Antibacterial activity of plant essential oils and extracts: The role of thyme essential oil,nisin, and their combination to control Listeria monocytogenes inoculated in minced fish meat

Food poisoning caused by Listeria monocytogenes leads to a 30% rate of mortality among patients. The antibacterial activity of cinnamon, thyme, and rosemary essential oils (EOs) and shallot and turmeric extracts was tested against L. monocytogenes using agar well and disc diffusion techniques. Results showed that thyme EO had the highest antimicrobial activity, followed by cinnamon and rosemary EOs, respectively. The antilisterial activity of thyme EO at 0.4%, 0.8%, and 1.2% levels, nisin at 500 or 1000 IU/g level, and their combination against L. monocytogenes was examined in minced fish samples. The antilisterial properties of nisin were also investigated in cooked minced fish treatments. Nisin at 500 or 1000 IU/g in the minced fish meat demonstrated bacteriostatic activity against L. monocytogenes. The use of thyme EO at 0.8% and 1.2% reduced the L. monocytogenes viable count below 2 log cfu/g after 6 days. Furthermore, simultaneous use of thyme EO at 0.8% and 1.2%, and nisin at 500 or 1000 IU/g level, reduced the L. monocytogenes viable count below 2 log cfu/g after the second day of storage. The antilisterial activity of nisin in the cooked minced fish samples was slightly stronger than that of the raw group.     

In vitro assessment of synthetic phenolic antioxidants for inhibition of foodborne Staphylococcus aureus,Bacillus cereus and Pseudomonas fluorescens

Minimal inhibitory concentrations (MICs) of seven synthetic phenolic compounds, five commonly used as antioxidants (TBHQ, BHA, BHT, propyl gallate and octyl gallate) and two as antimicrobials (propyl 4-hydroxybenzoate and n-heptyl 4-hydroxybenzoate) were assessed against several strains of two Gram-positive (Staphylococcus aureus and Bacillus cereus) and one Gram-negative (Pseudomonas fluorescens) bacteria, by using a standardized microdilution assay (ISO 20776-1, 2006). Octyl gallate was the most effective compound against the three genera/species of bacteria considered simultaneously (with the exception of four strains of B. cereus, which were resistant for this compound) with MIC values (≤100 μg/ml) lower than the concentrations usually used as antioxidants. TBHQ and n-heptyl 4-hydroxybenzoate were also effective in the control of S. aureus at very low concentrations (MIC of 3.1 μg/ml and 12.5 μg/ml, respectively). Propyl 4-hydroxybenzoate was the most inhibitory phenolic compound against all strains of B. cereus and both tested parabens (propyl- and heptyl-) were not effective for P. fluorescens (MIC > 1600 μg/ml). B. cereus was the bacterial genera that showed more intra-species variation, distinguishing two clearly groups of sensitivity among the strains to octyl gallate and n-heptyl 4-hydroxybenzoate (“sensitive” with mean MICs of 42.8 and 4.2 μg/ml, respectively; and “resistant” with MICs >1600 and >800 μg/ml, respectively). According to all that, octyl gallate would be an interesting phenolic compound for the food industry, not only because of its recognized antioxidant properties but also because of their effectivity as antimicrobial against S. aureus, B. cereus and P. fluorescens.     

Effect of detergents as antibacterial agents on biofilm of antibiotics-resistant Vibrio parahaemolyticus isolates

Vibrio parahaemolyticus (V. parahaemolyticus) is a halophilic, Gram-negative human pathogen known as a leading cause of seafood-derived food poisoning. Due to high contamination rate of seafood in Asian countries, V. parahaemolyticus is considered as a food safety concern. V. parahaemolyticus is able to produce biofilm which is more resistant toward disinfectants and antibodies than its planktonic form. Thirty six V. parahaemolyticus isolates from seafood were tested for their susceptibility using 18 different antibiotics. Two V. parahaemolyticus isolates were resistant to bacitracin, chloramphenicol, rifampin, ampicillin, vancomycin, nalidixic acid, penicillin and spectinomycin. Fourteen V. parahaemolyticus isolates were found to be resistant to bacitracin, tetracycline, rifampin, ampicillin, vancomycin, penicillin and spectinomycin. The remaining two isolates were resistant to more than 2 antibiotics. Majority of the V. parahaemolyticus isolates (97.2%) showed MAR index > 0.2, indicating that these isolates were originated from high risk sources. To investigate effect of three common detergents on antibacterial-resistant V. parahaemolyticus, 16 V. parahaemolyticus isolates resistant to more than 7 antibiotics were selected. V. parahaemolyticus (ATCC 17802) was used as reference strain. Detergents were tested for their minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) and time–kill curves were constructed to assess the concentration between MIC and bactericidal activity of detergents. Detergents D1 (Linear alkyl benzene based) was found to be the most effective with MIC and MBC ranged between 97.656 and 1562.5 μg/ml and 781.25–3125 μg/ml, respectively. The time–kill curves demonstrated that the bactericidal endpoint for resistant V. parahaemolyticus isolates reached after 30 min incubation with D1 at concentration 8 × MIC. The isolate VP003 was killed at 8 × MIC within 0.5 h and the reduction in CFU/ml was 3 log units (99.9%). V. parahaemolyticus biofilms were formed in 96 wells microtiter plates at 37 °C and 24 h-old biofilm were used to test antibacterial activity of detergents. Results showed that biofilm-producing ability of antibacterial-resistant V. parahaemolyticus isolates were inhibited at 1562.5–6250 μg/ml of D1 and eradicated at 3125 – ≥50,000 μg/ml of D1. Detergents showed potential antimicrobial activity against V. parahaemolyticus     

Surfactant assisted nisin loaded chitosan-carageenan nanocapsule synthesis for controlling food pathogens

Nisin loaded chitosan/carageenan nanocapsules were prepared using an ionic complexation method and evaluated for antibacterial activity. Concentration of polymers as well as surfactant affected particle size and encapsulation efficiency. The optimal parameters for synthesis of particles with least size and highest encapsulation efficiency were concentration of caragennan 0.75 mg/ml along with concentration of chitosan 10 mg/ml and surfactant 1 μg/ml. Release study conducted in vitro over a period of two weeks indicated slow and sustained release from the formulation. Furthermore, evaluation of antibacterial activity against Micrococcus luteus MTCC (Microbial Type Culture Collection) 1809, Pseudomonas aeruginosa MTCC 424 and Salmonella enterica MTCC 1253 and Enterobactor aerogenes MTCC2823 in vitro followed by antibacterial activity in tomato juice for six months, indicated that encapsulated nanocapsules showed better antibacterial effect on microbe's in vitro as well as in tomato juice for prolonged periods as compared to the components evaluated separately.     

Studies on antibacterial activity and antibacterial mechanism of a novel polysaccharide from Streptomyces virginia H03

A novel polysaccharide isolated from the broth of Streptomyces virginia H03 exhibited strong antibacterial activities on food spoilage and food poisoning microorganisms such as Staphylococcus aureus, Bacillus subtilis, Listeria monocytogenes, Escherichia coli, Zygosaccharomyces bailii and Candida utilis. The minimal inhibitory concentrations of the polysaccharide determined by the double broth dilution method were 15.6, 31.3, 31.3, 31.3, 125 and 125 (μg/ml) against the microorganisms mentioned above, respectively. The possible targets of the polysaccharide in bacteria might be cell wall, cytoplasmic membrane and DNA as indicated by electron microscopy, leakage of protein, cytoplasmic membrane permeability and DNA binding, respectively. Moreover, the antibacterial activity remained unchanged after being treated at 100 °C for 10 min, which indicated that the polysaccharide had a good heat-stability. In addition, it was safe according to mouse toxicity. All of these suggested that the polysaccharide might be used as a potential antimicrobial in food.     

Antibacterial activity and chemical constitutions of Ziziphora clinopodioides

The essential oil and methanol extract obtained from aerial parts of Ziziphora clinopodioides Lam. harvested in the Eastern part of Turkey were evaluated for their chemical composition and antibacterial activity against 52 Gram-positive and Gram-negative bacteria. The GC–MS analyses allowed 18 compounds to be determined; the main constituents of the essential oils were (+)-pulegone (31.86%), 1,8-cineole (12.21%), limonene (10.48%), menthol (9.13%), β–pinene (6.88%), menthone (6.73%), piperitenone (5.30%) and piperitone (4.18%) The essential oil and methanol extract had a broad-spectrum antimicrobial activity against pathogens in broth microdilution bioassays. Maximum activity of essential oil (>22 mm) and methanol extract (>11 mm) was observed against Acidovorax facilis, Bacillus flexus, Bacillus spp, Bacillus sphaericus, Brevibacillus brevis, Corynebacterium ammoniagenes, Enterobacter sakazakii, Erwinia carotovora carotovora, Moraxella catarrhalis and Xanthomonas arboricola.     

Antimicrobial peptides derived from hen egg lysozyme with inhibitory effect against Bacillus species

In food industry, Bacillus species are encountered in deteriorating many food products thus shortening their shelf-life. Moreover, Bacillus cereus and the subtilis group (Bacillus subtilis, Bacillus licheniformis, and Bacillus pumilus) have been recognized as food poisoning agents. Lysozyme peptides preparation (LzP) is a commercially available as a natural food preservative. Although, LzP derived from lysozyme yet it showed only 11% of the lysozyme lytic activity. LzP at a concentration of 100 μg ml⁻¹ completely inhibited B. subtilis, B. licheniformis, B. megaterium, B. mycoides, B. pumilus, B. coagulans, B. amyloliquefaciens, B. polymexa and B. macerans. However, B. cereus and B. stearothermophilus showed a slightly higher resistance. Interestingly, LzP at concentration ⩾10 μg ml⁻¹ showed inhibitory effect on both vegetative and spore forms of B. subtilis. Moreover, LzP was stable at 95 °C for 30 min and at different pH values (4.5–7). In conclusion, LzP may be useful to control growth of Bacillus spoilage organisms.     

Endophytic bacterial diversity in Korean kimchi made of Chinese cabbage leaves and their antimicrobial activity against pathogens

The diversity of bacterial endophytes associated with Chinese cabbage (CC) leaves from two cultivation areas in Korea, namely, Seosang-gun (SS) and Haenam-gun (HN), and from a laboratory transgenic plant (TP) was investigated. A total of 5.76 log CFU/g isolates were identified from the interior leaves of the three cultivars of CC, which were grouped into 5.08, 5.36, and 5.34 log CFU/g from SS, HN, and TP samples, respectively. Phylogenetic analysis based on the 16S rRNA sequences indicated that the isolates belonged to four major phylogenetic groups: high-G + C Gram-positive bacteria (HGCGPB), low-G + C Gram-positive bacteria (LGCGPB), Proteobacteria, and Bacteroidetes. The most predominant group of species in the leaves of the SS, HN and TP CC cultivars were LGCGPB (50%), LGCGPB (65.2%), and HGCGPB (43%), respectively. A total of 23 bacterial genera were identified from the three cultivars of Chinese cabbage. Most extracellular hydrolytic enzyme-producing colonies among the isolates belong to the genus Bacillus those were predominant in the HN cultivars. In addition, some isolates especially Bacillus sp. (TPL08), Bacillus subtilis (SSL16, HNL10) exhibited potential activity as biocontrol agents against food-borne pathogenic bacteria and phytopathogenic fungi tested in this study. This study first revealed the endophytic bacterial communities in leaves of Chinese cabbage (main ingredients of kimchi) grown in Korea.     

Isolation, identification and monitoring of contaminant bacteria in Iranian Kefir type drink by 16S rDNA sequencing

Iranian Kefir type drink (IKTD) is a highly consumed, traditional Iranian, fermented milk product. To improve monitoring procedures for food safety 32 industrial Kefir type drinks from 4 brands and 8 different production dates as well as 32 samples from pasteurized milk of the same Kefir manufacturers and air of the production sites were analyzed for contaminations. 16S rDNA extraction from Kefir samples as well as 16S rDNA obtained from samples incubated on Columbia agar were analyzed using PCR/DGGE, cloning, sequencing and phylogenetic classification. Already DGGE analysis indicated contaminations including Bacillus strains. Subsequently analysis of cultured clones indicated contaminations with Bacillus sp. including Bacillus cereus, Bacillus thuringiensis and Paenibacillus sp. in 9 (28%) from all analyzed samples. Also 38% of pasteurized milk samples were contaminated with B. cereus. The average count of B. cereus was 74 ± 19 cfu/ml. B. cereus and B. thuringiensis were found as contaminant bacteria in the air of the all manufacturing sites. These results suggest that milk is one of the most important sources of contamination with Bacillus sp., especially B. cereus for Kefir products in Iran. But bacterial contamination in Kefir samples might also originate from the air of the production sites. 16S rDNA analysis accelerates monitoring strategies.     

Inhibition of foodborne pathogens by Hf-1, a novel antibacterial peptide from the larvae of the housefly (Musca domestica) in medium and orange juice

Antibacterial molecules from edible insects may serve as a potentially significant group of food preservatives. Hf-1, a novel peptide from larvae of housefly (Musca domestica) with molecular weight approximately 8.0 KD, inhibited food pathogens Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27553, Salmonella typhimurium 50013, Shigella dysenteriae 51302, Staphylococcus aureus 6538 and Bacillus subtilis 9372 with MIC ranging from 18 to 72 μg/ml. The presence of EDTA can enhance Hf-1 activity against Gram-negative bacteria. Hf-1 was bactericidal in orange juice, showing an antibacterial activity approximately equivalent to sodium benzoate. The mode of action of Hf-1 involved decreasing the cell surface hydrophobicity and damaging the cell membrane. The results suggested Hf-1 has the potential to be used as the food preservative.     

Essential oils composition and bioactivities of two species leaves used as packaging materials in Xishuangbanna,China

This study reports the essential oils chemical composition and antioxidant and antimicrobial potentials of the leaves of Phrynium pubinerve Blume and Thysanolaena latifolia (Roxb. ex Hornem.) Honda, which are used as natural packaging materials by ethnic groups in Xishuangbanna, southwest China. GC–MS analysis identified 46 and 21 components, representing 88.6% and 93.4% of the essential oils of P. pubinerve and T. latifolia, respectively. The major constituents for P. pubinerve were (Z)-3-hexen-1-ol (17.31%), (E)-2-hexenal (9.01%) and 1-hexanol (8.61%). The major constituents for T. latifolia were (Z)-3-hexen-1-ol (28.79%), phytol (12.30) and (E)-β-ionone (9.54%). Both the essential oils and ethanol extracts showed antioxidant activity in DPPH test (IC₅₀ values = 192.47–706.07 μg/ml), ABTS assay (IC₅₀ values = 35.54–134.97 μg/ml) and FRAP assays. The essential oils showed considerable antimicrobial activity against pathogenic bacteria and spoilage organisms, with MIC and MBC values in the ranges of 64–3072 μg/ml and 64–4096 μg/ml, respectively. The bioactivities of these two plant species validate the traditional use of these two plants, suggesting that both could be new sources of natural antioxidant and antimicrobial agents for the packaging, medical and functional food industries.     

Inhibition of Listeria monocytogenes by pomegranate (Punica granatum) peel extract in meat paté at different temperatures

Natural antimicrobials are being more and more considered as alternative approach for controlling growth of microorganisms in food. The objective of this study was to evaluate the pomegranate extract’s (PE) potential to be used as a natural preservative in ready to eat meats. Listeria monocytogenes was the main target. In a preliminary assessment with the disk diffusion method PE showed inhibitory effect against all five tested species, in the following order of increasing sensitivity: L. monocytogenes, Bacillus subtilis, Bacillus cereus, Escherichia coli and Staphylococcus aureus.No viable cells of L. monocytogenes were detected after incubation in BHI broth in presence of 7.5% v/v of the liquid PE (or 24.7 mg dry PE/ml). This concentration was considered as the Minimal Bactericidal Concentration (MBC) of the tested PE. Two pure components commonly found in PE, namely gallic and ellagic acids were also tested in BHI broth, however they did not show considerable inhibition of L. monocytogenes. PE in a concentration equal to the measured MBC was tested against L. monocytogenes in meat paté at different temperatures. At 4 °C during 46 days the extract inhibited the growth in meat paté by 4.1 log CFU/g compared to the control, which had reached log 9.2 CFU/g already on the 18th day. Inhibition was less pronounced at higher temperatures. The results indicate that the PE has a potential to be used as a natural preservative in meat products.     

Antibacterial activity and mechanism of pinoresinol from Cinnamomum Camphora leaves against food-related bacteria

Pinoresinol from the leaves of Cinnamomum camphora used as a model natural compound was extracted and isolated. The chemical characterization of the compound was confirmed using UV, IR, MS, H NMR and C HMR. The purity of the compound (86%) was determined using HPLC technique and it was used for further experiments. Its antibacterial activities against five food-related bacteria (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus subtilis, Salmonella enterica) were detected using both broth dilution method and disk diffusion method. Results indicated that 62.5 μg/mL pinoresinol completely inhibit the growth of five bacteria. Much effort was focused on elucidating the mechanism of antibacterial action of pinoresinol against P. aeruginosa and B. subtilis by determination of cell permeability and observing the changes of cell microstructure using scanning electron microscope (SEM). The leakage of the soluble saccharides and proteins were detected and up to 13.89 and 21.76 μg/mL, respectively for P. aeruginosa and 11.08 and 19.05 μg/mL, respectively for B. subtilis. Measurements of the release of the soluble saccharides and proteins confirmed the disruptive action of pinoresinol on cytoplasmic membrane. The damages of the pinoresinol on cell wall were confirmed using Gram staining and scanning electron microscopy (SEM). We concluded that the antibacterial action mechanism of the pinoresinol is attributed to the physiological and morphological changes in bacterial cells.     

Antimicrobial activity of rhamnolipids against Listeria monocytogenes and their synergistic interaction with nisin

The rhamnolipids (RL) are biodegradable biosurfactants which have low toxicity and surface active properties that can be useful for food processing industries. The objective of this study was to evaluate the antimicrobial potential of rhamnolipids against Listeria monocytogenes. Susceptibility tests were performed by the minimal inhibitory concentration (MIC) using the micro-broth dilution technique. The MIC values varied from 78.1 μg/mL to 2500 μg/mL with the 2500 μg/mL being the predominant value. Among the 32 tested cultures, 90.6% were susceptible to RL. Results showed that the rhamnolipid activity was primarily bacteriostatic. The interaction of rhamnolipid with nisin was also investigated. The combined effect of nisin and RL was evaluated against two wild-type isolates of L. monocytogenes, L12 sensitive to RL (MIC 156.2 μg/mL) and L17 less sensitive to RL (2500 μg/mL). The FIC indexes for the isolates were 0.18 and 0.078 for L12 and L17 respectively, indicating a strong synergistic effect. The survival curve of isolates L12 and L17 showed that the combination between nisin and RL was bactericidal at lower concentration than for the individual antimicrobials. For the L12 isolate 78.1 μg/mL of RL and 160 IU/mL of nisin eliminated the population after 30 min of incubation. The combination of 156.2 μg/mL of RL and 320 IU/mL of nisin reduced completely L17 population after 2 h of incubation. Rhamnolipids showed antimicrobial activity against L. monocytogenes and presented a synergistic effect when combined with nisin.     

Efficacy of pulsed electric fields for the inactivation of indicator microorganisms and foodborne pathogens in liquids and raw chicken

This study investigated the ability of pulsed electric fields (PEF) to inactivate a range of microorganisms in liquid media and on raw chicken meat. The susceptibilities of ten Campylobacter isolates (seven Campylobacter jejuni isolates and three Campylobacter coli isolates), Escherichia coli (ATCC 25922) and Salmonella Enteritidis (ATCC 13076) to PEF in liquid media were investigated. All Campylobacter isolates tested in liquid were susceptible to PEF treatment (65 kV/cm, 5 μs, 500 Hz) with reductions of between 4.33 and 7.22 log₁₀ CFU/mL observed for the longest treatment (30 s). Significant differences in susceptibility were observed between Campylobacter isolates subjected to equivalent PEF treatments ranging from 2.41 to 5.19 log₁₀ CFU/mL. Campylobacter isolates processed in liquid media were found to be more sensitive to PEF than E. coli and S. Enteritidis (P < 0.05). The application of PEF (3.75 and 15 kV/cm, 10 μs, 5 Hz) to inoculated samples of raw chicken did not result in any significant reductions in total viable counts, Enterobacteriaceae, C. jejuni, E. coli or S. Enteritidis. Therefore, under the conditions used in this study, PEF technology may not be suitable as a food safety intervention measure for the control of microbial contaminants on broilers during processing although it may have potential to reduce contamination of process water (e.g. scald or spin chill water).     

Molecular characterisation and antimicrobial activity of bacteria associated with submerged lactic acid cassava fermentation

Molecular identification of microorganisms associated with submerged cassava fermentation was carried out and isolates of lactic acid bacteria (LAB) were examined for antimicrobial activity, including ability to produce antimicrobial peptides as a first step to define starter cultures for controlled cassava fermentations. A total of 75 isolates, including 41 LAB, 31 aerobic bacteria (AB) and three anaerobic bacteria were isolated from unfermented and fermenting cassava roots, cassava leaves and fermented cassava dough and identified by a combination of phenotypic tests and sequencing of 16S rRNA, rpoA, rpoB and pheS genes. Microbial diversity at interspecies and intraspecies level was screened by, respectively, PCR amplification of the 16S-23S rDNA intergenic transcribed spacer (ITS-PCR) and repetitive sequence based PCR (rep-PCR). Antimicrobial activity of LAB cultures and supernatants against indicator bacteria; Escherichia coli, Salmonella enterica serotype Typhimurium (S. Typhimurium), Bacillus cereus and Staphylococcus aureus was studied using agar diffusion tests. Furthermore, inactivation of indicator bacteria was investigated in both liquid medium and during controlled cassava fermentation. Results revealed a diversity of bacterial genera, species and subspecies associated with submerged cassava fermentation. DNA sequencing enabled identification of LAB isolates as Lactobacillus plantarum, Weissella confusa, Weissella paramesenteroides, Lactobacillus rhamnosus, Lactobacillus hilgardii, Lactobacillus paracasei, Leuconostoc mesenteroides, Enterococcus faecium, Enterococcus casseliflavus, and Pediococcus acidilactici. Lactobacillus spp. were the predominant LAB and were present in all cassava samples studied. Aerobic bacteria were predominantly Bacillus spp., including Bacillus subtilis, Bacillus amyloliquefaciens and B. cereus. Other species identified included Staphylococcus pasteuri and Clostridium beijerinkii. Cells, supernatants and cell free supernatants (CFS) of selected LAB isolates were able to inhibit both Gram positive and Gram negative pathogenic bacteria. LAB isolates inactivated all indicator organisms during controlled cassava fermentations, with a 4–6 log reduction after 48 h fermentation. The antimicrobial effect of the LAB was attributed to acid production.     

Antilisterial activity and stability of nanovesicle-encapsulated antimicrobial peptide P34 in milk

Bacillus sp. P34, a strain isolated from aquatic environments of Brazilian Amazon basin, produces a bacteriocin-like substance (BLS) which was encapsulated in nanovesicles prepared from partially purified soy lecithin. The efficiency of free and encapsulated BLS P34 to control the development of L. monocytogenes and maintenance of antimicrobial activity was assessed over time in milk. The antimicrobial activity of free and encapsulated BLS P34 decreased approximately 50% after 4 days of storage (<4 °C) in skim and whole milk. After this period there was not significant loss of activity up to 21 days. The viable counts of Listeria monocytogenes in skim and whole milk containing 3200 AU/ml of free or encapsulated BLS P34 were always lower than those observed in controls without bacteriocin at both 30 °C and 7 °C. At 1600 AU/ml concentration, free and encapsulated BLS P34 were inhibitory to L. monocytogenes in skim milk, when compared with the control at 7 days. Nanovesicle-encapsulated and free BLS P34 shows potential use as biopreservative for application in milk-derived products.     

Thyme oil to control Alternaria alternata in vitro and in vivo as fumigant and contact treatments

This study was conducted to evaluate the efficacy of thyme oil in suppressing Alternaria alternata (Fr.:Fr.) Keissl in vitro and in vivo as fumigant and contact treatments. Thyme oil possessed great fumigant and contact toxicity against A. alternata at different concentrations in vitro. Irreversible inhibition of fungal growth could be caused by exposure to 500 μL/L thyme oil as a fumigant for 6 and 12 days. The spores of A. alternata were completely suppressed 6 h after incubation in potato dextrose broth (PDB) by 2000 μL/L of thyme oil. The thyme oil as a fumigant at 66.7 μL/L showed a significant inhibition effect on A. alternata of cherry tomatoes stored at 25 °C for 5 days. Thyme oil at 500 μL/L showed a significant contact inhibition effect on A. alternata of cherry tomatoes stored at 25 °C for 3 days.     

Susceptibility of Clostridium perfringens to antimicrobials produced by lactic acid bacteria: Reuterin and nisin

The effectiveness as antimicrobials of lactic acid bacteria produced compounds reuterin and nisin was assessed against vegetative cells and spores of Clostridium perfringens isolates (from ovine milk obtained in farms with diarrheic lambs) and C. perfringens CECT 486 (type A toxin producer). We also tested the inhibitory effect of lysozyme and sodium nitrite on Clostridium. Minimal inhibitory concentrations (MIC) of antimicrobials were determined in modified RCM (mRCM) and milk by using a broth microdilution method, after 7 d at 37 °C under anaerobic conditions. The sensitivity of C. perfringens to the tested antimicrobials was strain and culture medium-dependent. In general, vegetative cells exhibited higher sensitivity than spores. Reuterin (MIC values 2.03–16.25 mM) inhibited the growth of vegetative cells and the outgrowth of spores of all tested C. perfringens, both in mRCM and milk, with higher resistance in milk. Nisin (MIC values 0.78–12.5 μg/ml) was also effective against vegetative cells and spores of tested C. perfringens in both culture media. However, lysozyme (up to 400 μg/ml) did not control the growth of any of the tested Clostridium. Sodium nitrite only inhibited the outgrowth of spores of two C. perfringens isolates at the maximum concentration assayed (300 μg/ml) exclusively in mRCM medium. These results suggest that reuterin and nisin have the potential to control the growth of C. perfringens, and might help to ensure safety at different stages of the food chain. Future studies in food/feed products would be necessary to further corroborate this hypothesis.     

Antibacterial characteristics and mechanisms of ɛ-poly-lysine against Escherichia coli and Staphylococcus aureus

The aim of this study was to investigate the antibacterial characteristics and antibacterial mechanisms of ɛ-poly-lysine against Escherichia coli and Staphylococcus aureus. The diameters of inhibition zones of E. coli (10 ± 0.5 mm) and S. aureus (12 ± 0.1 mm) treated by 200 μg/ml ɛ-poly-lysine were much larger than control (5 ± 0.3 mm) (p < 0.05). Minimum inhibition concentration of ɛ-poly-lysine against E. coli and S. aureus was 12.5 μg/ml. Scanning electron microscopy showed that ɛ-poly-lysine damaged the morphology of tested bacterial cells. The increase in electric conductivity of bacterial cells suspension indicated that the cytoplasmic membranes were broken by ɛ-poly-lysine, which caused leakage of ions in cells. SDS-PAGE of bacterial proteins demonstrated that ɛ-poly-lysine could damage bacterial cells through the destruction of cellular proteins. These results indicated that ɛ-poly-lysine has good potential to be as a natural food preservative.