Efficacies of Nisin A and Nisin V Semipurified Preparations Alone and in Combination with Plant Essential Oils for Controlling Listeria monocytogenes

Abstract

The food-borne pathogenic bacterium Listeria is known for relatively low morbidity and high mortality rates, reaching up to 25 to 30%. Listeria is a hardy organism, and its control in foods represents a significant challenge. Many naturally occurring compounds, including the bacteriocin nisin and a number of plant essential oils, have been widely studied and are reported to be effective as antimicrobial agents against spoilage and pathogenic microorganisms. The aim of this study was to investigate the ability of semipurified preparations (SPP) containing either nisin A or an enhanced bioengineered derivative, nisin V, alone and in combination with low concentrations of the essential oils thymol, carvacrol, and trans-cinnamaldehyde, to control Listeria monocytogenes in both laboratory media and model food systems. Combinations of nisin V-containing SPP (25 g/ml) with thymol (0.02%), carvacrol (0.02%), or cinnamaldehyde (0.02%) produced a significantly longer lag phase than any of the essential oil-nisin A combinations. In addition, the log reduction in cell counts achieved by the nisin V-carvacrol or nisin V-cinnamaldehyde combinations was twice that of the equivalent nisin A-essential oil treatment. Significantly, this enhanced activity was validated in model food systems against L. monocytogenes strains of food origin. We conclude that the fermentate form of nisin V in combination with carvacrol and cinnamaldehyde offers significant advantages as a novel, natural, and effective means to enhance food safety by inhibiting food-borne pathogens such as L. monocytogenes. The growing consumer demand for food products that are minimally processed and free of chemical preservatives presents a difficult challenge for food processors. Consequently, there has been a focus on the application of naturally produced antimicrobial compounds as a more acceptable means to control the growth of undesirable microorganisms in food (1, 2). Bacteriocins (ribosomally produced, small, heat-stable peptides that are active against other bacteria) derived from organisms generally regarded as safe provide one potential solution. However, only two bacteriocins have been made commercially available to any extent. These are nisin, produced by Lactococcus lactis, and pediocin PA-1, produced by Pediococcus acidilactici (3, 4). Of these, nisin is used in a wide variety of dairy and nondairy products, including cream and cheese products, soups, liquid egg, mayonnaises, salad dressings, tomato products, and beer (5). Nisin A exhibits antibacterial activity against a wide range of Gram-positive bacteria, including food-borne pathogens such as staphylococci, bacilli, clostridia, and Listeria (6, 7). Indeed, the success of nisin A from discovery (8) through to regulatory approval and finally to commercial application has spurred researchers to exploit its geneencoded nature and to attempt to "bioengineer" variants with altered biological, chemical, and physical properties. Over the last decade, several studies have described the discovery of new nisi...

A Bioengineered Nisin Derivative, M21A, in Combination with Food Grade Additives Eradicates Biofilms of Listeria monocytogenes

“…This is approximately 50% greater than the MIC previously published for cinnamaldehyde against a number of different L. monocytogenes strains. The MIC for carvacrol was, however, in keeping with results targeting two strains from this same study (Field et al, 2015a). Following initial broth-based growth curves the most notable effects were observed however when citric acid or cinnamaldehyde were used in combination with nisin.…”

“…Of the variant strains tested the observed zone sizes for three derivative producers, AAA (where the three hinge amino acids in positions 20–22 have been replaced with alanines), M21A and M21V were consistently enhanced when compared to the corresponding isogenic nisin A producer ( Figure 1 ). To date a number of studies have been conducted that have highlighted the efficacy of M21V against strains of L. monocytogenes (Field et al, 2008, 2015a; Campion et al, 2013). Here, we noted that the antimicrobial efficiency of the M21A producer appeared equivalent to that of M21V, while they both had a greater degree of activity against the indicator strain than that of the producer of the AAA derivative.…”

“…By including bioengineered versions of nisin in this study we sought to and achieved increased antimicrobial potency. Due to the documented sensitivity of Listeria to nisin and the relevance of both to the food industry (Thomas and Wimpenny, 1996; Arqués et al, 2008; Pinto et al, 2010; Field et al, 2015a), strain F6854, which has been linked to contaminated turkey frankfurters, represented an ideal target for nisin and food-grade antimicrobial combinations. The food additives sodium benzoate, sodium acetate, potassium sorbate, lactoferrin, carvacrol, citric acid and cinnamaldehyde were all tested initially.…”

“…Biofilm growth was performed as previously described by Field et al (2015a), but with modifications as follows. TSB supplemented with 1% D -(+)-glucose (Sigma) (TSBg) was used in order to aid biofilm formation (Choi et al, 2013).…”

“…Essential oils are natural, volatile liquids obtained from plant material (Gálvez et al, 2007) that have aromatic qualities (Burt, 2004). A number of studies have also been carried out that demonstrate the enhanced antimicrobial activity of nisin in combination with essential oils such as cinnamaldehyde (Field et al, 2015a) and carvacrol (Pol and Smid, 1999) against a broad range of pathogens including Gram-positive organisms, such as Listeria monocytogenes and Bacillus cereus (Pol and Smid, 1999; Field et al, 2015a), and Gram-negative bacteria, including Salmonella enterica and Escherichia coli (Phillips and Duggan, 2001; Govaris et al, 2010; Norhana et al, 2012; Prudêncio et al, 2015). …”