**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** LISTERIA MONOCYTOGENES (LM) IS A HUMAN PATHOGEN AND WORLD-WIDE, THE LEADING CAUSE OF BACTERIAL LINKED FOODBORNE MORTALITY. OUTBREAKS OF LISTERIOSIS AFFECT PRIMARILY AT- RISK PEOPLE (PREGNANT WOMEN, ELDERLY, IMMUNOCOMPROMISED INDIVIDUALS) AND CAN BE TRACED TO CONSUMPTION OF SMOKED MEATS, SEAFOOD, AND DAIRY PRODUCTS AS WELL AS PREHARVEST FRESH PRODUCE (CABBAGE, CORN, LETTUCE, CELERY, RADISH, AND SPINACH) AND THE PRESENCE OF LM IN THE FARM ENVIRONMENT. ANTIBIOTICS HAVE BEEN USED TO INHIBIT THE GROWTH OF PATHOGENIC BACTERIA FOR MANY DECADES, REDUCING THEIR THREAT TO HUMAN HEALTH. HOWEVER, DUE TO THE INCREASING NUMBER OF ANTIBIOTIC RESISTANT AND MULTI-DRUG RESISTANT STRAINS OF BACTERIA IT HAS BECOME NECESSARY TO EXPLORE NEW ALTERNATIVES FOR ERADICATING PATHOGENS. BACTERIOPHAGE (PHAGE) ENDOLYSINS ARE ENZYME ANTIMICROBIALS THAT DIGEST THE MAJOR BUILDING BLOCKS OF THE BACTERIAL CELL WALL. IN GRAM-POSITIVE BACTERIA INCLUDING LM, THE CELL WALL IS SURFACE EXPOSED, SO THE PURIFIED ENDOLYSIN WHEN EXPOSED EXTERNALLY, CAN CAUSE DEATH OF THE PATHOGEN BY DEGRADING THE CELL WALL. ENDOLYSINS ARE HIGHLY SPECIES-SPECIFIC AND REFRACTORY TO RESISTANT STRAIN DEVELOPMENT. DESPITE NUMEROUS EFFORTS TO FIND THEM, NOT A SINGLE ENDOLYSIN-RESISTANT BACTERIAL STRAIN HAS BEEN IDENTIFIED SO FAR. WE BELIEVE THAT ENDOLYSINS CAN PREVENT OR ERADICATE LM ON LEAFY VEGETABLES. THIS PROPOSAL WILL EXAMINE THE KNOWN LM PHAGE ENDOLYSINS FOR THEIR EFFICACY AGAINST LM STRAINS FOUND ON SPINACH. HOWEVER, ENDOLYSIN PURIFICATION IS NOT COST-EFFECTIVE FOR TREATING LARGE QUANTITIES OF LEAFY VEGETABLES WITH A PURIFIED LM PHAGE ENDOLYSIN, AND THIS TREATMENT ALONE WOULD NOT CONTROL OTHER FOOD-BORNE BACTERIAL PATHOGENS SUCH AS SALMONELLA OR E. COLI. TO ADDRESS THIS CONCERN, CURRENT TREATMENTS TO ERADICATE FOOD BORNE PATHOGENS INCLUDE CHEMICAL SANITIZERS. HOWEVER, FOOD BORNE PATHOGENS HAVE BEEN REPORTED TO DEVELOP RESISTANCE TO CHEMICAL SANITIZERS, INDICATING THAT LM MIGHT ALSO EVENTUALLY DEVELOP RESISTANCE TO THESE CHEMICAL AGENTS. THUS, IT IS THE GOAL OF THIS PROJECT TO IDENTIFY INDIVIDUAL OR PAIRED LM ENDOLYSINS WITH THE HIGHEST EFFICACY AGAINST SPINACH BORN LM AND TO THEN TEST FOR INCREASED KILLING EFFICACY BY PAIRING THE ENDOLYSIN TREATMENT WITH EXISTING FOOD INDUSTRY CHEMICAL TREATMENTS. THIS SHOULD IDENTIFY A MORE SUSTAINABLE TREATMENT OF FOOD BORNE PATHOGENS THAT DOES NOT INCUR SIGNIFICANTLY INCREASED TREATMENT COSTS WHILE REDUCING THE RISK OF CREATING TREATMENT-RESISTANT STRAINS OF LM.