The microbiota of wooden cheese ripening boards is a rich source of antimicrobial-producing bacteria against Listeria monocytogenes

Abstract

Wooden boards are essential tools in cheese ripening and there are accumulating observations suggesting their antimicrobial effect against foodborne bacterial pathogens, such as Listeria monocytogenes. However, poor bacterial recovery of bacteria from wood can confound quantification of pathogen burdens. To assess L. monocytogenes survival on wooden cheese boards, we applied a disruptive grinding method and tracked native board-associated bacterial counts as controls. Our data revealed that L. monocytogenes declines on clean zones of wooden boards, but can replicate on areas where there is suitable cheese. Our microbiota analysis revealed diverse bacterial communities on wooden board surfaces, with a prominent presence of Brevibacterium, Brachybacterium, and Staphylococcus genera. We further identified seven bacterial species that inhibit L. monocytogenes, belonging to Bacillus, Staphylococcus, and Serratia phyla, as well as Lactococcus lactis. We focused on a Bacillus safensis isolate as a novel biocontrol agent candidate, and found it to potently inhibit L. monocytogenes via secreted antimicrobial factors. Our genomic, bioinformatic, and biochemical analyses indicate that those factors are likely antimicrobial peptides encoded by multiple biosynthetic gene clusters, several of which are unique to B. safensis and have not been characterized. A sub-inhibitory concentration of B. safensis supernatant induces a significant down-regulation of prophage elements and up-regulation antimicrobial stress response in L. monocytogenes. Taken together, our findings indicate that the wooden board microbiota is a rich source of antimicrobial-producing bacteria with potential applications in foodborne pathogen control strategies. IMPORTANCEDespite stringent food safety measures, L. monocytogenes foodborne outbreaks remain frequent with high hospitalization and mortality rates. Removal of L. monocytogenes from food processing environments is extremely challenging, because this pathogen is ubiquitous and encodes a wide array of stress response mechanisms that enable it to thrive under harsh conditions. Our study found that clean wooden boards used in cheese ripening inhibit L. monocytogenes, causing a noticeable decline in pathogen population following surface inoculation. Bacterial communities on wooden cheese boards are rich and diverse, and harbor many species that produce antimicrobial compounds against L. monocytogenes, with the example of a new B. safensis isolate. Therefore, wooden board microbiota is a promising source for future antimicrobial discovery efforts.