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Autoaggregation (%) of isolates is calculated after 2 h, 6 h, 12 h and 24 h of incubation. Each value represents the mean value ± standard deviation (SD) (n = 3). Bars with different lower-case letters denoted significantly different (p < 0.05).
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The present study was designed to isolate Bifidobacterium strains from raw camel milk and to investigate their probiotic characteristics. Among 35 isolates, 8 were identified as Gram-positive, catalase negative, non-spore forming, non-motile and V or Y shaped rods. B-2, B-5, B-11, B-19 and B-28 exhibited good survival at low pH and high bile salt c...
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Bifidobacterium and Lactobacillus are known to be common members of the human intestinal microbiota, which play important roles in maintaining the homeostasis of host gut microenvironment. Several bifidobacterial and lactobacilli strains have been used as probiotics for health benefits. The exopolysaccharides (EPSs) produced by strains from Bifidob...
Citations
... At the end of the incubation, the hemolytic activity of probiotic strains was classified according to zone color around the bacterial colonies. Yellow and green zones were accepted as β-hemolytic and α-hemolytic, respectively, and red zones were accepted as safe (γ-hemolytic) [21]. All experiments were performed in duplicates including the control group. ...
Traditional Turkish fermented foods like boza, pickles, and tarhana are recognized for their nutritional and health benefits, yet the probiotic potential of lactic acid bacteria (LAB) strains isolated from them remains underexplored. Sixty-six LAB strains were isolated from fermented foods using bacterial morphology, Gram staining, and catalase activity. The isolates were differentiated at strain level by RAPD-PCR (Random Amplification of Polymorphic DNA—Polymerase Chain Reaction) and twenty-five strains were selected for further evaluation of acid and bile salt tolerance. Among these, ten strains exhibited high tolerance and were subsequently assessed for adhesion to Caco-2 colorectal carcinoma cells, antimicrobial activity, exopolysaccharide (EPS) production, lysozyme resistance, and hemolytic activity. Using k-means clustering, three strains: Lactiplantibacillus plantarum ES-3, Pediococcus pentosaceus N-1, and Enterococcus faecium N-2 demonstrated superior probiotic characteristics, including significant acid (100% survival at pH3.0) and 0.3% bile salt tolerance (57%, 64%, 67%), strong adhesion to intestinal cells (65%, 88%, 91%), high lysozyme resistance (88%, 88%, 77%), and produced high amounts of EPS. These strains show promising potential as probiotics and warrant further investigation to confirm their functional properties and potential applications.
... L. reuteri MG4722 was grown in MRS broth, streaked onto a TSA plate, and incubated for 48 h at 37°C. After 24 h, hemolytic activity was determined by evaluating the presence or absence of hemolysis around the colonies (Yasmin et al. 2020). ...
Background
Oral diseases with high prevalence worldwide are recognized as severe health problems. Probiotics are used to prevent oral diseases, including dental caries, oral malodor, periodontitis, and subgingival plaque. In this study, we aimed to confirm the antibacterial effect of probiotics on oral pathogens and to assess their characterization and safety as probiotics.
Methods
The antibacterial effects of Lacticaseibacillus rhamnosus MG4706, Lacticaseibacillus paracasei MG4715, and Limosilactobacillus reuteri MG4722 on the growth biofilm formation of Streptococcus mutans , Aggregatibacter actinomycetemcomitans , and Porphyromonas gingivalis were evaluated. We also investigated the production of antibacterial substances (H 2 O 2 and reuterin) by these strains and their ability to adhere to oral epithelial cells. The safety of L. reuteri MG4722 was verified through whole-genome sequencing analysis and antibiotic susceptibility, lactate dehydrogenase activity, hemolytic activity, and bile acid hydrolase activity. The reuterin biosynthesis genes of L. reuteri MG4722 were identified using genomic analysis.
Results
L. reuteri MG4722 significantly inhibited the growth of S . mutans , A. actinomycetemcomitans , and P. gingivalis and suppressed the biofilm formation by A. actinomycetemcomitans . In addition, it showed considerable adhesion ability to oral epithelial cells. L. reuteri MG4722 produced H 2 O 2 and reuterin as antibacterial substances, as confirmed by the presence of genes encoding the antibacterial compounds reuterin, reuteran, and reutericyclin. L. reuteri MG4722 showed no hemolysis, bile salt hydrolase activity, antibiotic resistance or toxicity to HT-29 cells, and no antibiotic-resistance genes were identified.
Conclusion
L. reuteri MG4722 demonstrated antibacterial effects on oral pathogens by producing antibacterial substances and adhering to oral epithelial cells. These results suggest that L. reuteri MG4722 could be an effective probiotic for oral health.
... Sodium nitrite reduction abilities of the isolated LAB strains were evaluated as previously described [28]. 100 µl of probiotic culture (1 × 10 8 CFU/ml) was added to the prepared sodium nitrite solution of 150 µg/ml and incubated anaerobically for 12 h at 37 °C. ...
Lactic acid bacteria (LAB), traditionally consumed as fermented foods, are now being applied to the medical field beyond health-functional food as probiotics. Therefore, it is necessary to continuously discover and evaluate new strains with suitable probiotic characteristics, mainly focusing on safety. In this study, we isolated eight new strains from postmenopausal vaginal fluid using culturomics approaches, an emerging area of interest. Data showed that most strains possessed significant cell surface hydrophobicity (≥ 76%), auto-aggregation capacity (17 to 61%), strong adhesion activity (8 to 34%), and excellent resistance to gastric acid, bile salt, and digestive enzyme, enhancing their survival in the gastrointestinal tract. Moreover, the strains exhibited functional characteristics, including substantial antibacterial activity with a minimal inhibitory concentration (MIC) ranging from 12.5 to 50%. They also harbored bacteriocins genes, produced short-chain fatty acids (acetate and propionate), exhibited significant phagocytic activity, possessed high antioxidative properties, rapidly depleted sodium nitrite, and exhibited proteolysis and β-glucosidase activity. In addition, heat-killed LAB strains significantly reduced the gene expressions of proinflammatory cytokines such as IL-β, IL-6, and iNOS in macrophages. Safety assessment revealed no cytotoxicity in macrophage cell lines. All strains tested negative for biogenic amine or H2O2 production, displayed no gelatinase or hemolytic activity, lacked virulence genes or detrimental enzymes, and displayed antibiotic susceptibility. In summary, these newly isolated strains demonstrate excellent probiotic functionality with a strong focus on safety, making them promising candidates for future drug development in the relevant fields.
... A green zone around the colony was considered indicative of α-hemolysis, a clear zone around the colony was considered as β-hemolysis, and the absence of a zone around the colony was considered as c-hemolysis. Only isolates with c-hemolysis were used in the subsequent analysis [15]. ...
Alpha-glucosidase inhibitors are one of the therapies used for treating type 2 diabetes by inhibiting the absorption of carbohydrates in the gastrointestinal tract. In addition to antimicrobial activity, some probiotic species show α-glucosidase inhibitor activity, making them potential alternative therapies for type 2 diabetes. This study aimed to characterize probiotics from “trites,” a traditional food from North Sumatra, Indonesia, that exhibit α-glucosidase inhibition, potentially useful for type 2 diabetes treatment. The probiotic potential of the isolates was evaluated through antagonistic activity, acid tolerance, bile tolerance, and susceptibility to antimicrobial agents. α-Glucosidase inhibition was tested with acarbose as a control. The best-performing isolate, LBSU8, was identified as Pediococcus acidilactici through 16S rRNA gene sequencing. Gene analysis using genome sequencing for LBSU8 revealed antimicrobial secondary metabolites, including RiPPs, polyketide, and NRP, while capsular polysaccharide might contribute to its antidiabetic activity. Though no specific α-glucosidase inhibitory secondary metabolites were identified, enzymes like dTDP-glucose 4,6-dehydratase, transketolase, and glucose-1-phosphate thymidylyltransferase may contribute to this activity. P. acidilactici LBSU8 shows potential as an alternative diabetes therapy in the food and drug industries. Further studies are needed to elucidate the exact mechanism behind its α-glucosidase inhibitory activity and to explore its efficacy in clinical settings.
... The spread of bacteria on defibrinated sheep blood agar plate is a standard method for assessing hemolytic activity of potential probiotic strains. The absence of hemolytic activity of lactobacilli suggests that they do not produce hemolytic enzymes, which could otherwise damage the mucosal membrane and facilitate the infiltration of pathogens and toxins [57]. To further investigate the safety of vaginal lactobacilli and nanofibers, we co-incubated them with Caco-2 cells. ...
Electrospun nanofibers offer a highly promising platform for the delivery of vaginal lactobacilli, providing an innovative approach to preventing and treating vaginal infections. To advance the application of nanofibers for the delivery of lactobacilli, tools for studying their safety and efficacy in vitro need to be established. In this study, fluorescent (mCherry and GFP) and luminescent (NanoLuc luciferase) proteins were expressed in three vaginal lactobacilli (Lactobacillus crispatus, Lactobacillus gasseri and Lactobacillus jensenii) and a control Lactiplantibacillus plantarum with the aim to use this technology for close tracking of lactobacilli release from nanofibers and their adhesion on epithelial cells. The recombinant proteins influenced the growth of the bacteria, but not their ability to produce hydrogen peroxide. Survival of lactobacilli in nanofibers immediately after electrospinning varied among species. Bacteria retained fluorescence upon incorporation into PEO nanofibers, which was vital for evaluation of their rapid release. In addition, fluorescent labelling facilitated efficient tracking of bacterial adhesion to Caco-2 epithelial cells, while luminescence provided important quantitative insights into bacterial attachment, which varied from 0.5 to 50% depending on the species. The four lactobacilli in dispersion or in nanofibers were not detrimental for the viability of Caco-2 cells, and did not demonstrate hemolytic activity highlighting the safety profiles of both bacteria and PEO nanofibers. To summarize, this study contributes to the development of a promising delivery system, tailored for local administration of safe vaginal lactobacilli.
... The hemolytic activity of strain WP12 was assessed as previously described with few modifications (Yasmin et al., 2020). WP12 was streaked onto a blood agar plate containing 5% blood and incubated at 37 • C for 48 h. ...
... These phenotypes are common in LAB. 36 Thus, it was inferred that these putative LAB are safe for further utilization as probiotics that can be incorporated into drugs or functional foods. The colonies of the two isolates were translucent, round, shiny, and convex (Figure 1b), consistent with the characteristics of a LAB isolate described in other studies. ...
Probiotics are microorganisms that are safe and stable under product development conditions and are used as adjuncts to food and drugs to promote health, including the acceleration of nutrient digestion. Here, we present evidence on the ability of two isolates, Lacticaseibacillus paracasei BCRC-16100 and L. paracasei ZFM54, to (1) enhance carbohydrate digestion, (2) tolerate processing conditions, and (3) demonstrate safety in terms of antimicrobial resistance (AMR). These approaches include whole-genome sequence (WGS) analysis, gene expression, and bioactivity assays. WGS revealed genes encoding enzymes involved in carbohydrate digestion, tolerance to processing conditions, and AMR. The ability of the two strains to digest carbohydrates was confirmed by glucose release when cultured alongside starch. The isolates also showed versatility across a range of temperatures and alcohol concentrations, indicating their suitability for product development. Genes cause AMR, particularly against vancomycin, through three mechanisms: transporter control, transcriptional regulation, and efflux pumps. Furthermore, promoter, gene expression, and transposable element analyses showed that some upregulated AMR genes in the presence of antimicrobials were transposable. Altogether, we show the potential of the two isolates for incorporation into products as probiotics to improve carbohydrate digestion, while considering precautions regarding mobile AMR genes that may compromise safety.
... Selected colonies were subjected to a primary screening that involved morphology of the colony, Gram staining, negative staining, oxidase, catalase, and other biochemical tests. The isolates were also tested for safety aspects via experiments like hemolytic activity (Yasmin et al. 2020). ...
The present research was aimed to isolate potential probiotic organisms from dairy products locally made in and around the Saurashtra region of Gujarat. A total of 224 colonies were screened for primary attributes. Based on the results, 70 isolates were carried further for secondary screening. Out of these, only 23 isolates were further tested for antioxidant activities. Only 6 potential probiotic strains were found to have all the probiotic attributes. These isolates demonstrated survivability up to 4 h at pH ≤ 3, bile concentration ≥ 1.5%, autoaggregation ability ≥ 81.08%, and cell surface hydrophobicity more than 70% while using toluene as the test hydrocarbon. The promising six isolates were subjected to 16S rRNA sequencing for species‐level identification and found to be belonging to the genus Bacillus, Enterococcus, and Lactobacillus. The isolates demonstrated higher antioxidant potential as determined by ABTS, DPPH, and FRAP methods. For all three methods, L. rhamnosus was taken as a positive control that showed 85.61%, 39.56%, and 78.18% reduction of free radicals as determined by the ABTS, DPPH, and FRAP methods, respectively. Compared to this, Limosilactobacillus fermentum BAB 7912 demonstrated the highest reduction of ABTS radicals (83.45%), while Bacillus subtilis BAB 7918 reduced 29.95% DPPH free radicals and Bacillus spizizenii BAB 7915 reduced 80.93% ferric ions as determined by the FRAP method. Isolates were subjected to 16S rRNA sequencing for species‐level identification and found to be belonging to genus Bacillus, Enterococcus, and Lactobacillus.
... Lactic acid bacteria isolate that will be applied to food products need to be tested for pathogenicity by observing the hemolysis activity of LAB on blood agar media. The absence of a clear zone around the bacterial colony indicates that the LAB isolate is not pathogenic because it cannot hemolyze (ˠ-hemolysis), namely the inability of bacteria to lyse red blood cells on blood agar media (Yasmin et al., 2020). Tallapragada et al. (2018) stated that the absence of hemolysis activity is a safe prerequisite for selecting LAB isolates for food products because LAB must be non-pathogenic. ...
Sago starch production in local industries is still carried out traditionally and uses poor-quality water. This production causes sago starch to be fermented spontaneously, resulting in sour sago and possibly contamination by pathogenic bacteria. Lactic acid bacteria (LAB) can produce lactic acid and are suitable for use as a starter. Adding LAB as a starter in sago starch fermentation is expected to reduce the number of pathogenic bacterial growths, thereby increasing food safety in sago starch. This research aimed to obtain LAB and evaluate their use in sago starch fermentation to improve food safety. LAB selection was conducted by testing the LAB tolerance ability to low pH and the adaptability of the LAB growth in sago starch. This study was carried out using and without a LAB liquid starter. The water source during the fermentation originated from drinking water and the sago starch industrial factory. The fermentation was carried out for ten days at room temperature with an observation every two days. The results showed that fermented sago starch using drinking water did not harbor E. coli, Salmonella, or Shigella bacterial contamination. In contrast, sago starch fermented using water from the factory harbored these bacterial contaminations. Adding LAB IL1 isolate as a starter in fermentation showed the ability to reduce the number of pathogenic bacteria in sago starch.
... Hemolytic activity was determined using Columbia agar (Oxoid, UK) containing 5% sheep blood (MBcell, Korea) [24]. LAB strains were streaked and cultured at 37 o C for 48 h. ...
Current cancer burden caused by persistent infection with human papillomaviruse genotype 16 (HPV16) cannot be ignored. The related mechanisms of oncoproteins E6 and E7 from HPV16 on keratinocyte malignant transformation need to be further elucidated. GSE3292 dataset analysis revealed the upregulation of ETS transcription factor 3 (ELF3) and cyclin E2 (CCNE2). To verify whether there is an interaction between ELF3 and CCNE2, E74 like ELF3 and CCNE2 expression profiles as well as their putative binding sites were analyzed using bioinformatics. Retroviruses encoding HPV16 E6 and E7 genes were used to induce immortalization of human foreskin keratinocytes (HFKs) in vitro. Dual luciferase reporters assay was used to verify the binding of ELF3 and CCNE2. The effect of ELF3 on the immortalized cells was investigated using CCK-8 assay, cell cycle analysis and western blot. ELF3 and CCNE2 presented overexpression patterns in head and neck squamous cell carcinoma. HPV16 E6/E7-expressing HFKs showed enhanced viability, accelerated cell cycle as well as upregulated ELF3 and CCNE2. ELF3 overexpression enhanced the activity of CCNE2 promoter. ELF3 silencing reduced viability, induced cell cycle arrest and suppressed expressions of CCNE2, E6 and E7 in HPV16 E6/E7-expressing HFKs. Downregulation of ELF3 played an inhibiting role in the malignant transformation of HPV16 E6/E7-immortalized HFKs by decreasing CCNE2 expression.