Objectives: The primary objective of this study was to determine the minimum inhibitory concentrations (MIC) of USDA-FSIS approved food antimicrobials necessary for the inhibition of the foodborne pathogen Listeria monocytogenes. The second major purpose of this work was to determine the interactions of food antimicrobials with one another for L. monocytogenes inhibition.

Study Methods: These objectives were completed by the use of a broth dilution microassay. Antimicrobials were serially diluted in sterile water immediately prior to sample inoculation. A single strain of the pathogen was incubated at 35C for 24 hours in a non-selective microbiological medium to which the test antimicrobial was added. Following incubation, experimental samples were removed and the change in sample optical density (turbidity) at 630 nm (OD630) was determined. If experimental samples exhibited no change in OD630 (<0.05 OD630) following incubation in the presence of the antimicrobial, the pathogen was pronounced inhibited. With respect to testing of combined antimicrobials, single strains of L. monocytogenes were again incubated at 35C in a nutritious medium containing two antimicrobials. The fractional inhibitory concentrations (FICs) were calculated following determination of MICs from combinations that contained the lowest concentrations of antimicrobials A and B that resulted in a <0.05 OD630. Calculated FIC values were then used to prepare FIC index values (FICI), used to classify the type of interaction observed between antimicrobials in sample wells. The FICIs were used to classify the interaction of antimicrobials as synergistic, additive, or antagonistic.
Study Results: With respect to antimicrobial interactions, the combination of Nisin and Acidic Calcium Sulfate as well as Octanoic Acid and Acidic Calcium Sulfate produced synergistic-type inhibition of L. monocytogenes strains within experimental parameters. Additive-type inhibition of L. monocytogenes was observed with the application of Nisin and Lauramide Arginine Ethyl Ester. Conversely, pairing of Nisin with Octanoic Acid resulted in an increase in the minimum concentrations of Nisin required for inhibition as compared to the MIC of Nisin required for pathogen inhibition when applied singly. Similar trends were observed for other antimicrobial pairings, whereby the minimum concentration of one or both antimicrobials capable of inhibiting the pathogen in combination was more than what was needed for L. monocytogenes inhibition when one or both antimicrobials were applied alone. Furthermore, the resulting antagonistic-type interactions of many antimicrobial pairings indicates that use of dual antimicrobials does not necessarily guarantee the user significantly enhanced inhibition of foodborne bacterial pathogens over the use of a single compound.

Industry Significance: Results of this study indicate that some experimental combinations of food antimicrobials produced synergistic-type inhibition of L. monocytogenes when incubated at 35C at pH 7.3 or 5.0, whereas others produced additive-type and antagonistic type behavior. However, it is recommended that before processors use a food antimicrobial or a combination of antimicrobials they must first experimentally validate the actions and interactions of antimicrobials in vitro and on surfaces of the food product and their ability to produce enhanced inhibitory effects against the target microorganism.
Principal Investigator Thomas M. Taylor, Ph.D., Department of Animal Science, Texas AgriLife Research, may be contacted at 979-862-7678 (Phone), 979-862-3475 (Fax), or [email protected]. Contact via U.S. mail at: 310-C Kleberg Center, College Station, TX 77843-2471.