OR16

The knowledge about epigenetic regulation of M. tuberculosis (MTB) is still blurry. We performed RNAseq from MTB H37Rv exposed to drug-induced stress and analyzed the expression profiling of regulatory smallRNAs (sRNAs). A candidate, named ncRv0842c, antisense (AS) to the efflux pump Rv0842, emerged as a possible factor involved in rifampicin (R) resistance.

The expression of both the sRNA and its target was carried out by qPCR in different MTB ancient (L1, L5, L6) and modern (L2, L4) lineage representatives in both basal and R-induced stress. Overexpression of ncRv0842c was achieved using a pMV261 plasmid (MOCK= empty; MUT= pMV261_ncRv0842c). R minimum inhibitory concentration (MIC) was determined by MABA assay. Statistical analysis was performed by a linear regression model.

qPCR showed that the sRNA is not expressed in ancient lineages that carry a silent mutation in Rv0842 that abrogates the sRNA promoter. qPCR for H37Rv under R stress showed upregulation of Rv0842 (x3, p-val<0.05), and downregulation of the sRNA (x0.25). MOCK-L4 isolates showed even marked downregulation of the sRNA during R stress (p<0.001; β=2.5, 95% CIs 1.7;3.3). A similar trend was observed even in respective MUT-strains during the R challenge (p<0.05; β=0.9, 95% CIs 0.1;1.7). MABA performed on MUT-L4 showed a consistent 1-dilution reduction of the MIC compared to their respective MOCK, while MUT-L1 and MUT-L5 strains did not show any MIC shift.

Our results suggest a correlation between sRNAs and drug-related tolerance mechanisms induced by antibiotic treatment. Unraveling of sRNAs may provide new insights on MTB lineage-specific adaptation to drug-related stress.