Efficacy of bacteriocin-containing cell-free culture supernatants from lactic acid bacteria to control Listeria monocytogenes in food

Abstract

Consumer demands have led to an increased interest in the use of natural antimicrobials for food protection. With the objective of developing novel products for enhancing the microbial safety of food, we have tested cell-free culture supernatants (CFS's) of eight antagonistic bacterial strains for their efficacy to inhibit Listeria monocytogenes in different food matrices. The antagonistic strains represented different members of the order Lactobacillales as well as one isolate of Staphylococcus sciuri and all showed strong inhibition of L. monocytogenes on agar plates. Cell-free supernatants were obtained after growing the bacteria in a yeast extract-glucose broth. In six of the CFS's, different class IIa bacteriocins, namely leucocin A, leucocin B, mundticin L, pediocin PA-1, sakacin A, and sakacin X, were identified as the major anti-listerial compounds. For the other two strains, the active substances could not be ascertained conclusively. The minimal effective concentration (MEC) of the individual CFS's to achieve a 2.3 log(10) reduction of L. monocytogenes was determined in culture broth, whole milk, and ground beef at 4°C. While all bacteriocin-containing CFS's were effective in broth at concentrations from 52 to 205 AU/ml, significant higher concentrations were needed when applied in food. Best results were obtained using CFS's containing pediocin PA-1, that displayed only three- and ten-times higher MEC's in milk (307 AU/ml) and ground meat (1024 AU/g) compared to broth, respectively. A twenty-fold increase in the MEC (2048 AU/ml) was observed for a mundticin L-containing fermentate, and a CFS containing leucocin A and B was inactivated more than fifty-fold (>1280 AU/ml) in both food matrices. Remarkably, the sakacin A and sakacin X containing CFS's displayed very selective inactivation rates, in which sakacin A was only effective in meat (512 AU/g), while sakacin X was only effective in milk (2048 AU/ml). In all cases, inhibition of L. monocytogenes was only transient and surviving or resistant bacteria started growing after prolonged storage. These results highlight the importance of careful testing the effectiveness of bacteriocins in the food systems for which they are intended to be applied against the selected target and non-target bacteria. Furthermore, the outgrowth of surviving or resistant bacterial populations points out that the tested bacteriocins are not suited to assure full inhibition of L. monocytogenes in a food product, if not applied in combination with additional preservative measures.