Antibacterial and Modulation Effects of Levilactobacillus brevis KU202399 Derived from Traditional Korean Fermented Foods, Baechu (Napa Cabbage) Kimchi Against Intestinal Pathogens

This study aims to determine the antibiofilm effect of cell-free supernatant (CFS) of Lactobacillus brevis strains against Streptococcus mutans strains. To study the antibiofilm mechanism against S. mutans strains, antibacterial effects, cell surface properties (auto-aggregation and cell surface hydrophobicity), exopolysaccharide (EPS) production, and morphological changes were examined. The antibiofilm effect of L. brevis KCCM 202399 CFS as morphological changes were evaluated by scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM), compared with the control treatment. Among the L. brevis strains, L. brevis KCCM 202399 showed the highest antibiofilm effect on S. mutans KCTC 5458. The antibacterial effect of L. brevis KCCM 202399 against S. mutans KCTC 5458 was investigated using the deferred method (16.00 mm). The minimum inhibitory concentration of L. brevis KCCM 202399 against S. mutans KCTC 5458 was 25.00%. Compared with the control treatment, L. brevis KCCM 202399 CFS inhibited the bacterial adhesion of S. mutans KCTC 5458 by decreasing auto-aggregation, cell surface hydrophobicity, and EPS production (45.91%, 40.51%, and 67.44%, respectively). L. brevis KCCM 202399 CFS inhibited and eradicated the S. mutans KCTC 5458 biofilm. Therefore, these results suggest that L. brevis KCCM 202399 CFS may be used to develop oral health in the probiotic industry.

Purpose: The purpose of this study is to analyze nutrient intake and frequently consumed foods according to the intake frequency of Baechu (Chinese cabbage) kimchi by Korean adults. Methods: Data from analysis of 7,122 subjects aged 19~64 years from 2012, 2013 KNHANES were obtained from health behavior interviews and the 24-hour dietary recall method. Results: With the daily intake frequency of Baechu kimchi, subjects with two times a day were the highest proportion among total subjects. As Baechu kimchi intake frequency of breakfast, lunch, and dinner increased, the rate of skipping meals decreased significantly (p < 0.05). In addition, all food intake and daily nutrient intake increased significantly with increasing frequency of daily Baechu kimchi intake (Crude p for trend < 0.05, Adjusted p for trend < 0.05). With the frequently consumed foods by daily Baechu kimchi intake frequency in 2012 and 2013, among 509 foods top ranked foods were white rice, Baechu kimchi, beer, distilled alcoholic drink, milk, and pork. Conclusion: The results of this study could be used in the future to promote consumption of kimchi in the Korean population.

Although rare, the ability to produce surface S-layer proteins is beneficially associated with particular Lactobacillus strains being investigated as probiotics. Therefore, this work aimed to study specific probiotic functionalities of selected Levilactobacillus brevis strains MB1, MB2, MB13 and MB20, isolated from human milk microbiota, and to assess the contribution of S-proteins. Firstly, Rapid Annotation using Subsystem Technology revealed that cell wall-related genes were abundant in analysed L. brevis genomes. Furthermore, the results demonstrated that S-proteins mediate aggregation capacity and competitive exclusion of selected pathogens by L. brevis strains. The improvement of Caco-2 epithelial monolayer barrier function was demonstrated by the increase in JAM-A and occludin expressions when L. brevis strains or S-proteins were added, with the effect being most pronounced after treatment with MB2 and S-proteins of MB1. L. brevis strains, especially MB20, exerted the potential to adhere to recombinant human ZG16. Strain MB2 and MB20-S-proteins improved the barrier function of HT29 epithelial monolayer, as evidenced by increased ZG16 expression. Analysed L. brevis strains and S-proteins differentially affected the protein expression of IL-1β, IL-6 and IL-8, and IL-10 cytokines. The most prominent effect was observed by S-proteins of MB20, since IL-1β production was decreased while IL-10 production was significantly increased.

Application of pharmaceutical waste sludge compost alters the antibiotic resistome in soil under the Chinese cabbage system

This study was performed to mine biogenic amine (BA)-degrading lactic acid bacteria (LAB) from kimchi and to investigate the effects of the LAB strains on BA reduction in Baechu kimchi fermentation. Among 1448 LAB strains isolated from various kimchi varieties, five strains capable of considerably degrading histamine and/or tyramine were selected through in vitro tests and identified as Levilactobacillus brevis PK08, Lactiplantibacillus pentosus PK05, Leuconostoc mesenteroides YM20, L. plantarum KD15, and Latilactobacillus sakei YM21. The selected strains were used to ferment five groups of Baechu kimchi, respectively. The LB group inoculated with L. brevis PK08 showed the highest reduction in tyramine content, 66.65% and 81.89%, compared to the control group and the positive control group, respectively. Other BA content was also considerably reduced, by 3.76–89.26% (five BAs) and 7.87–23.27% (four BAs), compared to the two control groups, respectively. The other inoculated groups showed similar or less BA reduction than the LB group. Meanwhile, a multicopper oxidase gene was detected in L. brevis PK08 when pursuing the BA degradation mechanism. Consequently, L. brevis PK08 could be applied to kimchi fermentation as a starter or protective culture to improve the BA-related safety of kimchi where prolific tyramine-producing LAB strains are present.

Kimchi is a Korean traditional fermented vegetable food and is produced through a series of processes, including brining of major vegetables, blending with various other ingredients and fermentation. There are two major groups of kimchi, with many kinds in each group. Baechu (Chinese cabbage) kimchi is the most popular product. Kimchi fermentation is initiated by various microorganisms naturally present in raw materials, and the fermentation is later dominated by lactic acid bacteria. Salt, sugar, temperature, oxygen and population of desirable microflora are major factors that influence the fermentation behavior and kimchi quality. Complex biochemical changes occur before, during, and after fermentation. The characteristics of kimchi differ depending on the raw materials used, brining process, fermentation and storage methods. Kimchi is a nutritional food, and it contains high level of vitamins (ascorbic acid, carotene, B-complex), minerals (calcium, iron, potassium) and dietary fiber. Kimchi has demonstrated many health benefits besides nutritional functions, such as antioxidant, antimutagenic, anticarcinogenic activities. Several biologically active components including carotene, capsaicin, chlorophylls, dietary fibers, phenolic compounds, ascorbic acid, allylsulfides and lactic acid are considered to be the active agents of the health benefits of kimchi.

In poultry production, antibiotics have been excessively used as growth promoters to support well-being and decrease mortality caused by pathogenic microorganisms. The overuse of antibiotics has led to the emergence of antibiotic-resistant bacteria and the presence of antibiotic residues in poultry products. To counteract this problem, probiotics could be used as adjuncts or as substitutes for preserving a diverse and balanced microflora to prevent the colonization and multiplication of pathogenic bacteria in the GI tract. This study aimed to isolate and characterize the potential probiotic properties of lactic acid bacteria from the small intestine of 23-week-old broiler breeders, with the goal of identifying potential probiotic candidates. Four phenotypically healthy broiler breeders were selected, and intestinal contents were aseptically collected and cultured on MRS agar. From the initial pool of 39 colonies, six isolates were identified based on Gram-positive and catalase-negative characteristics and further classified using 16S rRNA sequencing as Levilactobacillus brevis (n = 3), Pediococcus pentosaceus (n = 2), and Streptococcus salivarius (n = 1). These strains were further evaluated for probiotic properties such as transit resistance to simulated upper gastrointestinal conditions, antagonist activity, haemolytic activity, and cell surface properties such as autoaggregation, co-aggregation and hydrophobicity, in vitro. L. brevis NKFS8 showed good tolerance to pH 3, while P. pentosaceus NKSF10 exhibited good tolerance to pH 4 acidic conditions. All isolates demonstrated good survivability in bile salt concentration of 3% (w/v), with P. pentosaceus NKSF10 exhibiting the highest tolerance. The isolates showed a wide range of antagonistic activity against the test pathogens Pseudomonas aeruginosa (ATCC 27853), Salmonella typhimurium, Salmonella enterica (ATCC 13314), Staphylococcus aureus (ATCC 29213), and Listeria monocytogenes (ATCC 7644). Furthermore, these strains exhibited good auto-aggregation, co-aggregation, and hydrophobicity properties. In conclusion, lactic acid bacteria from the small intestine of broiler breeders present a valuable prospect for the development of effective probiotics. These probiotics can be utilized as a supplementary inclusion in poultry feed, obviating the need for antibiotics as growth promoters. Nevertheless, additional in vivo studies are required to closely monitor and assess the effects of probiotics on the gastrointestinal system of chickens.

The metabolic versatility of Levilactobacillus brevis, a heterofermentative lactic acid bacterium, could benefit environmentally compatible and low salt cucumber fermentation. The biodiversity of Lvb. brevis autochthonous to cucumber fermentation was studied using genotypic and phenotypic analyses to identify unique adjunct cultures. A group of 131 isolates autochthonous to industrial fermentations was screened using rep-PCR-(GTG)5 and a fermentation ability assay under varied combinations of salt (0 or 6%), initial pH (4.0 or 5.2), and temperature (15 or 30°C). No apparent similarities were observed among the seven and nine clusters in the genotypic and phenotypic dendrograms, respectively. A total of 14 isolates representing the observed biodiversity were subjected to comparative genome analysis. The autochthonous Lvb. brevis clustered apart from allochthonous isolates, as their genomes lack templates for citrate lyase, several putative hypothetical proteins, and some plasmid- and phage-associated proteins. Four and two representative autochthonous and allochthonous Lvb. brevis, respectively, were subjected to phenotype microarray analysis using an Omnilog. Growth of all Lvb. brevis strains was supported to various levels by glucose, fructose, gentiobiose, 1,2-propanediol, and propionic acid, whereas the allochthonous isolate ATCC14890 was unique in utilizing citric acid. All the Lvb. brevis genomes encode for 1,2-propanediol utilization microcompartments. This study identified a unique Lvb. brevis strain, autochthonous to cucumber, as a potential functional adjunct culture for commercial fermentation that is distinct in metabolic activities from allochthonous isolates of the same species.

An antimicrobial peptide produced by Bacillussubtilis KU25, KU43, and KU44 was isolated from traditional Korean fermented foods and characterized. It was named as subtilin KU25, KU43, and KU44. Subtilin KU25, KU43, and KU44 were sensitive against α-chymotrypsin, protease XIII, and various proteinase enzymes, respectively. B. subtilis KU43 was selected as the producer with the broadest antimicrobial spectrum. Subtilin KU43 was stable at a pH range of 3 to 9 for 4 h, and withstood exposure to temperatures of 50–90°C for 30 min. The mode of inhibition against Listeria monocytogenes ATCC 15313 involved a bactericidal effect by a reduction in the cell numbers and breakage of the indicator cell membranes. The molecular mass of subtilin KU43 was measured at approximately 3.5 kDa. These results demonstrate the development of novel strains from traditional Korean fermented foods, and illustrate the possibility that some of these strains might generate a natural preservative compound.

BackgroundOf the many neurotransmitters in humans, gamma-aminobutyric acid (GABA) shows potential for improving several mental health indications such as stress and anxiety. The microbiota-gut-brain axis is an important pathway for GABAergic effects, as microbially-secreted GABA within the gut can affect host mental health outcomes. Understanding the molecular characteristics of GABA production by microbes within the gut can offer insight to novel therapies for mental health.ResultsThree strains of Levilactobacillus brevis with syntenous glutamate decarboxylase (GAD) operons were evaluated for overall growth, glutamate utilization, and GABA production in typical synthetic growth media supplemented with monosodium glutamate (MSG). Levilactobacillus brevis Lbr-6108™ (Lbr-6108), formerly known as L. brevis DPC 6108, and Levilactobacillus brevis Lbr-35 ™ (Lbr-35) had similar growth profiles but differed significantly in GABA secretion and acid resistance. Lbr-6108 produced GABA early within the growth phase and produced significantly more GABA than Lbr-35 and the type strain Levilactobacillus brevis ATCC 14869 after the stationary phase. The global gene expression during GABA production at several timepoints was determined by RNA sequencing. The GAD operon, responsible for GABA production and secretion, activated in Lbr-6108 after only 6 h of fermentation and continued throughout the stationary phase. Furthermore, Lbr-6108 activated many different acid resistance mechanisms concurrently, which contribute to acid tolerance and energy production. In contrast, Lbr-35, which has a genetically similar GAD operon, including two copies of the GAD gene, showed no upregulation of the GAD operon, even when cultured with MSG.ConclusionsThis study is the first to evaluate whole transcriptome changes in Levilactobacillus brevis during GABA production in different growth phases. The concurrent expression of multiple acid-resistance mechanisms reveals niche-specific metabolic functionality between common human commensals and highlights the complex regulation of GABA metabolism in this important microbial species. Furthermore, the increased and rapid GABA production of Lbr-6108 highlights the strain’s potential as a therapeutic and the overall value of screening microbes for effector molecule output.

Kimchi is a Korean traditional vegetable fermented food with lactic acid bacteria (LAB). Fermented kimchi contains live LAB (probiotics), dietary fibers (prebiotics) and dead LAB and fermented metabolites (postbiotics), and it may have various health benefits. The taste and functionalities of kimchi are dependent on its main ingredients and subingredients, fermentation conditions, LAB in the fermentation, etc. There are many types of kimchi, but Baechu kimchi is the most popular and commonly used kimchi in Korea. Baechu kimchi is prepared by LAB fermentation of Baechu cabbage mixed with other subingredients such as radish, green onion, red pepper powder, garlic, ginger and fermented small sea fishes. Kimchi contains high levels of vitamins, minerals, dietary fibers and other functional components and fermented metabolites. Various studies have reported on kimchi and its antimutagenic and anticancer, antioxidative and antiaging, antiatherosclerotic, antidiabetic, antiobesity, and anti-inflammatory effects. Fermented kimchi contains high levels of LAB (108–9 CFU/g) and LAB also contribute functionalities to kimchi. We will discuss the process of manufacturing kimchi, fermentation of kimchi and related microorganisms. Though it briefly discuss on the general functionalities of kimchi, we will focus on the history and functions of the antiobesity, anticancer, and anti-inflammatory effects of kimchi in detail with better taste and preservation period. To increase the antiobesity and anticancer functions of kimchi we will introduce kimchi recipes, salt kinds, other added ingredients, etc. It was finally discussed on postbiotics in kimchi and their health benefits.

Gajami-sikhae is a traditional Korean fermented fish food made by naturally fermenting flatfish (Glyptocephalus stelleri) with other ingredients. This study was the first to investigate the diversity and dynamics of lactic acid bacteria in gajami-sikhae fermented at different temperatures using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A total of 4824 isolates were isolated from the fermented gajami-sikhae. These findings indicated that Latilactobacillus, Lactiplantibacillus, Levilactobacillus, Weissella, and Leuconostoc were the dominant genera during fermentation, while the dominant species were Latilactobacillus sakei, Lactiplantibacillus plantarum, Levilactobacillus brevis, Weissella koreensis, and Leuconostoc mesenteroides. At all temperatures, L. sakei was dominant at the early stage of gajami-sikhae fermentation, and it maintained dominance until the later stage of fermentation at low temperatures (5 °C and 10 °C). However, L. plantarum and L. brevis replaced it at higher temperatures (15 °C and 20 °C). The relative abundance of L. plantarum and L. brevis reached 100% at the later fermentation stage at 20 °C. These results suggest that the optimal fermentation temperatures for gajami-sikhae are low rather than high temperatures. This study could allow for the selection of an adjunct culture to control gajami-sikhae fermentation.

This study aimed to investigate the effects of Levilactobacillus brevis, Lacticaseibacillus paracasei, and Lacticaseibacillus rhamnosus strains isolated from Mihalic cheese, also known as “weeping cheese”, on fermentation kinetics, microbial viability, and textural and aromatic properties of the butter matrix. The effects of the isolates were determined on acidification kinetics (Vmax, Tvmax, pHvmax), viability proportion index (VPI), textural parameters (firmness, work of shear, stickiness, work of adhesion), and volatile aroma compounds (GC-MS) formation. This study found that the BLR sample containing Lacticaseibacillus rhamnosus maintained its limited viability under acidic stress conditions despite its high fermentation rate and low pHvmax values. The BLP sample containing Lacticaseibacillus paracasei exhibited high viability due to its low acidification rate and limited pH change. Determining the chemical classes to which the aroma compounds in the BLP sample belonged revealed a composition rich in fatty acids. The BLB sample containing Levilactobacillus brevis produced a high ΔpH value and an aroma profile rich in aldehyde compounds. Examination of the macro-structural properties of the butter samples revealed that the sample containing Lacticaseibacillus rhamnosus, similar to the control sample (BMC), was more compact and rigid during storage. In contrast, samples containing Lacticaseibacillus paracasei and Levilactobacillus brevis had a softer/spreadable texture. These findings demonstrate the potential of lactic acid bacteria isolates from the traditional Mihalic cheese microbiota as biological catalysts for the development/improvement of texture, aroma, and sensory quality in high-fat dairy products and for the industrial production of products modified to meet consumer preferences.

Validation and inhibition study for toxic expression of quinolone antibiotic resistance genes in agricultural soils of eastern China

Traditional Bulgarian fermented foods are prominent for their uniqueness of local ingredients, production methods, and endemic microbial species. The present research investigated the diversity and beneficial biological potential of lactic acid bacteria (LAB) isolated from various types of unique Bulgarian fermented foods. Species identification was performed via 16S rDNA sequencing. Biological activity was evaluated by determining antibacterial activity (via agar well diffusion assay), H2O2 production, spectrophotometrically determined auto- and co-aggregation, microbial adhesion to hydrocarbon, and biofilm formation. The biosafety of the isolated lactic acid bacteria was established based on hemolytic activity and phenotypic and genotypic antibiotic susceptibility. Forty-five strains were isolated from fermented foods (sauerkraut, fermented green tomatoes, fermented cucumbers, kefir, white cheese, and Izvara (curdled milk)). Five species were detected: Lactiplantibacillus plantarum, Levilactobacillus koreensis, Levilactobacillus brevis, Lactobacillus helveticus, and Levilactobacillus yonginensis. The most prominent species was L. plantarum, at 47%. For the first time, L. koreensis and L. yonginensis, isolated from unique Bulgarian fermented foods, are reported in this study. The antibacterial effect of the cell-free supernatants was evaluated. An antagonistic effect was observed against Escherichia coli (57%) and Salmonella enterica subsp. enterica serotype Enteritidis (19%) for several L. plantarum strains. Only one L. brevis (Sauerkraut, S15) strain showed activity against E. coli. The best autoaggregation ability at hour 4 was observed for L. koreensis (fermented cucumbers, FC4) (48%) and L. brevis S2 (44%). The highest percentage of co-aggregation with Candida albicans, at hou 4 in the experiments, was observed for strains L. koreensis (fermented green tomato, FGT1) (70%), L. plantarum strains S2 (54%), S13 (51%), and S6 (50%), while at hour 24 for strains L. koreensis FGT1 (95%), L. brevis (Kefir, K7) (89%), L. plantarum S2 (72%), and L. koreensis FC2 (70%). Seven of the isolated LAB strains showed hydrophobicity above 40%. Our results showed that the ability of biofilm formation is strain-dependent. No hemolytic activity was detected. The antibiotic resistance to 10 antibiotics from different groups was tested phenotypically and genotypically. No amplification products were observed in any strains, confirming that the isolates did not carry antibiotic-resistance genes. Traditional fermented Bulgarian foods can be considered functional foods and beneficial LAB sources.