Purpose: Determine the effectiveness of Listeria phage endolysin PlyP100 against L. monocytogenes when incorporated into Queso Fresco.
Methods: His-tagged PlyP100 was overexpressed in Escherichia coli and subsequently purified. PlyP100 was added to miniature Queso Fresco achieving 10 U/g. Antilisterial activity of PlyP100 was tested by inoculating cheese curds with approximately 5 log CFU/g of L. monocytogenes cocktail, and survival of pathogen was measured during 28 days of storage at 4°C. Different inoculum sizes of L. monocytogenes cocktail were evaluated to determine the pathogen population killed by PlyP100 in cheese after 7 day of storage. The stability of PlyP100 in cheese was assessed by inoculating approximately 5 log CFU/g of L. monocytogenes cocktail on cheese surfaces at 0 – 28 days after manufacture, and pathogen survival was measured at 7 days after cheese inoculation.
Results: PlyP100 in Queso Fresco reduced viable counts of L. monocytogenes by approximately 0.5 log CFU/g and remained without regrowth during 28 day storage. Furthermore, PlyP100 was able to achieve no L. monocytogenes recovery after cheese enrichment when inoculation size was 1 log CFU/g. Finally, an approximate 0.5 log reduction of L. monocytogenes occurred regardless of whether the culture was added immediately upon manufacture or up to 14 days later, suggesting that PlyP100 remains active in cheese during refrigerated storage.
Significance: Our results support the efficacy of phage endolysins as a practical biocontrol measure against Listeria contaminants in Queso Fresco.