Indicator Strains Research Articles

Effect of Cocultivation on Lactobacillus plantarum Strains Growth and Antagonistic Activity

The use of bacterial starters for the production of fermented foods has several advantages over traditional spontaneous fermentation, as it provides a rapid and controlled decrease of pH, improves the microbiological quality of the product, and prolongs the shelf-life. Fermented foods are typically produced using mixed cultures of lactic acid bacteria (LAB) due to the synergism between their constituent bacterial cultures. So, the compatibility of the LAB strains decides the efficacy of a multi-strain starter. The purpose of this study was to investigate the effect of the cocultivation of Lactobacillus plantarum strains on the growth, acidification, and antagonistic activity to determine suitable strain combinations for fermented vegetable production. Methods. The effect of cocultivation on growth characteristics of four L. plantarum strains was determined in MRS medium and cabbage-based medium with 2.5% NaCl. After 8 h of cultivation at 30°C and 37°C, the number of viable cells (CFU/ml) and the pH of the medium were determined. The antagonistic activity of monocultures of L. plantarum and their six compositions against opportunistic pathogenic microorganisms was determined by the method of delayed antagonism. Results. During growth in MRS broth at 30°C cocultivation of L. plantarum 47SM with L. plantarum 691T or L. plantarum 1047K strains led to enhanced rates of growth compared to the monocultures, suggesting some degree of symbiosis between these strains. Viable cell counts of L. plantarum 47SM, 1047K and 691T strains and ΔpH values of L. plantarum 952K, 1047K, and 691T strains were higher after 8 h growth in the cabbage-based medium at 30°C compared to MRS broth. Despite the intensive growth of L. plantarum monocultures in cabbage-based medium, a significant decrease of viable cell counts and ΔpH values during cocultivation at 30°C were found. Cocultivation did not affect the average size of the growth inhibition zones of most of the indicator strains used. However, growth inhibition zones of Shigella flexneri, Escherichia coli, and Proteus vulgaris decreased in some L. plantarum mixed cultures compared to monocultures. Thus, the growth inhibition zones of E. coli and S. flexneri by mixed culture L. plantarum 47 SM+1047K were significantly smaller compared to the growth inhibition zones of L. plantarum monocultures. Conclusions. Thus, based on the data obtained in present work, we can assume that some of these L. plantarum strains used in the work may be bactericinogenic. Although the four L. plantarum strains studied are compatible when cocultivated in a standard rich MRS medium, the results of cocultivation in a cabbage-based medium with 2.5% NaCl does not allow to recommend the use of these L. plantarum strains simultaneously in the starter for vegetable fermentation. Further investigation of bacteriocinogenic properties and mechanisms of growth inhibition under cocultivation in vegetable-like conditions are needed, which will allow combining of some of these L. plantarum strains with LAB strains of other species or genera to create multi-starters for vegetable fermentation.

3-Hydroxypropionic acid contributes to the antibacterial activity of glycerol metabolism by the food microbe Limosilactobacillus reuteri

Strains of Limosilactobacillus reuteri are used as starter and bioprotective cultures and contribute to the preservation of food through the production of fermentation metabolites lactic and acetic acid, and of the antimicrobial reuterin. Reuterin consists of acrolein and 3-hydroxypropionaldehyde (3-HPA), which can be further metabolized to 1,3-propanediol and 3-hydroxypropionic acid (3-HP). While reuterin has been the focus of many investigations, the contribution of 3-HP to the antimicrobial activity of food related reuterin-producers is unknown. We show that the antibacterial activity of 3-HP was stronger at pH 4.8 compared to pH 5.5 and 6.6. Gram-positive bacteria were in general more resistant against 3-HP and propionic acid than Gram-negative indicator strains including common food pathogens, while spoilage yeast and molds were not inhibited by ≤ 640 mM 3-HP. The presence of acrolein decreased the minimal inhibitory activity of 3-HP against E. coli indicating synergistic antibacterial activity. 3-HP was formed during the growth of the reuterin-producers, and by resting cells of L. reuteri DSM 20016. Taken together, this study shows that food-related reuterin producers strains synthesize a second antibacterial compound, which might be of relevance when strains are added as starter or bioprotective cultures to food products.

Incidence and characterisation of Bacillus cereus bacteriophages from Thua Nao, a Thai fermented soybean product.

Bacillus cereus is considered to be an important food poisoning agent causing diarrhea and vomiting. In this study, the occurrence of B. cereus bacteriophages in Thai fermented soybean products (Thua Nao) was studied using five B. cereus sensu lato indicator strains (four B. cereus strains and one B. thuringiensis strain). In a total of 26 Thua Nao samples, there were only two bacteriophages namely BaceFT01 and BaceCM02 exhibiting lytic activity against B. cereus. Morphological analysis revealed that these two bacteriophages belonged to the Myoviridae. Both phages were specific to B. cereus and not able to lyse other tested bacteria including B. licheniformis and B. subtilis. The two phages were able to survive in a pH range between 5 and 12. However, both phages were inactive either by treatment of 50°C for 2 h or exposure of UV for 2 h. It should be noted that both phages were chloroform-insensitive, however. This is the first report describing the presence of bacteriophages in Thua Nao products. The characterization of these two phages is expected to be useful in the food industry for an alternative strategy including the potential use of the phages as a biocontrol candidate against foodborne pathogenic bacteria.

Antiadhesive, antibiofilm and dispersion activity of biosurfactants isolated from Bacillus amyloliquefaciens 3/22.

The aim of this work was to monitor the potential antibiofilm properties of biosurfactants (BS) isolated from Bacillus amyloliquefaciens 3/22 against biofilm formation of the indicator strain Staphylococcus aureus CCM 4223. In this work, the effect of BS 3/22 on biofilm growth during co-incubation, inhibition of biofilm-forming cell adhesion and biofilm dispersion was studied. BS 3/22 inhibited biofilm formation, with its formation decreasing significantly (p < 0.05; p < 0.01; p < 0.001) with increasing BS 3/22 concentration. BS 3/22 also showed antiadhesive activity, which correlated with the concentration used. The dispersing effect of isolated BS 3/22 on a 24-hour biofilm was also detected. BS 3/22 were effective in biofilm dispersion even at lower concentrations compared to antiadhesive activity and inhibition of biofilm formation.

The preliminary optimization of culture components for improving antifungal production by a wild-type soil bacteria Bacillus subtilis

Antifungal compounds are produced by Bacillus species under various growth conditions. While optimizing the antifungal production by using the one-factor-at-a-time (OFAT) and response surface methodology (RSM) approaches mannose and malt extract were identified as the most suitable carbon and nitrogen sources respectively. The RSM was applied to determine the optimum conditions of the three parameters such as pH, carbon and nitrogen sources for improved production. Optimum concentrations of carbon and nitrogen sources were 0.3% and 0.6% respectively with optimum media pH of 7.0 which showed optimum value of 40 AU/ml of antifungal compound against the Candida albicans SC5314 used as an indicator strain. In the present study, the F-value was determined as 0.0034 to imply that the model is significant. The goodness of the fit of the model was tested using coefficient of determination, R2 value, that tantamounts to 0.8562. The identification of antifungal compounds with their molecular masses was accomplished by MALDI-TOF mass spectrometry after n- butanol extraction. The present study thus has provided a platform to upgrade the yield of antifungal compounds which have got immense clinical potential to tread Candidosis.

A new xanthostatin analogue from the marine sponge-associated actinomycete Streptomyces sp. SCSIO 40064

A new cyclo-heptadepsipeptide xanthostatin B (1), together with isobutyryl hexapeptide (2), xanthostatin (3), TXS-1 (4) and TXS-2 (5), were isolated from the marine sponge derived Streptomyces sp. SCSIO 40064. The structures were elucidated by comprehensive spectroscopic data analyses and comparison with the literatures. The D-Val unit in 1 was assigned by Marfey’s method. The absolute configuration of 4 was determined by X-ray crystallographic analysis. Compounds 1‒5 were evaluated for the inhibitory activities against four pharmaceutical targets and six antibacterial indicator strains. Compound 5 displayed α-glucosidase inhibitory activity with IC50 value of 18.67 ± 1.27 µM.

Effects of biochar on bacterial genetic diversity in soil contaminated with cadmium

AbstractBiochar has various ecological effects on heavy metal‐contaminated soils. Biochar can be used to passivate the activity of heavy metals and improve environmental conditions for microbial growth. A greenhouse pot experiment was conducted to explore the diversity of microbes in red soil under cadmium (Cd) stress following the application of wheat straw biochar. The contaminant Cd was prepared at 2.5 mg kg‒1 with deionized water to simulate the heavy metal pollution of red soil under natural conditions, and the proportions of wheat straw biochar used were 2.5% and 10%. The complete genetic diversity of red soil bacteria in this study was evaluated using high‐throughput sequencing. The results showed that the bacterial genetic diversity of Cd‐contaminated red soils was restored with biochar treatment, and recovery with 2.5% biochar was significant. Biochar significantly affected the richness of soil bacteria by 6.79%–21.04%. Forty‐three phyla of bacteria, including Proteobacteria, Acidobacteria and Gemmatimonadetes, constituted the bacterial community in the red soils. Further principal component analysis showed that Cd pollution and biochar application collectively affected the bacterial genetic diversity. Hydrogenophaga, Rubrivivax, Haliscomenobacter, Citrobacter, Methylibium and Azospirillum were indicator strains for Cd‐contaminated red soils, while Steroidobacter, Bradyrhizobium, Anaerolinea, Chloronema Dubinina and Gorlenko were key strains for the biochar remediation of Cd contamination. In conclusion, for soil polluted with 2.5 mg kg Cd‒1, the application of 2.5% wheat straw biochar significantly increases soil microbial abundance and genetic diversity and plays an active role in improving the soil micro‐ecosystem.

천연물 유래 생리활성 소재의 개발

With the recent increase in health concerns, consumer needs for new health function materials have increased. Accordingly, physiological activity such as antibacterial and antioxidant effects of black garlic, black bellflower root, black deodeok, black ginseng, black leek, and pine powder, which have recently increased interest as a health functional material, was reviewed in this study. To examine the antibacterial activity of hot water extracts of each material, E. coli KCTC 2571, E. coli KCTC 1039, S. aureus KCTC 3881, S. aureus KCCM 1135, B. cereus KCCM 11204, L. monocytogenes KCCM 40307, P. aeruginoss KCTC 2513 were examined. The hot water extracts of black onion and black deodeok showed the growth inhibition of all the indicator strains except B. cereus KCCM 11204 and L. monocytogenes KCCM 40307. To examine the anti-oxidation activity of each material, the free radical scavenging test was used. The anti-oxidation activity of hot water extraction of each material was generally superior to that of 70% ethanol extract. And the hot water extracts of pine powder and the black bellflower root showed the excellent anti-oxidation activities, and are expected to be expected as new antioxidant materials.

Optimization of Cultivation Conditions for the Lactobacillus reuteri LR1 Strain to Improve the Biosynthesis of Bacteriocin-Like Substances

The article presents the results of optimization of the culturing conditions to increase the production of bacteriocin-like compounds (presumably including reuterin) by the Lactobacillus reuteri strain LR1. The strain Escherichia coli ATCC 25922, an opportunistic microorganism, the content of which is standardized in foods by the Food Safety Legislation, was selected as an indicator strain of antimicrobial activity. The nutrient medium for L. reuteri LR1 cultivation was optimized to increase the production of bacteriocin-like compounds, and the parameters of two-stage growth of the producer were established: (1) cultivation on the initial medium for biomass accumulation and (2) growth on a water–glycerol medium to convert glycerol to antimicrobial compounds, presumably reuterin. The first stage was carried out as follows: the L. reuteri LR1 culture (5%) was inoculated into a medium with the following composition: hydrolyzed milk—250 mL/L; yeast extract—10 g/L; peptone—5 g/L; glucose—20 g/L; sodium acetate—5.0 g/L; monosubstituted potassium phosphate—2.0 g/L; magnesium sulfate—0.2 g/L, and manganese sulfate—0.5 g/L, pH 6.4–6.6. The cultivation was carried out for 18 h at (37 ± 1)°С. The second stage included the incubation of the obtained biomass in 200 mM of glycerol-containing aqueous solution, pH 6.6, for 2 h at 37°С. The zone of E. coli inhibition caused by the produced bacteriocin-like compounds was 25 mm.

LACTOCOCCUS LACTIS STRAINS FROM INTESTINAL ORGAN OF BLACK TIPS SHARK CARCHARHINUS LIMBATUS PRODUCING NISIN-LIKE BACTERIOCIN ACTIVE AGAINST SHRIMP AND FISH PATHOGENS (VIBRIO PARAHAEMOLYTICUS AND VIBRIO ALGINOLYTICUS)

Shark can be a potential source for host-associated probiotics destined to be used in aquaculture. In this study, intestinal tissues of Black tip shark (Carcharhinus limbatus) was used as the source sample for inhibition screening based on spot-on-lawn and agar well diffusion methods. Out of the 80 isolates, only four showed antagonistic activities against selected indicator strains. All of the 4 isolates FA1, FA2, FA3 and FA4 were found to be Gram positive coccus, non-spore former, oxidase and catalase negatives, as well as lactose fermenters. The isolate FA1, FA2, FA3 and FA4 were found to demonstrate broad range of inhibitory spectrum on gram-positive indicator bacterium (Staphylococcus aureus and Bacillus cereus) and the gram-negative bacterium (Escherichia coli, Pseudomonas aeruginosa, Salmonella typhimurium, Vibrio alginolyticus and Vibrio parahaemolyticus); with the highest inhibition zone at 20.0±0.1 mm, recorded on V. parahaemolyticus. The genotype of the isolates was characterized using 16S rRNA sequencing. Each sequence with a given GenBank (NCBI) accession number (MN975529 for FA1; MN982712 for FA2; MN982711 for FA3; and MN982710 for FA4) showed at least 99% similarity with Lactococcus lactis subspecies. The crude cell free supernatant (CFS) of the isolates potentially contained a putative bacteriocin displaying nisin-like properties. The pH and catalase treatments showed that neither organic acid nor hydrogen peroxide (H2O2) was the inhibitory component. An almost 95% inactivation of the CFS’s antimicrobial activity was observed following treatment with trypsin suggested a presence of proteinaceous agent. Growth and inhibition studies on all isolated strains demonstrated that the inhibition properties were growth associated, with maximum inhibition achieved at ~20.0 ± 0.1 mm (or ~168 ± 17 AU/ml) at 17 hours by FA2 strain against V. parahaemolyticus. A study on the mode of inhibition demonstrated a bactericidal killing against V. parahaemolyticus. The isolation of Lactococcus lactis strain from shark is rather unique since it was commonly reported to be isolated from human and animals. These strains show bacteriocinogenic properties, broad spectrum inhibition, and more importantly, they are able to antagonise some pathogens implicated in fish or shrimp diseases. Therefore, these strains have potential for use as probiotics in aquaculture.

Enterococcus mundtii Isolated from Slovak Raw Goat Milk and Its Bacteriocinogenic Potential.

Enterococci are lactic acid bacteria. Most of them can adapt well to the food system due to their salt and acid-tolerance. Moreover, many enterococcal species have been found to produce antimicrobial substances of proteinaceous character, i.e., bacteriocins/enterocins. In this study, Enterococcus mundtii EM ML2/2 with bacteriocinogenic potential was identified in Slovak raw goat milk. This strain demonstrated inhibition activity against up to 36% of Gram-positive indicator bacteria, and in concentrated form the bacteriocin substance (pH 6.3) showed the highest inhibition activity (1600 AU/mL) against the principal indicator strain E. avium EA5. Semi-purified substance (SPS) EM ML2/2 produced inhibition activity up to 3200 AU/mL. Concentrated bacteriocin substance and SPS maintained active (inhibition activity up to 100 AU/mL) for three months under −20 °C storage conditions. The strain showed susceptible antibiotic profile, and it did not form biofilm. No production of damaging enzymes was noted. It was nonhemolytic, as well as DNase, and gelatinase-negative. It grew well in skim milk, and it was salt and acid-tolerant. The bacteriocin potential of E. mundtii species isolated from Slovak raw goat milk has not previously been detected, so this is an original contribution which may stimulate addtitional research and application studies.

Functional Properties and Sustainability Improvement of Sourdough Bread by Lactic Acid Bacteria.

Preventing food spoilage without the addition of chemical food additives, while increasing functional properties of wheat-based bakery products, is an increasing demand by the consumers and a challenge for the food industry. Within this study, lactic acid bacteria (LAB) isolated from sourdough were screened in vitro for the ability to utilize the typical wheat carbohydrates, for their antimicrobial and functional properties. The dual culture overlay assay revealed varying levels of inhibition against the examined fungi, with Lactiplantibacillus plantarum S4.2 and Lentilactobacillus parabuchneri S2.9 exhibiting the highest suppression against the indicator strains Fusarium graminearum MUCL43764, Aspergillus fumigatus, A. flavus MUCL11945, A. brasiliensis DSM1988, and Penicillium roqueforti DSM1079. Furthermore, the antifungal activity was shown to be attributed mainly to the activity of acids produced by LAB. The antibacillus activity was evaluated by the spot-on-the-lawn method revealing a high inhibition potential of the majority of LAB isolated from sourdough against Bacillus cereus DSM31, B. licheniformis DSM13, B. subtilis LMG7135, and B. subtilis S15.20. Furthermore, evaluating the presence of the glutamate decarboxylase gen in LAB isolates by means of PCR showed a strain dependency of a potential GABA production. Finally, due to improved functional activities, LAB isolated from sourdoughs exhibit promising characteristics for the application as natural preservatives in wheat-based bakery products.

Silver Antibacterial Synergism Activities with Eight Other Metal(loid)-Based Antimicrobials against Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus.

The present study surveys potential antibacterial synergism effects of silver nitrate with eight other metal or metalloid-based antimicrobials (MBAs), including silver nitrate, copper (II) sulfate, gallium (III) nitrate, nickel sulfate, hydrogen tetrachloroaurate (III) trihydrate (gold), aluminum sulfate, sodium selenite, potassium tellurite, and zinc sulfate. Bacteriostatic and bactericidal susceptibility testing explored antibacterial synergism potency of 5760 combinations of MBAs against three bacteria (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus) in three different media. Silver nitrate in combination with potassium tellurite, zinc sulfate, and tetrachloroaurate trihydrate had remarkable bactericidal and bacteriostatic synergism effects. Synergism properties of MBAs decreased effective antibacterial concentrations remarkably and bacterial cell count decreased by 8.72 log10 colony-forming units (CFU)/mL in E. coli, 9.8 log10 CFU/mL in S. aureus, and 12.3 log10 CFU/mL in P. aeruginosa, compared to each MBA alone. Furthermore, most of the MBA combinations inhibited the recovery of bacteria; for instance, the combination of silver nitrate–tetrachloroaurate against P. aeruginosa inhibited the recovery of bacteria, while three-fold higher concentration of silver nitrate and two-fold higher concentration of tetrachloroaurate were required for inhibition of recovery when used individually. Overall, higher synergism was typically obtained in simulated wound fluid (SWF) rather than laboratory media. Unexpectedly, the combination of A silver nitrate–potassium tellurite had antagonistic bacteriostatic effects in Luria broth (LB) media for all three strains, while the combination of silver nitrate–potassium tellurite had the highest bacteriostatic and bactericidal synergism in SWF. Here, we identify the most effective antibacterial MBAs formulated against each of the Gram-positive and Gram-negative pathogen indicator strains.

Antibacterial Bioactivity of Some Lactic Acid Bacteria Isolated from Various Egyptian Products

The current research aimed to isolate and screening of antibacterial bioactive lactic acid bacteria (LAB) from different foods such as raw milk, dairy products and fermented meat and evaluate their potential for production of antibacterial substances, namely bacteriocins. Ten samples were collected from different market foods, examined and used for LAB isolation on MRS medium. A total of 71 bacterial isolates were selected based on some variation properties, such as color, shape, size and oxygen requirements. These isolates were confirmed as lactic acid bacteria based on their microscopic observation and chemical properties, i.e. catalase negative and acid producing from glucose fermentation. All selected isolates were grown on MRS medium at 37oC for 48 h, and compared to identified LAB strains, i.e. Lactococcus lactis, Streptococcus thermophilus, Befidobacterium sp, Lactobacillus casi, Lb rhamnosus, Lb. acidophilus, Lb. helveticus and Lb. plantarum. Isolates and strains were surveyed for bacteriocin production as inhibitory activity test. Partial purification of bacteriocin was carried out by salt precipitation method; this method was done by ammonium sulphate 40-60% saturation and centrifuged at 9500 rpm for 20 minutes at 4oC. Some of microbial indicator strains found to test by inhibitory biological activity substance (partial purification of bacteriocin) produced by lactic acid cultures. Indicator strains such as staphylococcus aureus, Listeria monocytogenes and others were grown by nutrient agar at 37o C for 24h. An inhibitory activity test was executed by paper diffusion method. The inhibition effect was examined by form clear inhibition zone around indicator strains. Twenty-eight isolates and the 8 LAB strains showed bioactivities against the tested pathogens. Gram positive indicator bacteria were found to be the most sensitive to inhibitory substances produced by LAB culture as compared to gram negative indicator strains. Staphylococcus aureus was exhibited the highest sensitivity to it. The highest bacteriocins activities were produced by strain Lactobacillus rhamnosus while R5 and S6 from LAB isolates. They were shown broad spectrum against gram positive and gram-negative pathogenic bacteria. Diameters of inhibition zone refer to degree of sensitive between indicator strains and bacteriocin activity

Antibacterial mechanisms of bacteriocin BM1157 against Escherichia coli and Cronobacter sakazakii

E. coli and C. sakazakii are both opportunistic pathogenic bacteria that widely found in food. Emerging of antibiotic resistance makes the treatment of Gram-negative bacteria infection become ever more difficult. Therefore, it is urgent to find new antibacterial compounds against Gram-negative bacteria. Our previous study showed that bacteriocin BM1157 had broad antibacterial activity against both Gram-positive and Gram-negative bacteria. However, its antimicrobial mechanism against Gram-negative bacteria is still unknown. In this study, propidium iodide uptake and lactate dehydrogenase release experiments showed that the membrane integrity and permeability of the two indicator strains were not affected by BM1157 treatment. Observation of the ultrastructure of both bacteria by SEM and TEM also indicated that BM1157 did not damage the cell membrane. However, it significantly inhibited the biofilm formation of both bacteria. Furthermore, the lable-free proteome showed that BM1157 disturbed the carbohydrate metabolism, amino acid metabolism, lipid metabolism and nucleic acid metabolism, and then bioactive molecule synthesis and intracellular metabolism were blocked, resulting in cell death. Down-regulation of gene expression was further confirmed by real-time quantitative PCR. This study indicates that bacteriocin BM1157 inhibited Gram-negative bacteria by non-pore mechanisms.

PURIFICATION AND CHARACTERIZATION OF BACTERIOCIN FROM Lactobacillus acidophilus HT1 AND ITS APPLICATION IN A CREAM FORMULA FOR THE TREATMENT OF SOME SKIN PATHOGES

This study was aimed to purified and characterized the bacteriocin produced from Lactobacillus acidophilus HT1, in order to use it in a skin pharmaceutical formula. The optimal conditions for bacteriocin production was investigated and results showed that modified nutrient broth was the best medium with glucose (30 gm/L) and yeast extract (7 gm/L) with peptone (7 gm/L) were the optimum carbon and nitrogen sources. In addition, 2% inoculum size, 37C◦ and pH 6.4 were the optimal conditions to obtain maximum bacteriocin of 640 AU/ml after 24 hrs. The bacteriocin was purified using 70% ammonium salt saturation and gel filtration with sephadex G50 that resulted 20 % yield and 2560 AU/ml of activity, then the partial purified bacteriocin was characterized and found the bacteriocin was protein in nature and kept its activity after 10 min at 20, 30 and 40◦C, however 50% of the activity was lost at 50C◦. Moreover, it showed stability at pH 6 and 7 for 30 min whereas; no activity was observed at pH 4 and 9. In addition, results showed that bacteriocin has a bactericidal effect rather than bacteriostatic. A cream formula contained the bacteriocin was prepared which already examined in vitro and in vivo. The effectiveness of the formula was confirmed using Klebsiella sp., Staphylococcus aureus and Pseudomonas aeruginosa as indicator strains. Results established that treatment at the onset time was more effective and the time of healing was decreased.

Screening of Lactobacillus plantarum with broad-spectrum antifungal activity and its application in preservation of golden-red apples

Fungal food spoilage is a common problem that leads to both great economic losses and serious health problems. This study screened the antifungal activity of 137 Lactobacillus plantarum isolates against six common food spoilage indicator fungi using an overlay method and indicator strains of the species Aspergillus flavus, Fusarium moniliforme, Penicillium expansum, Penicillium chrysogenum, Cladosporium cladosporioides, and Aspergillus niger. Among Lactobacillus plantarum isolates, strain IMAU80174 was selected as the most effective based on the results of mycelium growth inhibition by its cell-free supernatant (CFS) and tolerance to simulated gastrointestinal juices and bile. The CFS of Lactobacillus plantarum IMAU80174 showed heat and protease resistance, and it was active only in a low pH environment. The application of the CFS to golden-red apples could slow down spoilage caused by inoculation of Penicillium expansum.

Reduction of the Carbapenemase Inactivation Method (CIM) assay time by real-time PCR

Carbapenemase Inactivation Method (CIM) is a test to detect presence of the carbapenemase in Gram-negative bacteria. Determination of the carbapenemase production by inactivation of meropenem requires that a zone of control E. coli inhibition be measured approximately 6–24 h after plating. We have modified the CIM test by developing a rapid method which instead measures the growth of E. coli indicator strain ATCC 25922 using real-time PCR, referred to as a nucleic acid testing CIM (natCIM). Our natCIM, therefore reduces the detecting time from 6 to 24 h to approximately 4 h.

New indicator Escherichia coli strain for rapid and accurate detection of supF mutations

BackgroundThe supF gene of Escherichia coli is useful for forward mutation analysis in bacterial and mammalian cells used in mutagenesis and DNA repair studies. Indicator E. coli strains, such as KS40/pOF105, have been used to analyze supF mutations. However, KS40/pOF105 is not enough to select supF mutants on nutrient-rich agar plates. Therefore, in this study, a new indicator E. coli strain for rapid and accurate detection of supF mutations was developed.ResultsThe gyrA and rpsL genes with an amber mutation were integrated into the chromosomal DNA of E. coli KS40 to produce a new indicator strain, RF01. RF01 cells transformed by the wild-type supF gene were sensitive to nalidixic acid and streptomycin on LB agar plates. supF mutant frequencies and mutation spectra in RF01 were similar to those in KS40/pOF105. In addition, some mutations in supF were only detected in RF01.ConclusionRF01 is a new and useful indicator E. coli strain for analyzing supF mutations.

Identification and characterization of a halocin-producing haloarchaeon isolated from Pachpadra salt lake.

Haloarchaea are known to produce antimicrobial proteins, halocins which are generally stable at extreme conditions suggesting their potential biotechnological applications. Here, we report a halocin-producing haloarchaeon isolated from salt lake and identified as Haloferax larsenii HA4 using partial 16S rDNA sequence and biochemical properties. Whole-cell methanolysate showed ether-linked lipids, which is a characteristic feature of haloarchaea. Strain HA4 was able to grow at pH 6·0-10·0 and 15-30% NaCl. The growth response was normal but antimicrobial activity was detected only during the log-phase. Crude halocin HA4 was active in the pH range of pH 2·0-10·0 with stability up to 100°C. Cell-free supernatant (CFS) was also stable in different organic solvents and detergents tested. However, halocin activity was reduced after treatment with proteinase K suggesting the proteinaceous nature of the active compound. Concentrated CFS showed the presence of several proteins from 6·5-66kDa but bioassay suggested ~14kDa protein as halocin. Crude halocin preparation showed cytocidal activity against indicator strain, H. larsenii HA10 and inhibited the growth of other related strains such as H. larsenii HA3, HA8, HA9 and HA10.