Evaluating the Biocontrol Potential of Bacillus subtilis Spores Against Listeria monocytogenes in Tryptic Soy Broth and Hummus

This study evaluated the antimicrobial potential of a Bacillus subtilis spore-based probiotic cocktail to reduce foodborne pathogens in both nutrient-rich laboratory media and a complex food matrix (hummus). Three common foodborne pathogens—Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium—were cultured individually in full-strength, half-strength, and quarter-strength tryptic soy broth (TSB) with or without the probiotic spores (~7 log CFU/mL). Additionally, a commercial hummus formulation was inoculated with L. monocytogenes (~3 log CFU/g) and B. subtilis spores (~7 log CFU/g) and stored at 30 °C to simulate temperature abuse. In TSB, E. coli and Salmonella grew to ~8.2 log CFU/mL in full-strength media, with no significant inhibition by the probiotics. However, L. monocytogenes showed substantial suppression: in nutrient-limited TSB, viable counts dropped below the detection limit of 1.48 log CFU/mL by 24 h in the presence of probiotics. In hummus, L. monocytogenes grew to an average of 8.22 log CFU/g in the absence of probiotics but remained significantly lower at an average of 5.03 log CFU/g when co-inoculated with B. subtilis (p < 0.05). Germination of probiotic spores was confirmed within 6 h under all conditions. These findings suggest that B. subtilis spores selectively inhibit Listeria, particularly under nutrient stress or abuse conditions. While the probiotic had limited impact on Gram-negative pathogens, its application may serve as a clean-label strategy for suppressing L. monocytogenes in ready-to-eat (RTE) foods. This dual-model approach provides insights into both mechanistic activity and practical limitations of spore probiotics in complex food matrices.