Potential of Bifidobacterium lactis IDCC 4301 isolated from breast milk-fed infant feces as a probiotic and functional ingredient.

Probiotics provide important health benefits to the host by improving intestinal microbial balance and have been widely consumed as dietary supplements. In this study, we investigated whether Bifidobacterium lactis IDCC 4301 (BL), isolated from feces of breast milk-fed infants, is safe to consume. Based on the guidelines established by the European Food Safety Authority (EFSA), safety tests such as antibiotic susceptibility, hemolysis, toxic compound formation (i.e., biogenic amine and d-lactate), single-dose acute oral toxicity, and extracellular enzymatic activities were performed. In addition, toxigenic genes, antibiotic resistance genes, and mobile genetic elements were investigated by analyzing the genome sequence of BL. BL was susceptible to eight antibiotics except for vancomycin and the absence of transferable resistance in the genome of this strain implied that vancomycin resistance is likely to be intrinsic. With regard to phenotypic characteristics, there was no concern of toxicity of this strain. Furthermore, BL utilized various carbohydrates and their conjugates through the activity of various endogenous carbohydrate-utilizing enzymes. Interestingly, the supernatant of the BL showed strong antipathogenic activity against various infectious pathogens. Therefore, we suggest that BL should be a safe probiotic and can be used as a functional ingredient in the food, cosmetic, and pharmaceutical industries.

Bifidobacterium lactis IDCC 4301 Exerts Anti-Obesity Effects in High-Fat Diet-Fed Mice Model by Regulating Lipid Metabolism.

SCOPE: Chronic hypernutrition promotes lipid accumulation in the body and excessive lipid accumulation leads to obesity. An increase in the number and size of adipocytes, a characteristic of obesity is closely associated with adipose dysfunction. Recent in vitro and in vivo studies have shown that probiotics may prevent this dysfunction by regulating lipid metabolism. However, the mechanisms of action of probiotics in obesity are not fully understood and their usage for treating obesity remains limited. METHODS AND RESULTS: Bifidobacterium lactis IDCC 4301 is selected for its anti-obesity potential after evaluating inhibitory activity of pancreatic lipase and cholesterol reducing activity. Next, this study investigates the roles of B. lactis IDCC 4301 on lipid metabolism in 3T3-L1 preadipocytes and high-fat diet (HFD)-fed mice. B. lactis IDCC 4301 inhibits cell differentiation and lipid accumulation by suppressing the expression of adipogenic enzymes in 3T3-L1 cells. Moreover, the administration of B. lactis IDCC 4301 decreases body and adipose tissue weight, improves serum lipid levels, and downregulates adipogenic mRNA expression in HFD-fed mice. Additionally, metabolomic analysis suggests that 2-ketobutyrate should be a possible target compound against obesity. CONCLUSIONS: B. lactis IDCC 4301 may be used as an alternative treatment for obesity.

Isolation, Characterization, and Safety Evaluation of the Novel Probiotic Strain Lacticaseibacillus paracasei IDCC 3401 via Genomic and Phenotypic Approaches.

This study aimed to explore the safety and properties of Lacticaseibacillus paracasei IDCC 3401 as a novel probiotic strain via genomic and phenotypic analyses. In whole-genome sequencing, the genes associated with antibiotic resistance and virulence were not detected in this strain. The minimum inhibitory concentration test revealed that L. paracasei IDCC 3401 was susceptible to all the antibiotics tested, except for kanamycin. Furthermore, the strain did not produce toxigenic compounds, such as biogenic amines and D-lactate, nor did it exhibit significant toxicity in a single-dose acute oral toxicity test in rats. Phenotypic characterization of carbohydrate utilization and enzymatic activities indicated that L. paracasei IDCC 3401 can utilize various nutrients, allowing it to grow in deficient conditions and produce health-promoting metabolites. The presence of L. paracasei IDCC 3401 supernatants significantly inhibited the growth of enteric pathogens (p < 0.05). In addition, the adhesion ability of L. paracasei IDCC 3401 to intestinal epithelial cells was found to be as superior as that of Lacticaseibacillus rhamnosus GG. These results suggest that L. paracasei IDCC 3401 is safe for consumption and provides health benefits to the host.

Safety Evaluation of Bacillus subtilis IDCC1101, Newly Isolated from Cheonggukjang, for Industrial Applications.

The present study aimed to evaluate the safety of Bacillus subtilis (BS) IDCC1101, newly isolated from Cheonggukjang in Korea. Genome sequencing of BS IDCC1101 was performed to investigate the presence of secondary metabolites, virulence, antibiotic resistance, and mobile elements. Its phenotypic safety analyses included antibiotic susceptibility, enzyme activity, carbohydrate utilization, production of biogenic amines (BAs) and D-/L-lactate, hemolytic activity, and toxicities in HaCaT cells and rats. The genome of BS IDCC1101 consisted of 4,118,950 bp with 3077 functional genes. Among them, antimicrobial and antifungal secondary metabolites were found, such as fengycin, bacillibactin, and bacilysin. Antibiotic resistance and virulence genes did not exhibit transferability since they did not overlap with mobile elements in the genome. BS IDCC1101 was susceptible to almost all antibiotics suggested for assessment of BS's antibiotic susceptibility by EFSA guidelines, except for streptomycin. BS IDCC1101 showed the utilization of a wide range of 27 carbohydrates, as well as enzyme activities such as alkaline phosphatase, esterase, esterase lipase, naphthol-AS-BI-phosphohydrolase, α-galactosidase, ß-galactosidase, α-glucosidase, and ß-glucosidase activities. Additionally, BS IDCC1101 did not exhibit the production of D-/L-lactate and hemolytic activities. Its toxicity in HaCaT cells and rats was also not detected. Thus, these genotypic and phenotypic findings indicate that BS IDCC1101 can be safely used for industrial applications.

Safety Evaluation by Phenotypic and Genomic Characterization of Four Lactobacilli Strains with Probiotic Properties.

Probiotic Lactobacillus species are known to exert health benefits in hosts when administered in adequate quantities. A systematic safety assessment of the strains must be performed before the Lactobacillus strains can be designated as probiotics for human consumption. In this study, we selected Lactobacillus fermentum IDCC 3901, L. gasseri IDCC 3101, L. helveticus IDCC 3801, and L. salivarius IDCC 3551 as representative Lactobacilli probiotic strains and investigated their probiotic properties and potential risks through phenotypic and genomic characterization. Various assays including antimicrobial resistance, biogenic amine production, L-/D-lactate production, acute oral toxicity, and antipathogenic effect were performed to evaluate the safety of the four Lactobacillus strains. Genomic analysis using whole genome sequencing was performed to investigate virulence and antibiotic resistance genes in the genomes of the selected probiotic strains. The phenotypes of the strains such as enzymatic activity and carbohydrate utilization were also investigated. As a result, antibiotic resistances of the four Lactobacillus species were detected; however, neither antibiotic resistance-related genes nor virulence genes were found by genomic analysis. Moreover, the four Lactobacillus species did not exhibit hemolytic activity or ß-glucuronidase activity. The biogenic amine production and oral acute toxicity were not shown in the four Lactobacillus species, whereas they produced D-lactate with minor ratio. The four Lactobacillus species exhibited antipathogenic effect to five pathogenic microorganisms. This study provides a way to assess the potential risks of four different Lactobacillus species and validates the safety of all four strains as probiotics for human consumption.

Anti-inflammatory and anti-pathogenic potential of Lacticaseibacillus rhamnosus IDCC 3201 isolated from feces of breast-fed infants.

OBJECTIVE: We investigated the anti-inflammatory and anti-pathogenic activities of Lacticaseibacillus rhamnosus IDCC 3201 isolated from the feces of breast-fed infants. METHODS: Cell viability, nitric oxide (NO) production, and expression of inflammatory markers by L. rhamnosus IDCC 3201 were quantitatively analyzed in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. The antibacterial and antifungal activities of L. rhamnosus IDCC 3201 against various pathogens were also investigated. RESULTS: Treatment of LPS-induced macrophages with cell-free supernatant of L. rhamnosus IDCC 3201 significantly decreased the expression levels of tumor necrosis factor (TNF-α) and interleukin-6 (IL-6). Nitric oxide synthase (iNOS) and cyclooxygenase (COX-2) levels also significantly decreased in LPS-induced macrophages. Phenotypically, the treatment of L. rhamnosus IDCC 3201 reduced the production of nitric oxide (NO) in LPS-induced macrophages. Furthermore, L. rhamnosus IDCC 3201 was proven to have potent inhibitory activities against various pathogens responsible for inflammatory responses in the gastrointestinal tract (i.e., Bacillus cereus, Enterococcus faecalis, Staphylococcus aureus, and Salmonella Typhimurium), respiratory system (i.e., Streptococcus pneumoniae), and vagina (i.e., Candida albicans). CONCLUSION: L. rhamnosus IDCC 3201 has anti-inflammatory activity in terms of decreased expression of cytokines, inflammation-inducible enzymes in LPS-induced macrophages, and anti-pathogenic activity.

Safety evaluation and anti-inflammatory activity of Lactobacillus johnsonii IDCC 9203 isolated from feces of breast-fed infants.

This study evaluated the safety of Lactobacillus johnsonii IDCC 9203 and investigated its anti-inflammatory activity in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. Genomic analysis revealed that this strain has no virulence and antibiotic resistance gene except tetW, which is a tetracycline resistance gene. Minimum inhibitory concentration data showed that the strain is resistant to tetracycline and aminoglycosides. Further analysis indicated that the transferability of the tetW gene is extremely low, and resistance to aminoglycosides is due to the intrinsic resistance of L. johnsonii IDCC 9203. Phenotypic safety assessment showed that the strain has neither ß-hemolytic nor ß-glucuronidase activity, and no biogenic amine production. When LPS-induced RAW 264.7 cells were treated with L. johnsonii IDCC 9203, the level of nitric oxide and expression of pro-inflammatory cytokines significantly decreased (p < 0.05). Therefore, L. johnsonii IDCC 9203 strain is considered as safe and beneficial probiotic for human consumption.

A Quadruple Coating of Probiotics for Enhancing Intestinal Adhesion and Competitive Exclusion of Salmonella Typhimurium.

Previously, our group showed that a quadruple coating of probiotics resulted in higher survivability of probiotics under high acid, bile salt, and thermal stresses. In this study, we evaluated the effect of the quadruple coating of probiotics on adhesive properties as well as on competitive exclusion of Salmonella Typhimurium in Caco-2 cells. We found that the quadruple coating of probiotics exhibited an overall increased adhesion property (up to 10.8-fold) and increased competitive exclusion of Salmonella Typhimurium (up to 4.3-fold). Thus, this study has significant implications and can lead to the development of methods that can improve the adhesive ability of probiotics as well as the adhesive inhibition of pathogens.

Influence of cellulose nanocrystal addition on the production and characterization of bacterial nanocellulose.

Bacterial nanocellulose (BNC) is characterized by high purity and excellent mechanical properties; however, its production is constrained by low yield. Therefore, efforts aimed at improving its yield and material properties are imperative. This study investigated the effect of adding different concentrations (0%, 0.5%, and 1.0%) of cellulose nanocrystal (CNC) in Hestrin-Schramm modified medium on the yield and properties of BNC produced by Komagataeibacter sp. SFCB22-18. The BNC yield increased as following an increase in added CNC concentration. Also, the morphology, structure, crystallinity, thermal stability, and mechanical properties of BNC improved after CNC incorporation. A low CNC concentration (0.1%) favored mechanical strength, whereas 0.5% gave the optimum morphology, structural, and thermal stability. These results showed that modifying BNC with CNC could help increase yield and improve its properties, and thus; the potentiality of BNC in various applications would be much enhanced.

Anti-inflammatory potential of Lactiplantibacillus plantarum IDCC 3501 and its safety evaluation.

This study investigated the anti-inflammatory activity of Lactiplantibacillus plantarum IDCC 3501 isolated from kimchi (Korean fermented food) and its safety. When lipopolysaccharide (LPS)-induced RAW 264.7 macrophages were treated with cell-free supernatant from L. plantarum IDCC 3501, the mRNA expression level of inflammatory markers (i.e., TNF-α, IL-1ß, and IL-6) was significantly reduced. In addition, the decreased cell viability by LPS was recovered and NO production in LPS-induced cell was also decreased. For the safety assessment, the genes responsible for antibiotic resistance and virulence were not detected from the genome analysis of this strain. Consistent with this, minimal inhibitory concentrations against various antibiotics, biogenic amines, and D-lactate production, as well as enzymatic and hemolysis activities, indicated that L. plantarum IDCC 3501 did not produce any harmful compounds during fermentation. Furthermore, no acute toxicity and mortality were observed in a murine mouse model. Based on our findings, L. plantarum IDCC 3501 is safe and beneficial for human consumption.

Addition of Various Cellulosic Components to Bacterial Nanocellulose: A Comparison of Surface Qualities and Crystalline Properties.

Bacterial nanocellulose (BNC) is a biocompatible material with a lot of potential. To make BNC commercially feasible, improvements in its production and surface qualities must be made. Here, we investigated the in situ fermentation and generation of BNC by addition of different cellulosic substrates such as Avicel and carboxymethylcellulose (CMC) and using Komagataeibacter sp. SFCB22-18. The addition of cellulosic substrates improved BNC production by a maximum of about 5 times and slightly modified its structural properties. The morphological and structural properties of BNC were investigated by using Fourier transform-infrared spectroscopy (FT-IR), scanning electron microscopy and X-ray diffraction. Furthermore, a type-A cellulose-binding protein derived from Clostridium thermocellum, CtCBD3, was used in a novel biological analytic approach to measure the surface crystallinity of the BNC. Because Avicel and CMC may adhere to microfibrils during BNC synthesis or crystallization, cellulose-binding protein could be a useful tool for identifying the crystalline properties of BNC with high sensitivity.

Genomic-, phenotypic-, and toxicity-based safety assessment and probiotic potency of Bacillus coagulans IDCC 1201 isolated from green malt.

Probiotics are beneficial microorganisms, and the evaluation of their safety for human use in the food industry has become critical. This study examines the safety of Bacillus coagulans IDCC 1201 isolated from green malt by analyzing its genomic and phenotypic characteristics and determining its toxicity. The presence of antibiotic resistance and toxigenic genes and gene transferability were investigated using whole-genome analysis. The strain's hemolytic and enzyme activities, minimum inhibitory concentrations of antibiotics, and biogenic amine and D-lactate production were also examined. Furthermore, the principal properties of B. coagulans IDCC 1201 as probiotics, such as resistance to abiotic stress and intestinal adhesion, were studied. The whole-genome analysis demonstrated that B. coagulans IDCC 1201 had no antibiotic resistance or toxigenic genes; the strain was susceptible to the nine antibiotics proposed by the European Food Safety Authority. Moreover, this strain lacked hemolytic and ß-glucuronidase activities. Additionally, it was confirmed that B. coagulans IDCC 1201 produced undesirable metabolites, including biogenic amines or D-lactate, at a safe level. Finally, the strain exhibited functional potential as a probiotic in terms of abiotic tolerance, such as bile tolerance and intestinal adhesion in in vitro experiments. In conclusion, B. coagulans IDCC 1201 can be considered as a safe probiotic with regard to human health.

Safety assessment of Streptococcus thermophilus IDCC 2201 used for product manufacturing in Korea.

Safety evaluation of probiotics has become increasingly important for human consumption in food industry. The aims of this study were to assess safety of Streptococcus thermophilus IDCC 2201 through in vitro and in vivo tests. In results, this strain was found to be negative for hemolytic and ß-glucuronidase activity. In addition, thermophilus IDCC 2201 was susceptible to nine antibiotics suggested by EFSA. In accordance with MIC tests, whole-genome analysis indicated that S. thermophilus IDCC 2201 neither harbors antibiotic resistance nor toxigenic genes. Furthermore, none of the biogenic amines including tyramine and histamine was produced and negligible amounts of D-lactate were produced by S. thermophilus IDCC 2201. Finally, it was confirmed that there was no mortality and toxicity throughout single-dose oral toxicity tests in rats. Therefore, we report that S. thermophilus IDCC 2201 is considered to be safe for human consumption as probiotics.

Cellulosic Nanomaterial Production Via Fermentation by Komagataeibacter sp. SFCB22-18 Isolated from Ripened Persimmons.

Bacterial nanocellulose (BNC) which is generally synthesized by several species of bacteria has a wide variety of industrial uses, particularly in the food and material industries. However, the low levels of BNC production during the fermentation process should be overcome to reduce its production cost. Therefore, in this study, we screened and identified a new cellulose-producing bacterium, optimized production of the cellulose, and investigated the morphological properties of the cellulosic materials. Out of 147 bacterial isolates from ripened fruits and traditional vinegars, strain SFCB22-18 showed the highest capacity for BNC production and was identified as Komagataeibacter sp. based on 16S rRNA sequence analysis. During 6-week fermentation of the strain using an optimized medium containing 3.0% glucose, 2.5% yeast extract, 0.24% acetic acid, 0.27% Na2HPO4, and 0.5% ethanol at 30°C, about 5 g/l of cellulosic material was produced. Both imaging and IR analysis proved that the produced cellulose would be nanoscale bacterial cellulose.