Review 4: "Xpert MTB/RIF Ultra Resistant and MTBDRplus Susceptible Rifampicin Results in People with Tuberculosis: Utility of FluoroType MTBDR and Deep Sequencing"

Bordin Butr-Indr

doi:doi:10.1162/2e3983f5.c2fd8b5d

Xpert MTB/RIF Ultra resistant and MTBDRplus susceptible rifampicin results in people with tuberculosis: utility of FluoroType MTBDR and deep sequencing

by Yonas Ghebrekristos, Aysha Ahmed, Natalie Beylis, Sarishna Singh, Christoffel Opperman, Fahd Naufal, Megan Folkerts, David Engelthaler, Erick Auma, Rouxjeane Venter, Ghowa Booley, John Metcalfe, Robin Warren, and Grant Theron

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Published on Oct 30, 2024
www.medrxiv.org

Description

Summary Background Xpert MTB/RIF Ultra (Ultra)-detected rifampicin-resistant tuberculosis (TB) is often programmatically confirmed using MTBDRplus. There are limited data on discordant results, including re-tested using newer methods like FluoroType MTBDR (FT-MTBDR) and targeted deep sequencing.Methods MTBDRplus rifampicin-susceptible isolates from people with Ultra rifampicin-resistant sputum were identified from a South African programmatic laboratory. FT-MTBDR and single molecule-overlapping reads deep (SMOR; rpoB, inhA, katG) on isolate DNA were done (SMOR reference standard).Findings Between 01/04/2021-30/09/2022, 8% (109/1347) of Ultra rifampicin-resistant specimens were MTBDRplus-susceptible. Of 89% (97/109) isolates with a sequenceable rpoB, SMOR resolved most in favour of Ultra [79% (77/97)]. Sputum with lower mycobacterial load was associated with Ultra false-positive resistance [46% (11/24) of “very low” Ultras had false-resistance vs. 12% (9/73; p=0.0004) in those ≥“low”], as were Ultra heteroresistance calls (all wild type probes, ≥1 mutant probe) [62% (23/37 vs. 25% (15/60) for Ultra without heteroresistance calls; p=0.0003]. Of the 91% (88/97) of isolates successfully tested by FT-MTBDR, 55% (48/88) were FT-MTBDR rifampicin-resistant and 45% (40/88) susceptible, translating to 69% (47/68) sensitivity and 95% (19/20) specificity. In the 91% (99/109) of isolates with inhA and katG sequenced, 62% (61/99) were SMOR isoniazid-susceptible.Interpretation When Ultra and MTBDRplus rifampicin results are discordant, Ultra is more likely to be correct and FT-MTBDR agrees more with Ultra than MTBDRplus, however, lower load and the Ultra heteroresistance probe pattern were risk factors for Ultra false rifampicin-resistant results. Most people with Ultra-MTBDRplus discordant resistance results were isoniazid-susceptible. These data have implications for drug-resistant TB diagnosis.Funding This work was supported by European & Developing Countries Trial Partnerships (EDCTP2; RIA2020I-3305, CAGE-TB), National Institutes of Health (D43TW010350; U01AI152087; U54EB027049; R01AI136894).

RR\ID Evidence Scale rating by reviewer:

Strong. The main study claims are very well-justified by the data and analytic methods used. There is little room for doubt that the study produced has very similar results and conclusions as compared with the hypothetical ideal study. The study’s main claims should be considered conclusive and actionable without reservation.

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Review: The content of the manuscript is appropriate. Research indicates some assumptions and also affects the development of tuberculosis testing in the future. The experiment's population was sufficient enough to conclude the study. The study will lead to future tuberculosis diagnostic tests.

Some Comments:

Figure 1. The study profile. Why wasn’t FT-MTBDR used with all MTBDRplus: rifampicin-susceptible (109/1347).
From the results of selection by FT-MTBDR, gradually testing the rifampicin resistance group by SMOR should make the differences in the HR group even more visible. In addition, when criticizing the study results, it should be mentioned that the target genes of interest in each individual test have limitations.

Overall the main takeaway is that in order to prevent the spread of tuberculosis, it is crucial to research techniques for precise and transparent testing. Furthermore, ongoing research into the fundamental pathogenicity of tuberculosis is necessary, as is long-term education.

Review 4: "Xpert MTB/RIF Ultra Resistant and MTBDRplus Susceptible Rifampicin Results in People with Tuberculosis: Utility of FluoroType MTBDR and Deep Sequencing"

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