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.

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.

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.

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.

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.

Actividad antimicrobiana de Weissella confusa y sus metabolitos frente a Escherichia coli y Klebsiella pneumoniae / Antimicrobial activity of Weisella confusa and its metabolites against Escherichia coli and Klebsiella pneumoniae

Con el fin evaluar el campo de aplicación potencial de una bacteria ácido láctica y de sus metabolitos, se realizó la cinética de la actividad antimicrobiana de W. confusa y de sus metabolitos contra E. coli, y K. pneumoniae, dos patógenos causantes de enfermedades transmitidas por alimentos. La producción de W. confusa se realizó por fermentación discontinua en sustrato comercial MRS. Se realizaron tres fermentaciones durante 6 horas, sin aireación, agitación continúa 33°C y 100 rpm. Cada hora de fermentación se separaron tres sustancias biológicas, W. confusa con sus metabolitos (W+W10b), células de W. confusa libres de metabolitos (W) y metabolito (W10b) y se midió la actividad antimicrobiana contra los patógenos E. coli, y K. pneumoniae. Se encontraron diferencias estadísticas significativas entre tratamientos y tiempo de fermentación. Para E. coli el tratamiento W presentó la mayor actividad antimicrobiana, la cual se obtuvo entre la cuarta y sexta hora de fermentación (2.45 cm de diámetro promedio de inhibición). Para K. pneumoniae, los tratamientos W y W+W10b presentaron actividad antimicrobiana entre la cuarta y quinta hora de fermentación, sin diferencia significativa entre ellos. W. confusa y el metabolito W10b demostraron poseer capacidad antimicrobiana contra E. coli y K. pneumoniae, lo cual sugiere que W. confusa y W10b podrían utilizarse como alternativa de bioconservación o bioprotección de alimentos frescos y procesados, para alimentación humana y animal; y podría convertirse en una alternativa al uso de antibióticos para enfermedades causadas por E. coli y K. pneumoniae.

The kinetic of antimicrobial activity of Weissella confusa and their metabolites against E. coli, and K. pneumoniae, (two pathogens causing foodborne illness) was evaluated, in order to know the possible use in food processing. W. confusa was produced by batch fermentation using MRS commercial substrate. Three fermentations, of 6 hours at 33 °C, without aeration, stirring continuously (100 rpm) were performed. Every hour of fermentation, three biological substances, W. confusa with their metabolites (W + W10b), W. confusa free cells metabolites (W), and metabolite (W10b) were separated, and subsequently the antimicrobial activity against pathogenic E. coli and K. pneumoniae was measured. Statistically significant differences between treatments and fermentation time were found. Treatment (W) against E. coli, showed the greatest antimicrobial activity, it was obtained between the fourth and sixth hours of fermentation (2.45 cm diameter average inhibition). In treatments W and W + W10b against K. pneumoniae, statistically significant differences between them were not found. The antimicrobial activity was shown between the fourth and fifth hour of fermentation. W. confusa and W10b have antimicrobial activity against E. coli and K. pneumoniae, suggesting that W and W10b could be used as an alternative to biopreservation or bioprotection of fresh and processed food for human and animal consumption, and could become an alternative to antibiotics used for diseases caused by E. coli and K. pneumoniae.

Guava Psidium guajava seed flour and dry Aspergillus niger mycelium as nitrogen sources for the production of biomass and antimicrobial compounds produced by Weissella confusa

Background: The fermentation substrate efficiency of glucose supplemented with guava seed flour (GGSF) or glucose supplemented with dry Aspergillus niger mycelium (GANM) was evaluated during the production of biomass and antimicrobial compounds by the lactic acid bacteria Weissella confusa. Results: The fermentation substrate efficiency was measured by comparing the biomass formation, substrate consumption, substrate conversion, antimicrobial activity and product yield. The antimicrobial activity was measured against a commercial Staphylococcus aureus strain. The results were compared against fermentations performed in a commercial substrate (CS), the MRS (Man-Rogosa-Sharpe) substrate. The fermentations were performed discontinuously for 4 hrs at 100 rpm and 32ºC. The biomass production exhibited a statistically significant difference (P ≤ 0.05) between treatments. The biomass production was 13.98% higher in the CS than in the GGSF and GANM substrates; however, there were no statistically significant differences for the specific growth rate. Conclusions: The GGSF and GANM substrates favored an antimicrobial activity against Staphylococcus aureus during the second and third hours of fermentation (inhibition diameter was 6.11% and 4.72%, respectively). The GGSF, GANM and CS substrates did not present statistically significant differences for the production of antimicrobial substances against Staphylococcus aureus. Therefore, GGSF and GANM can be considered as viable and economical alternative nitrogen sources for the production of the antimicrobial compounds formed by Weissella confusa in submerged fermentations.

Weissella confusa Bacteremia in an Immune-Competent Patient with Underlying Intramural Hematomas of the Aorta

No abstract available.

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.

Effects of fermentation substrates and conservation methods on the viability and antimicrobial activity of weissella confusa and its metabolites

Lactic acid bacteria produce metabolites with antagonistic activity against other bacteria. However, growth conditions and conservation methods may reduce the viability and antimicrobial activity of lactic acid bacteria. This study evaluated the effects of fermentation substrate, lyophilization (freeze-drying) and refrigeration on the viability and antimicrobial activity of Weissella confusa strain and its metabolites against pathogens responsible for bovine mastitis. W. confusa strain was grown in MRS broth and milk supplemented with yeast extract and glucose (MYEG). The collected fractions were preserved by lyophilization or under refrigeration at 4ºC. Every seven days, the viability of W. confusa strain and the stability of its metabolites were evaluated against Staphylococcus aureus and Streptococcus agalactiae by disc diffusion assays. In both fermentation substrates, the combination of lyophilized strain and metabolites retained antimicrobial activity against the two pathogens for 42 days. Also, W. confusa strain retained adequate viability and antimicrobial activity when grown in MYEG and stored under refrigeration conditions. It was concluded that MYEG and refrigeration are acceptable low cost options to preserve the viability of W. confusa for its potential commercial use in the prevention and treatment of bovine mastitis.

Comparison of Temperature and Additives Affecting the Stability of the Probiotic Weissella cibaria / 전남의대학술지

Daily use of probiotic chewing gum might have a beneficial effect on oral health, and it is important that the viability of the probiotics be maintained in this food product. In this study, we examined the stability of probiotic chewing gum containing Weissella cibaria. We evaluated the effects of various factors, including temperature and additives, on the survival of freeze-dried probiotic W. cibaria powder. No changes in viability were detected during storage at 4degrees C for 5 months, whereas the viability of bacteria stored at 20degrees C decreased. The stability of probiotic chewing gum decreased steadily during storage at 20degrees C for 4 weeks. The viability of the freeze-dried W. cibaria mixed with various additives, such as xylitol, sorbitol, menthol, sugar ester, magnesium stearate, and vitamin C, was determined over a 4-week storage period at 20degrees C. Most of the freeze-dried bacteria except for those mixed with menthol and vitamin C were generally stable during a 3-week storage period. Overall, our study showed that W. cibaria was more stable at 4degrees C than that at 20degrees C. In addition, menthol and vitamin C had a detrimental effect on the storage stability of W. cibaria. This is the first study to examine the effects of various chewing gum additives on the stability of W. cibaria. Further studies will be needed to improve the stability of probiotic bacteria for developing a novel probiotic W. cibaria gum.

Comparison of Temperature and Additives Affecting the Stability of the Probiotic Weissella cibaria / 전남의대학술지

Daily use of probiotic chewing gum might have a beneficial effect on oral health, and it is important that the viability of the probiotics be maintained in this food product. In this study, we examined the stability of probiotic chewing gum containing Weissella cibaria. We evaluated the effects of various factors, including temperature and additives, on the survival of freeze-dried probiotic W. cibaria powder. No changes in viability were detected during storage at 4degrees C for 5 months, whereas the viability of bacteria stored at 20degrees C decreased. The stability of probiotic chewing gum decreased steadily during storage at 20degrees C for 4 weeks. The viability of the freeze-dried W. cibaria mixed with various additives, such as xylitol, sorbitol, menthol, sugar ester, magnesium stearate, and vitamin C, was determined over a 4-week storage period at 20degrees C. Most of the freeze-dried bacteria except for those mixed with menthol and vitamin C were generally stable during a 3-week storage period. Overall, our study showed that W. cibaria was more stable at 4degrees C than that at 20degrees C. In addition, menthol and vitamin C had a detrimental effect on the storage stability of W. cibaria. This is the first study to examine the effects of various chewing gum additives on the stability of W. cibaria. Further studies will be needed to improve the stability of probiotic bacteria for developing a novel probiotic W. cibaria gum.

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.

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.