Inclusion of minor alleles improves catalogue-based prediction of fluoroquinolone resistance in Mycobacterium tuberculosis

Objectives Fluoroquinolone resistance poses a threat to the successful treatment of tuberculosis. WGS, and the subsequent detection of catalogued resistance-associated mutations, offers an attractive solution to fluoroquinolone susceptibility testing but sensitivities are often less than 90%. We hypothesize that this is partly because the bioinformatic pipelines used usually mask the recognition of minor alleles that have been implicated in fluoroquinolone resistance. Methods We analysed the Comprehensive Resistance Prediction for Tuberculosis: an International Consortium (CRyPTIC) dataset of globally diverse WGS Mycobacterium tuberculosis isolates, with matched MICs for two fluoroquinolone drugs and allowed putative minor alleles to contribute to resistance prediction. Results Detecting minor alleles increased the sensitivity of WGS for moxifloxacin resistance prediction from 85.4% to 94.0%, without significantly reducing specificity. We also found no correlation between the proportion of an M. tuberculosis population containing a resistance-conferring allele and the magnitude of resistance. Conclusions Together our results highlight the importance of detecting minor resistance-conferring alleles when using WGS, or indeed any sequencing-based approach, to diagnose fluoroquinolone resistance.

Automated summary

Fluoroquinolone resistance is a threat to tuberculosis treatment, and Whole Genome Sequencing (WGS) with detection of catalogued resistance-associated mutations is a promising method for susceptibility testing. However, the sensitivity of this method is often less than 90% due to the masking of minor alleles. It is important to detect these minor resistance-conferring alleles in WGS or any sequencing-based approach for diagnosing fluoroquinolone resistance. Importance: rating 7/10.