Efficacy of Antimicrobials and Their Combinations in Controlling Listeria monocytogenes in Broth and Milk Systems

Introduction: Dairy-related recalls and illnesses are often attributed to Listeria monocytogenes contamination of high-risk cheeses characterized by high water activity and pH levels. Although post-lethality interventions have been identified for more acidic foods, control options are limited for products where heating, controlling pH, and/or controlling water activity is not practical.

Purpose: The objective of this study was to determine the efficacy of GRAS antimicrobials, both singly and in combination, to control L. monocytogenes in a near-neutral broth system and in milk.

Methods: Broth micro-dilution checkerboard assays were utilized to identify minimum inhibitory (MIC) and bactericidal concentrations (MBC) of antimicrobials against L. monocytogenes in growth media (pH ~7.4) incubated at 37°C for 24 h. Fractional inhibitory and bactericidal concentration indices (FIC_I and FBC_I) were calculated to characterize antimicrobial interactions as additive, synergistic, or antagonistic. MICs and MBCs were also determined in UHT milk stored at 7°C for 21 days using a similar dilution assay.  

Results: When applied singly in broth, lauric arginate (LAE), hydrogen peroxide (HP), and e-polylysine (EPL) were most effective with MICs of 6.25, 40, and 20 ppm, and MBCs of 25, 50, and 60 ppm, respectively. At 200 ppm, EPL inhibited the growth of L. monocytogenes in milk throughout storage, while HP (200 ppm) rapidly reduced pathogen levels to below the detection limit, which remained undetectable for 21 days. Combinations of sodium octanoate (SO) with either EPL (FIC_I 0.406) or acidified calcium sulfate with lactic acid (ACSL) (FIC_I 0.479) worked synergistically to inhibit L. monocytogenes in broth and synergistic bactericidal activity was identified for combinations of LAE/EPL (FBC_I 0.286) and SO/EPL (FBC_I0.426). 

Significance: Together, these results at near neutral pH and within a milk matrix serve as a basis for the identification of antimicrobials and their combinations for use in post-lethality treatments to control L. monocytogenes in high-risk cheese and similar products.