Investigation of efflux pump genes in isoniazid resistant Mycobacterium tuberculosis isolates

Background: Tuberculosis (TB) is one of the most important infectious diseases worldwide. Resistance to antituberculosis drugs develops because of genetic mutations that render drug-activating enzymes inactive, changes in cell wall permeability, and increased expression of efflux pump genes and also combination therapy with efflux pump inhibitors may be more effective in drug-resistant TB patients.

Aims: To investigate the effect of verapamil (VR) on isonicotinic acid hydrazide (INH) resistance and the expression of 21 efflux pump genes in INH monoresistant MTBC clinical isolates.

Study design: In vitro study.

Methods: In our mycobacteriology laboratory, 10 INH monoresistant and 10 primary anti-TB drug-susceptible MTBC clinical isolates were selected. Drug susceptibilities for INH and VR were studied by resazurin microtiter plate method and minimum inhibitory concentration (MIC) was determined. Additionally, mRNA gene expressions were investigated by quantitative Real Time Polymerase Chain Reaction for 21 efflux gene regions.

Results: While no change was observed in INH MICs of susceptible isolates under VR effect, 6 (60%) of the 10 INH-resistant isolates showed a decrease of less than one dilution in INH MIC under VR effect. VR significantly reduced resistance in resistant isolates (p ​< ​0.05). INH monoresistant MTBC isolates showed a 2.85-fold expression increase in the Rv1634 region of the Major Facilitator Superfamily efflux family under INH stress (p ​= ​0.029). No statistically significant change was observed in other efflux gene regions. Herein, increased expression was observed in the Rv1634 region, consistent with other studies in the literature, and this was associated with drug resistance. No significant change in expression was detected in other gene regions.

Conclusion: The effect of efflux pump inhibitor VR on INH MIC levels is promising for the treatment of resistant TB. However, studies with more resistant strains are needed to evaluate the efficacy of efflux pump genes.