Unexpected high prevalence of resistance-associated Rv0678 variants in MDR-TB patients without documented prior use of clofazimine or bedaquiline

Abstract

Objectives: Resistance-associated variants (RAVs) in Rv0678 , a regulator of the MmpS5-MmpL5 efflux pump, have been shown to lead to increased MICs of bedaquiline (2- to 8- fold) and clofazimine (2- to 4-fold). The prevalence of these Rv0678 RAVs in clinical isolates and their impact on treatment outcomes are important factors to take into account in bedaquiline treatment guidelines.

Methods: Baseline isolates from two bedaquiline MDR-TB clinical trials were sequenced for Rv0678 RAVs and corresponding bedaquiline MICs were determined on 7H11 agar. Rv0678 RAVs were also investigated in non-MDR-TB sequences of a population-based cohort.

Results: Rv0678 RAVs were identified in 23/347 (6.3%) of MDR-TB baseline isolates. Surprisingly, bedaquiline MICs for these isolates were high (> 0.24 mg/L, n = 8), normal (0.03-0.24 mg/L, n = 11) or low (< 0.03 mg/L, n = 4). A variant at position -11 in the intergenic region mmpS5 - Rv0678 was identified in 39 isolates (11.3%) and appeared to increase the susceptibility to bedaquiline. In non-MDR-TB isolates, the frequency of Rv0678 RAVs was lower (6/852 or 0.7%). Competition experiments suggested that rifampicin was not the drug selecting for Rv0678 RAVs.

Conclusions: RAVs in Rv0678 occur more frequently in MDR-TB patients than previously anticipated, are not associated with prior use of bedaquiline or clofazimine, and in the majority of cases do not lead to bedaquiline MICs above the provisional breakpoint (0.24 mg/L). Their origin remains unknown. Given the variety of RAVs in Rv0678 and their variable effects on the MIC, only phenotypic drug-susceptibility methods can currently be used to assess bedaquiline susceptibility.

Figure 1

Mutations in Rv0678 and the intergenic region mmpS5–Rv0678 and lineage of their respective isolates. All the different DNA mutations found in C208 and C209 baseline isolates (MDR, purple markers) and in the Hamburg cohort (non-MDR, orange markers) are shown. Silent mutations are not shown. Mutations in isolates with high MICs are coloured in pink. The circle next to every mutation indicates the lineage of the isolate: blue, lineage 1; red, lineage 2; yellow, lineage 3; and green, lineage 4. When the mutation was found in more than one isolate, the number of isolates carrying that mutation is indicated in brackets. Some isolates carry more than one mutation, and the mutations found in the same isolate are indicated with the same symbol (*, ** or ***). The mutations co-existing with the mutation in position −11 are marked with the symbol ^.