Crude Bacteriocins Research Articles

PCR-DGGE analysis of population dynamics of lactic acid bacteria in kimchi by addition of bacteriocins.

The effects of two crude bacteriocins (DF01 and K10) on lactic acid bacteria (LAB) communities and pH during kimchi fermentation were analyzed by polymerase chain reactiondenaturing gradient gel electrophoresis (PCR-DGGE). Crude LAB bacteriocins, prepared by ammonium sulfate precipitation, were added at 5 AU/mL, and kimchi was incubated at 20°C for 7 days. The pH and titratable acidity of the kimchies were determined daily, and the amplified 16S rRNA products were analyzed by PCR-DGGE. The common and main LAB were Weissella spp., Leuconostoc spp., and Lactobacillus spp. from both control and bacteriocin-treated samples. Among them, W. koreensis, W. confusa, and Lb. sakei were the predominant microorganisms throughout the fermentation period. Some obligate and facultatively heterofermentative LAB were detected from the bacteriocin-treated samples. The pH of the kimchi samples treated with each bacteriocin was higher (ca. 0.8 unit) than that of the control at day 4 and 5.

Effects of PH and Storage Temperatures on Antibacterial Activity of Bacteriocin Produced by Lactic Acid Bacteria Isolated from OGI

Aims: Lactic acid bacteria are important organisms recognized for their fermentative ability. They produce various compounds including bacteriocins which are proteinaceous antimicrobial compounds. The purpose of this study was to evaluate the lactic acid bacterial content of ogi, extract crude bacteriocin from the lactic acid bacterial isolate(s) and determine the effects of pH and storage temperature on the antimicrobial properties of the crude bacteriocin extract. Study Design: The study was designed to isolate and characterise lactic acid bacteria from ogi, thereafter extract crude bacteriocins from the isolates and then determine the antibacterial activities of the bacteriocins so extracted against some known indicator organisms. Place and Duration of Study: Ogi samples were purchased from vendors who hawk it around the Benson Idahosa University campus and the study was done at the Department of Basic Sciences, Faculty of Basic and Applied Sciences, Benson Idahosa University, Benin City, Nigeria; between Short Communication Ohenhen et al.; BMRJ, 9(3): 1-9, 2015; Article no.BMRJ.13471 2 October 2012 and June 2013. Methodology: Preliminary isolation and characterization of lactic acid bacteria was done using standard microbiological methods after which the bacteriocins were extracted by propagating in 500ml MRS broth (pH 7.0 glucose 0.25% w/v) and incubated for 72 h at 30oC under anaerobic conditions. Extract was obtain by centrifuging cultures at 10,000 rpm for 20 min. Antimicrobial activity of the extract was determined using agar well diffusion method. Indicator organisms utilized were; Escherichia coli and Staphylococcus aureus. The effects of pH and storage temperatures of the crude bacteriocin, on the antimicrobial properties were determined using standard methods. Results: Lactobacillus plantarum, Lactobacillus casei, Lactobacillus brevis, Lactobacillus fermentum and Lactobacillus jensenii were the lactic acid bacteria species isolated and identified from the Ogi samples. Lactobacillus plantarum had the highest zone of inhibition during the screening test for antibacterial activity and was used to produce the bacteriocins used for the subsequent tests. At pH 2, there was a high antimicrobial activity but at pH 10, there was no antimicrobial activity. Crude bacteriocin extract stored at -20oC also showed the highest antimicrobial activity. Conclusion: This study has established that Ogi is a viable source of several lactobacilli which are capable of producing several antimicrobial compounds such as bacteriocins. It also established that bacteriocins recovered from Lactobacillus plantarum have a potent antimicrobial activity against a variety of both diarrhoeagenic and spoilage bacterial.

Amoebicidal Effects of Three Bacteriocin like Substances from Lactic Acid Bacteria against Acanthamoeba Polyphaga

We investigated the antiamoebic activity of three Bacteriocin Like Substances (BLS 39, BLS GS 54, BLS GS 16) produced by Lactic Acid Bacteria (LAB). The crude bacteriocins showed an amoebicidal effect against Acanthamoeba polyphaga, but with differences. BLS 39, produced by Lactobacillus pentosus, determined a prompt and progressive decrease of viable amoebal cell count, up to the end of the experiment (144 h), where the trophozoites were not detectable. A killing effect, but after a more prolonged contact time, was observed for BLS GS 54, produced by Lactobacillus paraplantarum, whereas the bacteriocin produced by Lactobacillus plantarum GS16 showed the lowest toxicity for A. polyphaga. For BLS GS 16 the maximum percentage of reduction in trophozoites count (45%) was obtained after 144 h, value much lower when compared to BLS GS54 and BLS 39, that showed values of 44,60% and 52,60% after only one hour of contact, with a maximum of 98% and 100% of non-viable cells, respectively, after 144 h. Morphological changes of the A. polyphaga cells as swollen cells, roundness and cellular lysis, were already observed after the first hours of contact with BLS and, at the end of the experiment, most of the cells were colored (blue), indicating their death. Currently there isn$prime;t evidence of BLS produced by LAB active against amoebas. In this study we have shown that all the three BLS secreted by the Lactic Acid Bacteria are endowed with amoebicidal effect against Acanthamoeba polyphaga, killing the protozoan with different effectiveness and at different times of contact.

Antibacterial activity and genotypic–phenotypic characteristics of bacteriocin-producing Bacillus subtilis KKU213: Potential as a probiotic strain

The antimicrobial activity and probiotic properties of Bacillus subtilis strain KKU213, isolated from local soil, were investigated. The cell-free supernatant (CFS) of a KKU213 culture containing crude bacteriocins exhibited inhibitory effects on Gram-positive bacteria, including Bacillus cereus, Listeria monocytogenes, Micrococcus luteus, and Staphylococcus aureus. The antibacterial activity of the CFS precipitated with 40% ammonium sulfate (AS) remained even after treatment at 60 and 100°C, at pH 4 and 10 and with proteolytic enzymes, detergents and heavy metals. When analyzed by SDS-PAGE and overlaid with the indicator strains B. cereus and S. aureus, the 40% AS precipitate exhibited inhibitory activity on proteins smaller than 10kDa. However, proteins larger than 25kDa and smaller than 10kDa were still observed on a native protein gel. Purified subtilosin A was prepared by Amberlite XAD-16 bead extraction and HPLC and analyzed by Nano-LC-QTOF-MS. Its molecular mass was found to be 3.4kDa, and it retained its antibacterial activity. These results are consistent with the detection of the anti-listerial subtilosin A gene of the sbo/alb cluster in the KKU213 strain, which is 100% identical to that of B. subtilis subsp. subtilis 168. In addition to stable and cyclic subtilosin A, a mixture of many extracellular antibacterial peptides was also detected in the KKU213 culture. The KKU213 strain produced extracellular amylase, cellulase, lipase and protease, is highly acid-resistant (pH 2) when cultured in inulin and promotes health and reduces infection of intestinally colonized broiler chickens. Therefore, we propose that bacteriocin-producing B. subtilis KKU213 could be used as a potential probiotic strain or protective culture.

Antimicrobial Activity of Cell Free Supernatant of Lactic Acid Bacteria Isolated from Fermented Durian Flesh against Multiple Antibiotic Resistance’s Salmonella Associated with Food Poisoning Cases in Malaysia

Epidemic of food poisoning cases and outbreaks associated with Salmonella spp. have been increased all over the world. Most of food poisoning cases in Malaysia related to Salmonella are associated with contaminated water compared to other sources. Finding alternative treatment to reduce foodborne cases becomes crucial, and bio-preservation using lactic acid bacteria (LAB) is one of strategy that can be used in food industries. The objective of this study was to determine the antimicrobial activities in vitro of cell free supernatant from eight LAB strains isolated from fermented durian flesh, which were 5 strains from Lactobacillus buchneri, 1 strain from each culture, Lactobacillus plantarum, Lactobacillus brevis 1 and Lactobacillus acidophilus 1 against 23 species of various serotypes of pathogenic Salmonella obtained from food poisoning cases in Malaysia. The antimicrobial activity of crude bacteriocins from cell free supernatant (CFS) of LAB strains were tested using well diffusion method. Susceptibility test of eight selected Salmonella strains was done using disk diffusion method (DDT) against twelve selected antibiotics. Results showed that crude bacteriocins of Lb. plantarum, Lb. buchneri, Lb. brevis 1 and Lb. acidophilus 1 from fermented durian flesh had strong inhibitory effect against most of 23 species of MAR's Salmonella associated with food poisoning cases with intermediate to high inhibitory zone by average mean from 16 mm to 22 mm diameter zone. All eight Salmonella strains were resistant to bacitracin, nalidixic acid and vancomycin and some strains were also resistant to ampicillin, penicillin, streptomycin and tetracycline. Further study could be utilised to understand the mechanism of selected LAB bacteriocin producer from fermented durian flesh as antimicrobial substance.

CRUDE BACTERIOCIN EXTRACT EFFECT ON THE MICROBIOLOGICAL AND SENSORIAL CHARACTERISTICS OF SOLOMO REDONDO (</I>Longissimus dorsi</I>) VACUUM-PACKAGING

The application of crude bacteriocins extract (CBE) produced by Lactobacillus plantarum LPBM10 is tested on the microbiological and sensory characteristics of rib filets (Longissimus dorsi) vacuum packing for 31 days storage at 3°C. As parameters of meat quality through time microbiology analysis, pH, cutting force, weight loss and sensorial are made. Significant differences are found in psychrotrophic counts in the fifth day storage, being better the treatment with crude bacteriocin extract. Levels of fecal coliforms persistaround 1.5 UFC/g in meat samples with crude bacteriocins extract. Significant difference in every day storage (p

Inhibition of Listeria monocytogenes by In Situ Produced and Semipurified Bacteriocins of Carnobacterium spp. on Vacuum-Packed, Refrigerated Cold-Smoked Salmon

Listeria monocytogenes inhibition by Carnobacterium strains and crude bacteriocins on sterile and commercial vacuum-packed cold-smoked salmon stored at 4°C and 8°C was investigated. Carnobacterium piscicola V1 was bactericidal against L. monocytogenes at the two temperatures, whereas Carnobacterium divergens V41 presented a bacteriostatic effect. C. piscicola SF668 delayed L. monocytogenes growth at 8°C and had a bacteriostatic effect at 4°C. Listeria growth was not affected by a non–bacteriocin-producing C. piscicola. Crude extracts of piscicocins were bactericidal at 4°C and 8°C. Listeria growth was delayed by divercin V41 at 8°C and was inhibited at 4°C. Nisin delayed Listeria growth at 8°C and was bacteriostatic at 4°C. The present study demonstrates that L. monocytogenes growth could be prevented on vacuum-packed cold-smoked salmon by Carnobacterium and associated bacteriocins at chilled temperatures. Moreover, no product spoilage could be observed with the use of such bacteriocin-producing strains as demonstrated by good sensorial analyses and low biogenic amine production.

Comparative antimicrobial activity of enterocin L50, pediocin PA-1, nisin A and lactocin S against spoilage and foodborne pathogenic bacteria

The present work compares, under the same stated experimental conditions, the antimicrobial activity of crude and purified enterocin L50, pediocin PA-1, nisin A and lactocin S, produced by lactic acid bacteria (LAB) isolated from Spanish dry-fermented sausages. The bacteriocins were purified to homogeneity by ammonium sulphate precipitation, gel filtration (for lactocin S), and cation-exchange, hydrophobic-interaction, and reverse-phase-chromatography; high yields of pure bacteriocins were obtained. Minimal inhibitory concentration (MIC) of pure enterocin L50, pediocin PA-1, nisin A and lactocin S was determined against a broad spectrum of Gram-positive bacteria, including spoilage and foodborne pathogenic bacteria. The purified bacteriocins showed a broad antimicrobial spectrum similar to that exerted by crude bacteriocins. Enterocin L50 and pediocin PA-1 were very active againstListeria monocytogenes, which was quite resistant to nisin A and lactocin S. Enterocin L50 also displayed antimicrobial activity againstStaphylococcus aureus,Clostridium perfringensandClostridium botulinum. However, these pathogens were weakly inhibited, or not at all, by the other pure bacteriocins.

Behavior of Listeria monocytogenes in Mozzarella cheese in presence of Lactococcus lactis

The behavior of Listeria monocytogenes (Scott A) on fully processed Italian Mozzarella cheese was examined in presence and in absence of bacteriocins produced by Lactococcus lactis ssp. lactis strains (DIP 15 and DIP 16). These strains, isolated from raw milk, produced heat stable bacteriocins that were inactivated by pronase, alpha- chymotrypsin and proteinase K, but not by pepsin, trypsin and catalase. The addition of crude bacteriocins to the growing culture of Listeria monocytogenes resulted in a significant reduction in cell number at 5 °C, but not at 30 °C. Mozzarella cheese was inoculated with the Listeria culture to obtain an initial level of approximately 30 CFU/cm 2 surface of Mozzarella and approximately 10 3 CFU/ml of the surrounding fluid and then packaged in bags containing the heat-treated neutralized-cultures of Lactococcus lactis ssp. lactis in skim milk (in Italy, Mozzarella is sold in small size pieces, individually packaged in bags containing some fluid). Bags were stored at 5 °C up to 21 days. The presence of bacteriocins resulted in apparent death of Listeria monocytogenes after 24 h storage. After 7 days of storage, a revival of Listeria monocytogenes was observed, followed by an increase in number. However, for a storage period of 2–3 weeks the number of Listeria monocytogenes remained significantly below the number observed for Mozzarella cheese packaged in absence of the heat-treated cultures of Lactococcus lactis.