P079

1:Institute of Tropical Medicine Antwerp; 2:Nottingham Trent University; 3:Antwerp University

Bedaquiline (BDQ) is essential for treating rifampicin-resistant tuberculosis (RR-TB), but its long-term efficacy is threatened by emerging resistance. Although clinical resistance is linked to mutations in mmpR5, a repressor of the mmpS5/mmpL5 efflux pump, the genotype–phenotype correlation remains challenging, suggesting additional mechanisms.

To explore the impact of diverse mmpR5 mutations and identify potential regulatory mechanisms, RNA-sequencing was performed on four BDQ-resistant strains (344delA, L43R, A84T, and the double mutant E138G/M139L) and three BDQ-susceptible strains. Samples were analyzed before and after BDQ exposure (0.5 µg/mL for 30 minutes and 2 hours).

Three of four resistant strains exhibited overexpression of the mmpS5/mmpL5 efflux system (padj<0.05). One resistant strain showed no significant upregulation (padj>0.05), suggesting alternative resistance pathways. Transcriptomic analysis revealed 711 differentially expressed genes (log2FC < -1 or >1) in resistant versus susceptible strains prior to BDQ exposure. Changes included upregulation of other efflux genes, increased lipid transport, redox stress tolerance, and respiratory chain reprogramming. Notably, all resistant strains upregulated cytochrome bd oxidase, suggesting a compensatory mechanism. At the concentration tested, BDQ exposure did not significantly alter mmpR5, mmpS5, or mmpL5 expression or pathway enrichment (padj>0.05).

These findings suggest BDQ resistance may result from both constitutive mmpS5/mmpL5 activation and broader metabolic adaptations. Further omics studies are needed to clarify these mechanims.