In silico prediction and in vitro analysis of bacteriocin and probiotic properties of Weissella cibaria NM1 isolated from Asian sea bass

Aims@#This study aims to predict the presence of bacteriocin- and probiotic-associated genes in the genome of Weissella cibaria NM1 isolated from Asian sea bass using a machine learning-based NeuBI prediction approach, followed by the investigation of the crude bacteriocin antimicrobial and probiotic properties via in vitro analysis.@*Methodology and results@#This study utilized the machine learning-based NeuBI prediction approach with a homology search of highly conserved bacteriocin-associated genes present in the genome of W. cibaria NM1. This approach discovered a putative bacteriocin gene (WC_2064) and bacteriocin operon with complete immunity, transporter, regulator and modifier genes. Furthermore, the genome of W. cibaria NM1 was found to harboured specific probiotic*associated genes that would contribute to acid and bile tolerance, adhesion on thehost cell and exhibited cholesterolreducing ability. On top of that, the genome also shows the absence of virulence and antibiotic resistance genes, which signifies the safety of W. cibaria NM1 as a potential probiotic candidate. In vitro study has confirmed the antipseudomonal activity of crude bacteriocin NM1 with MIC of 62.5 mg/mL. Weissella cibaria NM1 can tolerate 0.3% (v/v) of bile salt condition and the transit through the simulated gastric (pH 3 and 4) and small intestinal (pH 8) tract.@*Conclusion, significance and impact of study@#Current findings suggested in silico approach can speed up the search for putative bacteriocin and probiotic-associated genes from the genome of W. cibaria NM1. Nevertheless, further verification through experimental works will be deemed essential.

Optimization of the production of Weissella confusa MBF8-1 lysate in plant-based media by using response surface methodology

Aims@#Bacterial lysate has been reported to possess many health-care-related benefits. This study aimed to determine the optimum conditions for producing Weissella confusa MBF8-1 lysate in two plant-based modified De Man, Rogosa, and Sharpe (MRS) media using the response surface methodology (RSM). In this study, we applied several condition factors and compared them to standard MRS media. @*Methodology and results@#Weissella confusa MBF8-1 was grown in two modified MRS media, which are MRS Vegitone and soy peptone modified-MRS. The optimized fermentation condition factors such as nitrogen sources (i.e., soy peptone, proteose peptone), dextrose concentrations, and fermentation time were measured, and the responses, such as bacteriocin-like inhibitory substance (BLIS) activity and lysate pH were observed. RSM results showed the diameter of BLIS activity-inhibition zone and pH decreases of the lysate produced in MRS Vegitone containing 1.50% dextrose, 0.75% proteose peptone for 11.75 h fermentation and in soy peptone modified-MRS containing 2.05% dextrose, 1.05% soy peptone for 7.53 h fermentation, i.e., 7.41 mm at 7.36, and 7.80 mm at 7.30, respectively. Whereas, lysate produced in standard MRS medium containing 2% dextrose, 1% peptone for 8 h fermentation showed 7.85 mm diameter of BLIS activity-inhibition zone at pH 7.26. W. confusa MBF8-1 lysate showed slightly lower pH, but higher BLIS activity when grown in standard MRS media compared to those of the two modified MRS media. @*Conclusion, significance and impact of study@#The data obtained provide the optimum condition of W. confusa MBF8-1 lysate production in plant-based media. The pH and BLIS activity possessed by W. confusa MBF8-1 lysate produced in soy peptone modified-MRS showed a more similar result as the standard one than the other modified one. Thus, the soy peptone modified-MRS is recommended as a plant-based alternative medium replacing standard MRS.

Encapsulación de Weissella paramesenteroides y su aplicación en residuos de pescado / Weissella paramesenteroides encapsulation and its application in the use of fish wastes

The goal of the present study was to evaluate the encapsulation of Weissella paramesenteroides, isolated from bee bread, as a technological tool for its use in biological fish silage. The pH decrease in fish silages using the bacteria encapsulated and in a non-encapsulated form was compared. W. paramesenteroides showed a good performance in the development of bioEnc ap su lati on; logical fish silage. The alginate encapsulation method showed an encapsulation efficacy of 85% and provides a reliable technological application.

El objetivo del presente estudio fue evaluar la encapsulación de Weissella paramesenteroides, aislada a partir del pan de polen, como herramienta tecnológica para su uso en la elaboración de ensilado biológico de pescado. Se comparó el descenso de pH para los ensilados utilizando la bacteria encapsulada y no encapsulada. W. paramesenteroides mostró un buen desempeño en el desarrollo de ensilado biológico de pescado. El método de encapsulación con alginato mostró una eficacia del 85% y puede ser utilizado para su aplicación tecnológica.

Four amino acids as serum biomarkers for anti-asthma effects in the ovalbumin-induced asthma mouse model treated with extract of Asparagus cochinchinensis / 한국실험동물학회지

The butanol extract of Asparagus cochinchinensis roots fermented with Weissella cibaria (BAW) effectively prevents inflammation and remodeling of airway in the ovalbumin (OVA)-induced asthma model. To characterize biomarkers that can predict the anti-asthmatic effects induced by BAW treatment, we measured the alteration of endogenous metabolites in the serum of OVA-induced asthma mice after administration of low concentration BAW (BAWLo, 250 mg/kg) and high concentration BAW (BAWHi, 500 mg/kg) using ¹H nuclear magnetic resonance (¹H-NMR) spectral data. The number of immune cells and serum concentration of IgE as well as thickness of the respiratory epithelium and infiltration of inflammatory cells in the airway significantly recovered in the OVA+BAW treated group as compared to the OVA+Vehicle treated group. In the metabolic profile analysis, the pattern recognition showed completely separate clustering of serum analysis parameters between the OVA+Vehicle and OVA+BAW treated groups. Of the total endogenous metabolites, 19 metabolites were upregulated or downregulated in the OVA+Vehicle treated group as compared to the Control treated group. However, only 4 amino acids (alanine, glycine, methionine and tryptophan) were significantly recovered after BAWLo and BAWHi treatment. This study provides the first results pertaining to metabolic changes in the asthma model mice treated with OVA+BAW. Additionally, these findings show that 4 metabolites can be used as one of biomarkers to predict the anti-asthmatic effects.

Effect of Natural Extracts on Oral Care Probiotics Weissella cibaria CMU and Periodontal Pathogens

The purpose of this study is to determine if natural extracts could be used as an additive in oral health food made with Weissella cibaria CMU (oraCMU). Natural extracts of green tea, mulberry leaf, licorice, and propolis, which are reported to have antimicrobial activities, were selected and used in this study. The minimum inhibitory concentrations (MIC) of extracts on periodontal pathogens such as Fusobacterium nucleatum and Porphyromonas gingivalis and their synergy effects with oraCMU by the fractional inhibitory concentrations methods were measured. From the results obtained, all the extracts showed no effect on the growth of oraCMU. Green tea extract showed the best antibacterial activity with MIC of 1.8 mg/ml against both F. nucleatum and P. gingivalis. In addition, green tea extract had a synergistic effect with oraCMU against F. nucleatum. Therefore, these results suggested that green tea extract is available as an additive in oral health food made with oraCMU.

Oral malodor-reducing effects by oral feeding of Weissella cibaria CMU in Beagle dogs / 대한수의학회지

This study assessed the effects of Weissella cibaria (W. cibaria) CMU on oral health in male and female beagles (n = 18) by measuring oral malodor and periodontal disease-related parameters (calculus, plaque, and gingivitis indices). Oral malodor and indicators of periodontal disease were assessed in five treatment groups: negative control (scaling and 0.24 mg of maltodextrin, n = 3), positive control (0.24 mg of maltodextrin, n = 3), and W. cibaria CMU groups (each n = 4) at low (CMU-L, 2 × 10⁷ colony forming unit [CFU]), medium (CMU-M, 2 × 10⁸ CFU), and high (CMU-H, 2 × 10⁹ CFU) concentrations. After feeding with W. cibaria CMU for 6 weeks, total volatile sulfur compound concentrations in the CMU-L (2.0 ± 1.04 ng/10 mL), CMU-M (2.4 ± 1.05 ng/10 mL), and CMU-H (2.6 ± 1.33 ng/10 mL)groups were significantly lower than in the positive control group (3.2 ± 1.65 ng/10 mL). Also, CMU-L (1.4 ± 0.83 ng/10 mL) and CMU-H (1.9 ± 1.14 ng/10 mL) groups had methyl mercaptan levels lower than that in the positive control group (2.4 ± 1.21 ng/10 mL) at week 2. The plaque index was significantly lower in the CMU-H group (4.5 ± 0.28) than in the positive control group (5.9 ± 1.08) at week 6. W. cibaria CMU could be useful as a novel oral hygiene probiotics for reducing volatile sulfur compounds production and inhibiting plaque growth in companion animals.

Dose dependence and durability of the therapeutic effects of Asparagus cochinchinensis fermented extract in an ovalbumin-challenged asthma model / 한국실험동물학회지

The butanol extract of Asparagus cochinchinensis roots fermented with Weissella cibaria (BAfW) significantly suppressed the inflammatory response induced by lipopolysaccharide (LPS) treatment in RAW264.7 cells. To investigate the dose dependence and durability of BAfW on the anti-asthma effects, alterations in key parameters were measured in ovalbumin (OVA)-challenged Balb/c mice treated with the different doses of BAfW at three different time points. The number of immune cells, OVA-specific IgE level, thickness of respiratory epithelium and mucus score decreased significantly in a dose-dependent manner in response to treatment with 125 to 500 mg/kg BAfW (P < 0.05), although the highest level was detected in the 500 mg/kg treated group. Moreover, the decrease in these parameters was maintained from 24 to 48 h in the 500 mg/kg of BAfW treated group. At 72 h, the effects of BAfW on the number of immune cells, OVA-specific IgE level and thickness of respiratory epithelium partially disappeared. Overall, this study provides the first evidence that the anti-asthma effect of BAfW may reach the maximum level in OVA-challenged Balb/c mice treated with 500 mg/kg and that these effects can last for 48 h.

Characteristics and immuno-modulatory effects of Weissella cibaria JW15 isolated from Kimchi, Korea traditional fermented food, for probiotic use / 동물의과학연구지

In this study, characteristics and immuno-modulatory effects of Weissella cibaria JW15 isolated from Kimchi, traditional Korean fermented food, were examined for investigation of the capacity of potentially probiotic strains. We measured acid, bile, and heat tolerance, adhesive properties to intestinal epithelial cells, and inhibitory activity against pathogens. JW15 could survive at pH 3.0 for 2 hr, but not at pH 2.0. JW15 also showed tolerance to 0.3% oxgall bile salt, and heat tolerance at 70degrees C and 80degrees C for 5 min, respectively. Adhesive ability to Caco-2 cells was similar to that of Lactobacillus rhamnosus GG (LGG), a well-known commercial probiotic. JW15 exhibited antimicrobial activities to pathogenic bacteria such as Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, and Salmonella enteritidis. The immuno-modulatory effects of JW15 were compared with those of LGG, a well-known immune enhancer. For analysis, production of nitric oxide (NO), NF-kappaB (Nuclear factor kappaB), Interleukin-1beta (IL-1beta), and tumor necrosis factor-alpha (TNF-alpha) was measured. The concentration of NO induced by JW15 was higher than that by LGG at low concentration (1 x 10(7) cfu/mL). Low and high (5 x 10(7) CFU/mL) concentration of JW15 induced statistically higher production of NF-kappaB, IL-1beta, and TNF-alpha than that produced by LGG, respectively. In conclusion, Weissella cibaria JW15 had ability as a probiotic strain, including acid, bile, and heat tolerance, adhesive properties to intestinal epithelial cells, and inhibitory activity against pathogens. In addition, JW15 showed better immuno-modulatory effects than LGG when NO, NF-kappaB, IL-1beta, and TNF-alpha were measured. According to these results, the characteristics and immunomodulating activity of Weissella cibaria JW15 are suitable for consideration as a potential probiotic.

Optimization of Fermentation Medium for Enhanced Glucansucrase and Glucan Production from Weissella confusa

A glucan producing Weissella confusa isolated from fermented cabbage, as identified earlier, was used for optimization of its fermentation medium. The effects of various macronutrients such as sucrose, glucose as cosubstrate, yeast extract, beef extract, peptone, sodium acetate, K2HPO4 and Tween 80 were studied on glucansucrase and glucan production from Weissella confusa. The medium used as control gave 6.0 U/ml enzyme activity and 34 mg/ml glucan concentration. Sucrose (5 percent), glucose as cosubstrate (5 percent for glucansucrase and 3 percent for glucan production), Tween 80 (0.1 percent), yeast extract (1.5 percent), Peptone (2.0 percent) and K2HPO4 (1.5 percent) were effective nutrients displaying higher glucansucrase and glucan production giving 18.2, 18.0, 7.0, 6.4, 6.2 and 6.4 U/ml enzyme activity and 103, 100, 46, 41, 39 and 37 mg/ml glucan concentration, respectively. Sodium acetate and beef extract were not effective for enzyme and glucan production. The new strain Weissella confusa can be used for commercial production of glucansucrase and glucan using optimized medium.

Effect of Weissella cibaria on Fusobacterium nucleatum-induced Interleukin-6 and Interleukin-8 Production in KB Cells

Oral microorganisms, including pathogens together with commensals, interact with oral epithelial cells, which can lead to the activation and expression of a variety of inflammatory mediators in epithelial cells. Fusobacterium nucleatum is a filamentous human pathogen that is strongly associated with periodontal diseases. Our previous data suggest that Weissella cibaria, an oral commensal, inhibits the proliferation of periodontopathic bacteria including F. nucleatum. The aim of this study was to examine the effects of W. cibaria on the inflammatory mediators, interleukin (IL)-6 and IL-8, in KB cells stimulated by F. nucleatum. In a reverse transcription-polymerase chain reaction and an enzyme-linked immunosorbent assay, live F. nucleatum alone induced high levels of gene expression and protein release of IL-6 and IL-8, whereas W. cibaria alone did not induce IL-6 and IL-8 responses in KB cells. W. cibaria dose-dependently inhibited the increases of the IL-6 and IL-8 gene expression as well as IL-6 protein level in KB cells which was induced by F. nucleatum. Bacterial viability and its coaggregation with F. nucleatum are not essential in the inhibitory effect of W. cibaria. Visible effects of W. cibaria on the attachment and invasion of KB cells by F. nucleatum were observed. In conclusion, W. cibaria may exert immunomodulatory effects on the IL-6 and IL-8 responses to F. nucleatum-activated KB cells.

Isolation and characteristics of lactic acid bacteria isolated from ripe mulberries in Taiwan

The objective of this study was to isolate, characterize, and identify lactic acid bacteria (LAB) from ripe mulberries collected in Taiwan. Ripe mulberry samples were collected at five mulberry farms, located in different counties of Taiwan. Eighty-eight acid-producing cultures were isolated from these samples, and isolates were divided into classes first by phenotype, then into groups by restriction fragment length polymorphism (RFLP) analysis and sequencing of 16S ribosomal DNA (rDNA). Phenotypic and biochemical characteristics led to identification of four bacterial groups (A to D). Weissella cibaria was the most abundant type of LAB distributed in four mulberry farms, and Lactobacillus plantarum was the most abundant LAB found in the remaining farm. Ten W. cibaria and one Lactococcus lactis subsp. lactis isolate produced bacteriocins against the indicator strain Lactobacillus sakei JCM 1157T. These results suggest that various LAB are distributed in ripe mulberries and W. cibaria was the most abundant LAB found in this study.

Quantitative Analysis of Weissella cibaria against Periodontopathic Bacteria by Real-time PCR

The objective of this study was to analyze quantitatively whether Weissella cibaria could affect the proliferation of five periodontopathic bacteria, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Aggregatibacter actinomycetemcomitans, and Fusobacterium nucleatum, after incubation for 8~48 h. In addition, by using real-time PCR with a dual-labeled probe, each growth of bacteria was examined under different growth media conditions. The proliferation of periodontopathic bacteria was significantly inhibited by W. cibaria after incubation for 24~48 h (p < 0.05), whereas the growth of W. cibaria was not affected by these pathogenic bacteria. The growth of P. gingivalis, T. forsythia and T. denticola significantly increased in each growth media after incubation for 24 h (p < 0.05), as compared to the culture in mixed growth media. However, no differences in the growth of five periodontopathic bacteria were observed between each growth media and mixed media after incubation for 48 h. The growth and pH of W. cibaria culture significantly were changed in MRS after incubation for 24~48 h (p < 0.05), as compared to the bacterial culture in mixed growth media. The pH of P. gingivalis and F. nucleatum culture significantly was changed in both growth media and mixed media after incubation for 24~48 h (p < 0.05). Our data indicate that W. cibaria significantly inhibits the proliferation of five periodontopathic bacteria and each growth of bacteria is quantitatively analyzed under various media conditions by real-time PCR.

Adhesion of Weissella cibaria to the Epithelial Cells and Factors Affecting its Adhesion

We evaluated the ability of lactic acid bacteria, Weissella cibaria, isolated from the oral cavity to adhere to epithelial cells. W. cibaria efficiently adhered to KB cells and HeLa cells. In addition, W. cibaria efficiently adhered to Fusobacterium nucleatum. But the adhesiveness of W. cibaria disappeared upon exposure to LiCl or pronase, suggesting that the S-layer proteins of W. cibaria mediated the adhesiveness. The molecular mass of the S-layer proteins extracted from W. cibaria was approximately 50 kDa. When W. cibaria strains were washed with 0.45% saline, the bacteria were efficiently adhered to the epithelial cells. In conclusion, W. cibaria has the ability to adhere to epithelial cells through the S-layer proteins.