Inhibitors of biofilm formation by fuel ethanol contaminants

Industrial fuel ethanol production suffers from chronic and acute infections that reduce yields and cause “stuck fermentations” that result in costly shutdowns.  Lactic acid bacteria, particularly Lactobacillus sp., are recognized as major contaminants.  In previous studies, we observed that certain species persist over time, suggesting the presence of recalcitrant biofilms.  Subsequently, we demonstrated that some contaminants form biofilms under laboratory conditions.  In the current study, biofilm-forming strains of Lactobacillus brevis, L. fermentum, and L. plantarum were isolated from a dry-grind fuel ethanol plant and tested against a variety of potential inhibitors.  Microbial polysaccharides (alternan, pullulan, or schizophyllan) did not inhibit biofilm formation.  Neither did ferric ammonium citrate, liamocins, or xylitol. The antibiotic virginiamycin inhibited both growth and biofilm formation.  Commercial enzyme preparations (cellulase, β-glucosidase, and xylanase) generally reduced biofilm formation.  Crude culture supernatants from Bacillus cereus strain MR1 had little or no effect, but those from B. subtilis strains ALT3A and RPT-82412 reduced biofilm formation.  Supernatants from strain RPT-82412 also inhibited planktonic growth, while those from strain ALT3A did not.  MALDI-TOF mass spectra suggested the presence of surfactin-like molecules in culture supernatants of both B. subtilis strains, and the presence of iturin-like molecules in supernatants of strain RPT-82412.  Surfactins and iturins are surface-active lipopeptides that can have antimicrobial activities, and biosurfactants have been reported to inhibit the adhesion of certain biofilm-forming bacteria.  B. subtilis strains merit further examination as novel control agents for fuel ethanol contamination problems.