The 2021 WHO catalogue of Mycobacterium tuberculosis complex mutations associated with drug resistance: A genotypic analysis

Abstract

Background: Molecular diagnostics are considered the most promising route to achieving rapid, universal drug susceptibility testing for Mycobacterium tuberculosiscomplex (MTBC). We aimed to generate a WHO endorsed catalogue of mutations to serve as a global standard for interpreting molecular information for drug resistance prediction.

Methods: A candidate gene approach was used to identify mutations as associated with resistance, or consistent with susceptibility, for 13 WHO endorsed anti-tuberculosis drugs. 38,215 MTBC isolates with paired whole-genome sequencing and phenotypic drug susceptibility testing data were amassed from 45 countries. For each mutation, a contingency table of binary phenotypes and presence or absence of the mutation computed positive predictive value, and Fisher's exact tests generated odds ratios and Benjamini-Hochberg corrected p-values. Mutations were graded as Associated with Resistance if present in at least 5 isolates, if the odds ratio was >1 with a statistically significant corrected p-value, and if the lower bound of the 95% confidence interval on the positive predictive value for phenotypic resistance was >25%. A series of expert rules were applied for final confidence grading of each mutation.

Findings: 15,667 associations were computed for 13,211 unique mutations linked to one or more drugs. 1,149/15,667 (7·3%) mutations were classified as associated with phenotypic resistance and 107/15,667 (0·7%) were deemed consistent with susceptibility. For rifampicin, isoniazid, ethambutol, fluoroquinolones, and streptomycin, the mutations' pooled sensitivity was >80%. Specificity was over 95% for all drugs except ethionamide (91·4%), moxifloxacin (91·6%) and ethambutol (93·3%). Only two resistance mutations were classified for bedaquiline, delamanid, clofazimine, and linezolid as prevalence of phenotypic resistance was low for these drugs.

Interpretation: This first WHO endorsed catalogue of molecular targets for MTBC drug susceptibility testing provides a global standard for resistance interpretation. Its existence should encourage the implementation of molecular diagnostics by National Tuberculosis Programmes.

Funding: UNITAID, Wellcome, MRC, BMGF.

Figure 1

Number of tier 1 mutations by mutation group and drug Genes were divided into two tiers for analysis, in which tier 1 sequences were deemed to have a higher probability of association with resistance. For isolates with no tier 1 genomic explanation for resistance, tier 2 sequences were analysed. No tier 2 mutations were graded as group 1 or 2 so these are not shown for any groups. To reflect the minimum number of isolates a mutation must have been seen in for it to be graded as group 1 or 2, the mutations used here were seen in 5 or more isolates. Exceptions are mutations relevant to pyrazinamide (counted if seen in 2 or more isolates) and mutations subject to an expert rule (counted if seen in any number of isolates). The actual number of tier 1 group 3 mutations, regardless of the frequency with which these were seen, is written within each green bar. Group 4 mutations graded as such by an expert rule are not shown separately here, but can be viewed in appendix 2 (table S4). All expert rule mutations were group 2, with the exception of rpoB borderline mutations. Group 1=associated with resistance. Group 2=associated with resistance—interim. Group 3=uncertain significance. Group 4=not associated with resistance—interim. Group 5=not associated with resistance.

Figure 2

Number of isolates per mutation by mutation group and drug Error bars indicate upper and lower quartiles. Box and whisker plots exclude outside values. Mutations graded by expert rules are not shown as the number of isolates was not relevant to their classification. Group 1=associated with resistance. Group 2=associated with resistance—interim. Group 3=uncertain significance. Group 4=not associated with resistance—interim. Group 5=not associated with resistance.

Figure 3

Sensitivity and specificity for all drugs Sensitivity is represented by bars going upwards from zero, and specificity by bars going downwards from 100. The colour progression in each bar shows the incremental sensitivity gained, and corresponding specificity lost, by expanding the catalogue to include first group 1 and then group 2 mutations, in each case without the use of expert rules, and then adding in the expert rules. All mutations subject to an expert rule were graded as group 2 except for borderline rpoB mutations. Error bars indicate 95% CIs for the total effect of all mutations shown. Group 1=associated with resistance. Group 2=associated with resistance—interim.

Figure 4

Total number of isolates for each drug by sensitivity Each drug is represented by a coloured circle weighted by the prevalence of phenotypic resistance to that drug. The centre of each circle shows the intersection between values on the x-axis and y-axis. Group 1=associated with resistance. Group 2=associated with resistance—interim.