the non-synonymous uncharacterized variants that can represent up to 30% of the detected single nucleotide variants. We observed a good level of concordance between Deeplex Myc-TB-spoligotyping and MIRU-VNTR despite a lower discriminatory power for spoligotyping. The median time to obtain complete results from clinical samples was 8 days (IQR 7–13) provided a high-throughput NGS sequencing platform was available. Our results highlight that the GeneLEAD/Deeplex Myc-TB approach could be a breakthrough in rapid diagnosis of MDR TB in routine practice.
Highlights
In 2019, tuberculosis (TB) was responsible for more deaths than any other infectious disease in the world (Furin et al, 2019)
The bacterial load determined by microscopic examination of the 112 clinical specimens ranged from ≥100 acid-fast bacilli (AFB) per field (n = 10), 10 to 99 AFB per field (n = 18), 1 to 9 AFB per field (n = 12), to 10 on the slide (n = 7)
We assessed the performance of GeneLEAD combined to Deeplex Myc-TB test for identifying Mycobacterium tuberculosis complex (MTBC), predicting antiTB drug susceptibility and MTBC genotyping
Summary
In 2019, tuberculosis (TB) was responsible for more deaths than any other infectious disease in the world (Furin et al, 2019). DNA strip testing such as the MTBDRplus and MTBDRsl tests (Hain, Germany), and Sanger sequencing of genes involved in drug resistance can detect drug resistance mutations (Brossier et al, 2009; Brossier et al, 2016; Brossier et al, 2017). Both methods remain technically time-consuming, expensive, complex, and generally need to be performed on cultures to give good results. Some progress remains to be made in terms of the timeliness of results, performance when applied directly on clinical samples/data interpretation even if large systematic reviews have been published, and standardization to implement WGS in the routine clinical setting (Votintseva et al, 2017; Doyle et al, 2018; Meehan et al, 2019; World Health Organization, 2021)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
