Nisin高产菌株的高通量筛选

【目的】乳酸链球菌素(nisin)是一种天然生物活性抗菌肽,对包括食品腐败菌和致病菌在内的许多革兰氏阳性菌具有强烈的抑制作用,而用作食品的防腐剂。本研究通过建立高通量筛选方法,实现高效快速省力的高产菌株筛选,为工业上筛选高产菌株提供研究方案。【方法】通过对Lactococcus lactis ATCC11454菌株进行紫外诱变,获得2511株突变株。利用Biomek FXP自动工作站建立96微孔板的高通量筛选方法,突变株经高通量挑选、菌种培养及菌液稀释后,加入到生长至对数中期的藤黄微球菌中,采用改进后的比浊法快速检测nisin生物活性。用此方法对突变株进行初筛、复筛后可得到nisin高产菌株,并通过摇瓶发酵评估高通量筛选方法。【结果】确定比浊法检测的条件为:nisin活性稀释在10–25 IU/m L范围内,与藤黄微球菌反应2 h后检测藤黄微球菌的菌体量(OD600)。2511株突变株经过2轮高通量筛选,最终获得约50株产量提升的菌株,对其中8株进行摇瓶精确测量,显示产量均有提高,并且其中一株产量提升了30%,成功建立了高通量筛选nisin高产菌株的方法。【结论】利用比浊检测法,在其基础上成功建立高通量筛选高产nisin菌的方法,经过初筛复筛,整个周期由1人耗时5 d即可完成2511株突变株的筛选工作。相较于传统的选育方法,高通量筛选具有快速、稳定、高效的特点,提高了筛选效率,缩短了选育周期,是工业上筛选高产nisin菌的有效手段。     

Identification and Characteristics of Nisin Z-Producing <Emphasis Type="Italic">Lactococcus lactis</Emphasis> subsp. <Emphasis Type="Italic">lactis</Emphasis> Isolated from Kimchi

We isolated bacteriocin-producing Lactococcus lactis subsp. lactis from Kimchi. The bacteriocin inhibited strains of Clostridium perfringens, C. difficile, Listeria monocytogenes, vancomycin-resistant Enterococcus, and one out of four methicillin-resistant Staphylococcus aureus strains, as well as some closely related lactic acid bacteria. In tricine-SDS-PAGE, the bacteriocin migrated with an apparent molecular weight of about 4 kDa to the same location as nisin A and crude nisin Z. The gene encoding this bacteriocin was found to be identical to that of nisin Z with direct PCR sequence methods. The inhibitory activity was stable against heat and pH, but it was lost at 100°C for 1 h and at 121°C for 15 min. The bacteriocin was inactivated by proteolytic enzymes, but was not affected by lysozyme, lipase, catalase, or β-glucosidase. There were some differences in characteristics from those of nisins described previously. Received: 21 June 2002 / Accepted: 22 July 2002     

Chemical Modification of the Carboxyl Terminal of Nisin A with Biotin does not Abolish Antimicrobial Activity Against the Indicator Organism, Kocuria rhizophila

Nisin is an antimicrobial peptide that is widely used for food preservation. Although it has potent activity against a number of food pathogens, suggesting potential therapeutic applications, its potential for clinical use is limited by proteolytic susceptibility and poor oral bioavailability. Derivatization of nisin could overcome these issues; however, many nisin analogues, prepared by modification at the N-terminal and C-terminal have previously been shown to be inactive. A method for the C-terminal modification was developed using biotinylation as a model derivative. Purification of the modified nisin was carried out using reverse phase chromatography. Confirmation of nisin modification was confirmed by Mass Spectroscopy. The C-terminal modification of nisin resulted in only a twofold reduction in antimicrobial activity of the conjugate against the indicator organism, Kocuria rhizophila. The C-terminal modification could be used to increase the therapeutic potential of nisin by creating more favourable physicochemical characteristics. This is the first study that showed that nisin modification can be carried out successfully without destroying its antimicrobial activity.     

Induction of autolysis of staphylococci by the basic peptide antibiotics Pep 5 and nisin and their influence on the activity of autolytic enzymes

Pep 5 and nisin are cationic bactericidal peptides which were shown to induce autolysis in Staphylococcus cohnii 22. In contrast to nisin, Pep 5 induced lysis could be stimulated in the presence of glucose. Addition of lipoteichoic acids (LTA) (d-alanine:phosphorus=0.475:1) inhibited all effects of Pep 5 on susceptible cells in a molar ratio LTA:Pep 5 of 10:1. Treatment of S. cohnii 22 with Pep 5 or nisin for 20 min and subsequent washing with 2.5 M NaCl released autolysin activity. Crude preparations of the hydrolyzing enzymes produced free amino groups as well as polysaccharide fragments from the murein backbone, suggesting the presence of a muramidase or glucosamidase, and endopeptidase or amidase. Both enzyme activities were inhibited by lipoteichoic acid; they could be fully reactivated by addition of Pep 5 in sufficient concentrations. The velocity of hydrolysis was not influenced by nisin, whereas it was doubled in presence of Pep 5. The results are discussed in view of a possible mechanism of induction of lysis by Pep 5 and nisin.Abbreviations A.U. arbitrary unit - CCCP carbonylcyanide-m-chlorophenyl hydrazone - DNase deoxyribonuclease - CYG casein yeast extract glucose - IT initial turbidity - LTA lipoteichoic acid - RNase ribonuclease - TSB Tryptone Soy Broth     

Voltage-dependent depolarization of bacterial membranes and artificial lipid bilayers by the peptide antibiotic nisin

The peptide antibiotic nisin is shown to disrupt valinomycin-induced potassium diffusion potentials imposed on intact cells of Staphylococcus cohnii 22. Membrane depolarization occurred rapidly at high diffusion potentials while at low potentials nisin-induced depolarization was slower suggesting that nisin requires a membrane potential for activity. This assumption was proven in experiments with planar lipid bilayers (black lipid membranes). Macroscopic conductivity measurements indicated a voltage-dependent action of nisin. The potential must have a trans-negative orientation with respect to the addition of nisin (added to the cis-side) and a sufficient magnitude (ca. -100 mV). With intact cells the threshold potential was lower (-50 to -80 mV at pH 7.5 and below -50 mV at pH 5.5). Single channel recordings resolved transient multistate pores, strongly resembling those introduced by melittin into artificial bilayers. The pores had diameters in the range of 0.2–1 nm, and lifetimes of few to several hundred milliseconds. The results indicate that nisin has to be regarded as a membrane-depolarizing agent which acts in a voltage-dependent fashion.Abbreviations BLM Black lipid membranes - CCCP carbonyl cyanide m-chlorophenylhydrazone - DOPC dioleoyl phosphatidylcholine - PS phosphatidylserine - TPP⁺ tetraphenylphosphonium cation     

Biosynthetic characterization and biochemical features of the third natural nisin variant,nisin Q,produced by Lactococcus lactis 61‐14

Aims: To characterize the genetic and biochemical features of nisin Q. Methods and Results: The nisin Q gene cluster was sequenced, and 11 putative orfs having 82% homology with the nisin A biosynthesis gene cluster were identified. Nisin Q production was confirmed from the nisQ‐introduced nisin Z producer. In the reporter assay, nisin Q exhibited an induction level that was threefold lower than that of nisin A. Nisin Q demonstrated an antimicrobial spectrum similar to those of the other nisins. Under oxidizing conditions, nisin Q retained a higher level of activity than nisin A. This higher oxidative tolerance could be attributed to the presence of only one methionine residue in nisin Q, in contrast to other nisins that contain two. Conclusions: The 11 orfs of the nisin producers were identical with regard to their functions. The antimicrobial spectra of the three natural nisins were similar. Nisin Q demonstrated higher oxidative tolerance than nisin A. Significance and Impact of the Study:  Genetic and biochemical features of nisin Q are similar to those of other variants. Moreover, owing to its higher oxidative tolerance, nisin Q is a potential alternative for nisin A.     

Scalable purification of the lantibiotic nisin and isolation of chemical/enzymatic cleavage fragments suitable for semi‐synthesis

Herein, we describe a scalable purification of the lantibiotic nisin via an extraction/precipitation approach using a biphasic system, which can be carried out up to 40–80 gram scale. This approach results in an at least tenfold enrichment of commercially available preparations of nisin, which usually contain only 2.5% of the desired peptide, to allow further purification by preparative HPLC. As a follow‐up study, the enriched nisin sample was digested either by trypsin or chymotrypsin, or treated by CNBr, and these reactions were monitored by LC‐MS to identify and characterize the obtained fragments. Two previously unknown cleavage sites have been identified: Asn20–Met21 and Met21–Lys22 for trypsin and chymotrypsin, respectively. Furthermore, a novel and convenient enzymatic approach to isolate the native nisin C‐ring [nisin fragment (13–20)] was uncovered. Finally, by means of preparative HPLC, nisin fragments (1–12), (1–20), (22–34), and (22–31) could be isolated and will be used in a semi‐synthesis approach to elucidate the role of each fragment in the mode of action of nisin as an antimicrobial peptide. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

The Effect of Calcium and Magnesium on the Activity of Bovicin HC5 and Nisin

Some Gram-positive bacteria produce small peptides (bacteriocins) that have antimicrobial activity, but many bacteria can become bacteriocin resistant. Bovicin HC5, a lantibiotic produced by Streptococcus bovis HC5, has the ability to inhibit nisin-resistant bacteria. Because nisin resistance has in many cases been correlated with an alteration of lipoteichoic acids or the polar head groups of membrane phospholipids, we decided to examine the effect of divalent cations on nisin and bovicin HC5 activity. Both bacteriocins catalyzed potassium efflux from S. bovis JB1, a non–bacteriocin-producing strain. The addition of large amounts (100 mM) of calcium or magnesium increased the ability of S. bovis JB1 to bind Congo red (an anionic dye) and counteracted bacteriocin-mediated potassium loss. Calcium was more effective than magnesium in decreasing nisin activity, but the reverse was observed with bovicin HC5. Nisin-resistant S. bovis JB1 cells bound three times as much Congo red as nisin-sensitive cells, and this result is consistent with the idea that changes in cell surface charge can be a mechanism of bacteriocin resistance. The nisin-resistant cells were less susceptible to bovicin HC5, but bovicin HC5 still caused a 50% depletion of intracellular potassium. These results indicate that nisin and bovicin HC5 react differently with the cell surfaces of Gram-positive bacteria. Proprietary or names are necessary to report factually on available data; however, the United States Department of Agriculture (USDA) neither guarantees nor warrants the standard of the product, and the use of the name by the USDA implies no approval of the product, and exclusion of others that may be suitable.     

Optimisation of a rapid bioassay for nisin utilising potassium efflux from sensitivePediococcus sp. cells

Summary Addition of nisin to a K⁺-loaded, metabolising suspension ofPediococcus sp resulted in K⁺ efflux into the medium. The total K⁺ efflux was proportional to the logarithm of the nisin activity with a MEC of 0.7 IUml⁻¹ and resulted in total loss of cellular K⁺ at 4–5 IUml nisin. Conditions were optimised for the use of K⁺ efflux measurements as a rapid nisin bioassay using a combination of factorial and sequential simplex procedures. A nisin assay based on K⁺ efflux measurements was used to follow the course of a nisin-producing fermentation.     

Salt effect of nisin Z isolated from a marine fish on the growth inhibition of Streptococcus iniae, a pathogen of streptococcosis

A bacteriocin-producing Lactococcus lactis subsp. lactis was isolated from the intestine of olive flounder. The bacteriocin was identified as nisin Z. It was active against Gram-positive bacteria. Nisin Z at 3,200 arbitrary units (AU) was more effective in seawater than in PBS; growth of Streptococcus iniae was completely inhibited within 3 h. Nisin Z preparations with 3.5% (w/v) NaCl was the most effective against S. iniae being similar to nisin Z in seawater. Nisin Z is thus a good alternative to antibiotics to prevent streptococcosis caused by S. iniae aquaculture systems.     

Incorporation of nisin into a meat binding system to inhibit bacteria on beef surfaces

In two separate experiments, the bacteriocin, nisin, was incorporated into a commercially available meat binding system (Fibrimex^®) and applied to meat surfaces as a way of inhibiting the meat spoilage organism, Brochothrix thermosphacta during extended refrigerated storage. In experiment 1, pre-rigor lean beef carcass tissue (BCT) was inoculated with B. thermosphacta , left untreated (U), treated with 10 μg ml⁻¹ nisin (N), Fibrimex^® (F) or Fibrimex^® containing 10 μg ml⁻¹ nisin (FN), held aerobically at 4 °C for up to 7 d, and populations of B. thermosphacta and nisin activity determined. Experiment 2 determined the effects of the same treatments but on post-rigor, frozen and thawed lean BCT that was inoculated, vacuum-packaged, and stored at 4 °C for up to 14 d. In both experiments, N- and FN-treated tissues exhibited significantly lower populations of B. thermosphacta compared to U- and F-treated tissues, for the duration of refrigerated storage. Nisin activity was detected up to 7 d in N- and FN-treated samples from experiment 1. However, activity was detected only to days 0 and 2 in FN- and N-treated samples, respectively, from experiment 2. These studies indicate that the addition of a bacteriocin to a meat binding system and application to meat surfaces may be useful in reducing undesirable bacteria in restructured meat products.     

Potential application of the nisin Z preparation of Lactococcus lactis W8 in preservation of milk

Aims: The aim of the study is to evaluate the effectiveness of the preparation of nisin Z from Lactococcus lactis W8‐fermented milk in controlling the growth of spoilage bacteria in pasteurized milk. Methods and Results: Spoilage bacteria isolated from pasteurized milk at 8 and 15°C were identified as Enterococcus italicus, Enterococcus mundtii, Enterococcus faecalis, Bacillus thuringiensis, Bacillus cereus, Lactobacillus paracasei, Acinetobacter sp., Pseudomonas fluorescens and Enterobacter aerogenes. These bacteria were found to have the ability to survive pasteurization temperature. Except Enterobacter aerogenes, the spoilage bacteria were sensitive to the nisin Z preparation of the L. lactis W8. Addition of the nisin Z preparation to either the skim milk or fat milk inoculated with each of the spoilage bacteria reduced the initial counts (about 5 log CFU ml^?1) to an undetectable level within 8–20 h. The nisin Z preparation extended the shelf life of milk to 2 months under refrigeration. Conclusions: The nisin Z preparation from L. lactis W8‐fermented milk was found to be effective as a backup preservative to counteract postpasteurization contamination in milk. Significance and Impact of the Study: A rapid inhibition of spoilage bacteria in pasteurized skim and fat milk with the nisin Z preparation of L. lactis W8 is more significant in comparison with the commercially available nisin (nisin A). The nisin Z preparation can be used instead of commercial nisin, which is not effective in fat milk.     

PCR detection of the structural genes of nisin Z and lacticin 481 in Lactococcus lactis subsp. lactis INIA 415, a strain isolated from raw milk Manchego cheese

A Lactococcus strain with strong antimicrobial activity was isolated from raw milk Manchego cheese during a survey on the production of bacteriocins by lactic acid bacteria present in raw milk cheeses. It was identified as Lactococcus lactis subsp. lactis, phenotypically by its morphological and physiological characteristics and genotypically by a PCR technique. When tested for tolerance to known bacteriocins produced by lactococci, it was shown to be resistant to nisin A and nisin Z. The presence of genes encoding nisin and lacticin 481 was revealed by PCR techniques with specific probes. Sequences of the respective PCR amplified fragments matched sequences reported for nisin Z and lacticin 481.     

Nisin production of Lactococcus lactis N8 with hemin‐stimulated cell respiration in fed‐batch fermentation system

In this study, nisin production of Lactococcus lactis N8 was optimized by independent variables of glucose, hemin and oxygen concentrations in fed‐batch fermentation in which respiration of cells was stimulated with hemin. Response surface model was able to explain the changes of the nisin production of L. lactis N8 in fed‐batch fermentation system with high fidelity (R² 98%) and insignificant lack of fit. Accordingly, the equation developed indicated the optimum parameters for glucose, hemin, and dissolved oxygen were 8 g L^?1 h^?1, 3 μg mL^?1 and 40%, respectively. While 1711 IU mL^?1 nisin was produced by L. lactis N8 in control fed‐batch fermentation, 5410 IU mL^?1 nisin production was achieved within the relevant optimum parameters where the respiration of cell was stimulated with hemin. Accordingly, nisin production was enhanced 3.1 fold in fed‐batch fermentation using hemin. In conclusion the nisin production of L. lactis N8 was enhanced extensively as a result of increasing the biomass by stimulating the cell respiration with adding the hemin in the fed‐batch fermentation. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:678–685, 2015     

Inhibitory activity of natural antimicrobial compounds alone or in combination with nisin against Enterobacter sakazakii

Aim: To determine the antimicrobial activity of natural organic compounds alone and in combination with nisin on the growth of Enterobacter sakazakii in laboratory media. Methods and Results: The minimum inhibitory concentrations (MIC) of five natural organic compounds were determined, and their effects in combination with nisin were evaluated by comparing treatment with each natural organic compound alone and in combination with 25 mg ml^?1 nisin in tryptic soy broth. Among the tested natural organic compounds, the MIC of carvacrol and thymol was 1·25 mmol l^?1 and showed the strongest inhibitory activity against E. sakazakii, whereas the MIC of cinnamic acid was higher than 5 mmol l^?1, and therefore showed the weakest inhibitory activity. However, the combination of each compound with nisin did not result in the enhancement of their antimicrobial activities except when nisin was combined with diacetyl. Conclusions: The order of inhibition attributed to natural organic compounds was carvacrol = thymol > eugenol > diacetyl > cinnamic acid, and only the combination of diacetyl and nisin showed a synergistic effect of inhibiting the growth of E. sakazakii. Significance and Impact of the Study: This study shows the potential of natural organic compounds for controlling E. sakazakii.     

A Comparative Study Between the Antibacterial Effect of Nisin and Nisin-Loaded Chitosan/Alginate Nanoparticles on the Growth of Staphylococcus aureus in Raw and Pasteurized Milk Samples

The aim of this study was to evaluate the antibacterial effect of nisin-loaded chitosan/alginate nanoparticles as a novel antibacterial delivery vehicle. The nisin-loaded nanoparticles were prepared using colloidal dispersion of the chitosan/alginate polymers in the presence of nisin. After the preparation of the nisin-loaded nanoparticles, their physicochemical properties such as size, shape, and zeta potential of the formulations were studied using scanning electron microscope and nanosizer instruments, consecutively. FTIR and differential scanning calorimetery studies were performed to investigate polymer–polymer or polymer–protein interactions. Next, the release kinetics and entrapment efficiency of the nisin-loaded nanoparticles were examined to assess the application potential of these formulations as a candidate vector. For measuring the antibacterial activity of the nisin-loaded nanoparticles, agar diffusion and MIC methods were employed. The samples under investigation for total microbial counts were pasteurized and raw milks each of which contained the nisin-loaded nanoparticles and inoculated Staphylococcus aureus (ATCC 19117 at 10⁶ CFU/mL), pasteurized and raw milks each included free nisin and S. aureus (10⁶ CFU/mL), and pasteurized and raw milks each had S. aureus (10⁶ CFU/mL) in as control. Total counts of S. aureus were measured after 24 and 48 h for the pasteurized milk samples and after the time intervals of 0, 6, 10, 14, 18, and 24 h for the raw milk samples, respectively. According to the results, entrapment efficiency of nisin inside of the nanoparticles was about 90–95%. The average size of the nanoparticles was 205 nm, and the average zeta potential of them was ?47 mV. In agar diffusion assay, an antibacterial activity (inhibition zone diameter, at 450 IU/mL) about 2 times higher than that of free nisin was observed for the nisin-loaded nanoparticles. MIC of the nisin-loaded nanoparticles (0.5 mg/mL) was about four times less than that of free nisin (2 mg/mL). Evaluation of the kinetic of the growth of S. aureus based on the total counts in the raw and pasteurized milks revealed that the nisin-loaded nanoparticles were able to inhibit more effectively the growth of S. aureus than free nisin during longer incubation periods. In other words, the decrease in the population of S. aureus for free nisin and the nisin-loaded nanoparticles in pasteurized milk was the same after 24 h of incubation while lessening in the growth of S. aureus was more marked for the nisin-loaded nanoparticles than the samples containing only free nisin after 48 h of incubation. Although the same growth reduction profile in S. aureus was noticed for free nisin and the nisin-loaded nanoparticles in the raw milk up to 14 h of incubation, after this time the nisin-loaded nanoparticles showed higher growth inhibition than free nisin. Since, generally, naked nisin has greater interactions with the ingredients present in milk samples in comparison with the protected nisin. Therefore, it is concluded that the antibacterial activity of nisin naturally decreases more during longer times of incubation than the protected nisin with the chitosan/alginate nanoparticles. Consequently, this protection increases and keeps antibacterial efficiency of nisin in comparison with free nisin during longer times of storage. These results can pave the way for further research and use of these nanoparticles as new antimicrobial agents in various realms of dairy products.     

Influence of growth conditions on the nisin production of bioengineered <Emphasis Type="Italic">Lactococcus lactis</Emphasis> strains

Nisin production of three bioengineered strains, (LAC338, LAC339 and LAC340) with immunity (nisFEG) and/or regulation (nisRK) genes of nisin biosynthesis on plasmids in the Lactococcus lactis LL27 nisin producer, was evaluated under pH-controlled and pH-uncontrolled batch fermentations. Optimization studies showed that fructose and yeast extract yielded the highest nisin activity. The strains LAC338, LAC339, and LAC340 produced 24, 45, and 44% more nisin, respectively, than wild-type L. lactis LL27 after 12-h incubation. However, sharp decreases in the yield of nisin were observed at the late phase of fermentation with LAC339 and LL27 in contrast to LAC340 and LAC338 strains for which the high level of nisin could be maintained longer. Obviously, increasing the copy number of the regulation genes together with immunity genes in the nisin producers retarded the loss of nisin in the late phase of the fermentation.     

Comparative effectiveness of nisin and bacteriocin J46 at different pH values

E. HUOT, C. BARRENA-GONZALEZ AND H. PETITDEMANGE. 1996. A Comparative study of the inhibitory activity of nisin, the well-known lantibiotic produced by certain strains of Lactococcus lactis subsp. lactis , and of the bacteriocin produced by L. lactis subsp. cremoris J46, a strain previously isolated from fermented milk, was conducted. For both bacteriocins, the activity against L. lactis subsp. cremoris decreased with increasing pH. In addition, the bacteriocin preparations were more stable at 4 than at 20°C. The influence of the storage temperature was more crucial for nisin. Essentially the same activity was observed for bacteriocin J46 stored for 3 h at 4 or 20°C. More interesting was the observed stability of bacteriocin J46 at pH values between 5.8 and 6.8. For example, about 23% of nisin activity was lost at pH 6.4 whereas no loss of bacteriocin J46 activity was observed.     

Influences on the antimicrobial activity of surface-adsorbed nisin

The efficacy of the antimicrobial peptide nisin was examined after adsorption to silica surfaces. Three protocols were used to evaluate nisin's activity against adhered cells ofListeria monocytogenes: bioassay usingPediococcus pentosaceous FBB 61-2 as the sensitive indicator strain; visualization and enumeration of cells by microscopic image analysis; and viability of adhered cells as determined by lodonitrotetrazolium violet uptake and crystallization. The activity of adsorbed nisin was highly dependent upon conditions of adsorption. The highest antimicrobial activity of adsorbed nisin occurred with high concentrations of nisin (1.0 mg ml^–1) and brief contact times (1 h) on surfaces of low hydrophobicity. Sequential adsorption of a second protein (-lactoglobulin or bovine serum albumin) onto surfaces consistently resulted in decreased nisin activity. These data provide direction for the development of applications to limit microbial attachment on food contact surfaces through the use of adsorbed antimicrobial peptides.     

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

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 μg/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.