Abstract: Objective To investigate the inhibitory effects of levofloxacin (LEV) combined with cellulase against bacille Calmette-Guerin (BCG) biofilms in vitro. Methods The mature growth cycle of BCG biofilms was determined using the XTT method and crystal violet staining. BCG planktonic bacteria and BCG biofilms were treated with different concentrations of LEV and cellulose alone or jointly, and the changes in biofilm biomass were quantified with crystal violet staining. The mature BCG biofilm was then treated with cellulase alone for 24 h, and after staining with SYTO 9 and Calcofluor White Stain, the number of viable bacteria and the change in cellulose content in the biofilm were observed with confocal laser scanning microscopy. The structural changes of the treated biofilm were observed under scanning electron microscopy. Results The MIC, MBC and MBEC values of LEV determined by broth microdilution method were 4 μg/mL, 8 μg/mL and 1024 μg/mL, respectively. The combined treatment with 1/4 ×MIC LEV and 2.56, 5.12 or 10.24 U/mL cellulase resulted in a significant reduction in biofilm biomass (P<0.001). Cellulase treatments at the concentrations of 10.24, 5.12 and 2.56 U/mL all produced significant dispersion effects on mature BCG biofilms (P<0.001). Conclusion LEV combined with cellulose can effectively eradicate BCG biofilm infections, suggesting the potential of glycoside hydrolase therapy for improving the efficacy of antibiotics against biofilm-associated infections caused by Mycobacterium tuberculosis.

Key words: biofilm inhibition; levofloxacin; cellulase; bacille Calmette-Guerin; antibiotics