Potentiation of the antimycobacterial activity of bedaquiline, clofazimine, and doxycycline against Mycobacterium smegmatis by several natural product-based compounds is putatively via efflux inhibition

<ns3:p> Background <ns3:italic>Mycobacterium tuberculosis</ns3:italic> ( <ns3:italic>Mtb</ns3:italic> ), the causative agent of tuberculosis (TB), is the leading cause of death due to a single infectious pathogen globally. The increasing prevalence of drug-resistant <ns3:italic>Mtb</ns3:italic> strains underscores the pressing need for the development of new antimycobacterial drugs with novel mechanisms of action. Targeting pathogen drug efflux, a key antimycobacterial drug resistance mechanism, is an attractive, viable strategy for the development of new TB therapeutics. Methods In this study, we utilised <ns3:italic>Mycobacterium smegmatis</ns3:italic> ( <ns3:italic>Msm</ns3:italic> ), a non-pathogenic <ns3:italic>Mtb</ns3:italic> surrogate, to delineate the ability of natural-product based compounds to augment the efficacy of bedaquiline (BDQ), clofazimine (CFZ) and doxycycline (DOX), probably via efflux inhibition (EI). Literature reporting the plant sources of the known efflux inhibitors (EIs) reserpine (RES), berberine (BER) and piperine (PIP) was scoped and additional compounds, (+)-lyoniresinol-3-Alpha-O-Beta-D-glucopyranoside (LYO-3) and lyoniresinol (LYO), isolated from the same plant species were chosen for testing. <ns3:italic>In vitro</ns3:italic> screening of the selected compounds was performed using the two-dimensional (2-D) checkerboard assay in which each likely efflux disruptor was tested in combination with BDQ, CFZ and DOX against <ns3:italic>Msm</ns3:italic> and the effect of the combinations ascertained. Thereafter, compounds that exhibited probable EI activity were docked onto potential targets namely MSMEG_5187, a <ns3:italic>Msm</ns3:italic> homologue of <ns3:italic>Mtb</ns3:italic> Rv1258c efflux pump (EP) and MmpL5. Results Molecular docking revealed that the EIs avidly bound to the EPs with docking scores of &lt;-7 kcal/mol while the checkerboard combination assays demonstrated strong synergistic interactions for BDQ plus BER, LYO-3 and LYO. Conclusion These results point to a probable disruption of the <ns3:italic>Msm</ns3:italic> efflux system by <ns3:italic>Berberis species</ns3:italic> derivatives. </ns3:p>