Hispanic-style cheeses (HSC) are a varied group of cheeses that are originally developed and manufactured in Mexico and other Latin American countries. In particular, Queso Fresco (QF) is the most widespread HSC in the United States. Most of the HSC are characterized by being fresh, having a delicate flavor and texture, and a short-shelf life that requires to store them under refrigeration. Different from aged cheeses, fresh cheeses provide conditions for the growth of foodborne pathogens, notably Listeria monocytogenes.HSCs have been linked to historical and recent listeriosis outbreaks. Since 2008, about one out of every five listeriosis outbreaks in the U.S. have been associated with the consumption of HSC, this is higher than for other ready-to-eat foods and other types of cheeses, highlighting the health risk associated between HSCs and L. monocytogenes.Similar to other types of cheeses, HSC can be manufactured safely; however, pasteurization is the only step in the process that eliminates L. monocytogenes, and post-pasteurization interventions are needed to ensure their safety if contamination occurs. To date, multiple potential interventions to inhibit L. monocytogenes in QF have been evaluated, including non-thermal processes such as High Pressure Processing, or the addition of ingredients with antilisterial activity such as organic acids, bacteriocins, fermentates, bacteriophages or antimicrobial packaging. Unfortunately, those treatments have not been effective to eliminate L. monocytogenes in QF, and it is necessary to find antimicrobial treatments that can work in QF. Nisin is an antimicrobial peptide produced by certain strains of Lactococcus lactis, it is widely used in the food industry and it is able to inhibit L. monocytogenes, however it has limited activity in QF. Therefore, the overall goal of this study is to enhance the antilisterial effectiveness of nisin in QF by exploring its synergy with novel antimicrobials, and the use of microencapsulation technology and bioengineering approaches.In Chapter 2, the potential synergy of nisin with the Listeria phage endolysin PlyP100 against Listeria in QF was evaluated. By combining the commercial preparation of nisin with the purified endolysin PlyP100 in QF, the treatment reduced the viable populations of the pathogen to below the detection limit of enumeration, and in some samples, recovery of the pathogen was not possible, indicating elimination of the pathogen in the cheese. The combination of nisin with PlyP100 could be used as a preservative in QF due to the listeriacidal synergy observed both antimicrobials.In Chapter 3, a droplet-based microfluidic process was used to microencapsulate nisin with zein for antilisterial activity enhancement. Microencapsulation of nisin with zein allowed for sustained release of nisin in solution, and when added into QF, microencapsulated nisin displayed significantly enhanced listeriacidal effect over the first 3 days of storage at 4 ⁰C. Fabrication of nisin-zein microcapsules could be further optimized for tuning the release rate and microcapsule properties for extended antilisterial effect of nisin in cheese.In Chapter 4, a series of nisin A derivatives in which the hydrophobic residues of the C-terminal region were replaced with positively charged amino acids was created. All nisin derivatives created displayed altered antilisterial and biochemical properties relative to unmodified nisin, notably reduced antilisterial activity but significantly higher stability at neutral pH and lower absorption to milk fat. Single substituted nisin derivatives were stable enough in QF to exhibit a listeriastic effect at early cold storage of the cheese despite of their reduced antilisterial activity. Bioengineering of nisin can result in the creation of nisin derivatives with higher stability and extended antimicrobial activity in non-acidic and high-fat food matrices.
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Improving the efficacy of nisin to control Listeria monocytogenes in queso fresco
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