Characterisation of the probiotic qualities exhibited by lactobacilli strains isolated from the anogenital tract

Introduction: Lactobacilli are well-documented probiotics that exert health benefits on their host. They exhibit characteristics that make them potential alternative treatments to address the antimicrobial resistance conundrum and diseases. Their mechanism of action varies with strain and species. Five lactobacilli strains previously isolated from the anogenital region were subjected to several assessments highlighted in the FAO/WHO document, ‘Guidelines for the Evaluation of Probiotics in Food’ to determine its suitability as potential probiotics. Methods: The five lactobacilli strains were subcultured onto Man de Rogosa agar (MRS). Their ability to auto- and co-aggregate was determined spectrophotometrically. Simultaneously, the cell surface hydrophobic properties of these strains towards xylene and toluene were evaluated using the microbial adhesion to hydrocarbon (MATH) test. The lactobacilli strains were also tested for their ability to withstand acid, bile and spermicide to determine their level of tolerance. Results: Lact. reuteri 29A, L. delbrueckii 45E and L. reuteri 29B exhibited the highest degree of auto- and co-aggregation properties. These lactobacilli strains also demonstrated high cell surface hydrophobicity, with the exception of L. delbrueckii 45E. Further tests to evaluate the isolated lactobacilli tolerance identified L. reuteri 29B as the most tolerant strain towards low pH (pH 2.5 for 4 h), high bile concentration (0.5% for 4 h) and high spermicides concentration (up to 10%). Conclusion: Out of the five lactobacilli strains which possessed high antimicrobial activities, L. reuteri 29B portrayed the best probiotic qualities with good auto- and co-aggregation abilities and high tolerance against acid, bile and spermicide

Safety evaluation of lactobacillus strains as potential probiotics for human use

Lactobacillus species are generally regarded as safe (GRAS) by the United State Food and Drug Administration (FDA) due to their long history of safe consumption in traditional fermented foods. However, there have been few reported cases of Lactobacillus species associated with clinical conditions such as bacteremia, endocarditis and localized infections. In view of this, it is relevant to have a more thorough safety assessment of new strains before use as probiotics. Therefore, the aim of this study was to evaluate the safety of a new potentially probiotic lactobacilli strains. In this study, safety assessment of five lactobacilli strains, previously characterized as potential probiotic bacteria was carried out to determine their suitability for human use. Antibiotic resistance patterns were determined using agar overlay disc diffusion method. Hemolytic activity, bile salt deconjugation and amino acid decarboxylase activity were determined in vitro on agar plates. Enzymatic activities and carbohydrate fermentation profile were determined using API systems. Bile salt hydrolase (BSH) activity and D-/L-lactic acid production were quantified spectrophotometrically. Among the five strains, Lactobacillus reuteri S29 PFB was evaluated for safety in vivo in a 28 day sub-acute oral toxicity study in 40 BALB/c mice. The mice were randomized into 4 groups and orally fed with L. reuteri S29 PFB (1 x 108, 109 and 1010 CFU/day). Blood and organs collected at the end of the intervention period were analyzed for health parameters (blood biochemistry, hematology and histopathology) and incidence of translocation to blood and organs. Results from this study demonstrated that all the strains were able to utilize glucose and lactose and the enzymatic profiles of the strains correlated with the carbohydrate fermentation. Harmful bacterial enzymes such as beta glucoronidase was not detected in all the strains. the tested lactobacilli strains were shown to be suceptible to penicillins, chloramphenicol and tetracycline. None of the strains were shown to be hemolytic or decarboxylate histidine, tyrosine, lysine or arginine to form biogenic amines. The concentrations of D-/L- lactic acid and bile salt hydrolase activity units were comparable to that of the commercial strain, L. reuteri RC-14. Results from the in vivo study demonstrated that oral administration of different doses of L. reuteri 29 PFB for 28 days had no adverse toxicological effect on the mice general health status, hematological and blood biochemical parameters and intestinal histological parameters. There were no incidence of translocation to blood and DNA fingerprinting patterns of isolates recovered from tissues did not match with that of the administered strain. Overall, the result from this study suggests that L. reuteri 29 PFB was shown to be a better candidate for probiotic development, and would likely be safe for human use

Isolation and safety characterisation of lactobacilli strains with antimicrobial properties as potential probiotics for human use

The present study focuses on the antimicrobial activity and safety aspect of lactobacilli strains isolated from the anogenital region. Sixty-two lactobacilli strains were isolated from 52 healthy, non-menstruating women from a local health clinic in Puchong, Selangor, Malaysia and confirmed by 16S rDNA sequencing. The samples were collected by sterile cotton swabs and directly streaked onto De Man, Rogosa, Sharpe agar prior to transportation to the lab on ice. A selective preliminary screening against several Candida albicans and Candida glabrata strains identified 5 promising lactobacilli strains for the subsequent assays; L. delbreuckii 45E, L. fermentum 28E, L. mucosae 28C, L. reuteri 29A and L. reuteri 29B. A follow-up antibacterial assay identified L. delbreuckii 45E, L. reuteri 29A and L. reuteri 29B as the lactobacilli strains with the highest inhibitory activity. Subsequent assays including haemolytic activity, susceptibility to antibiotics, quantification of D/l-lactic acid and H2O2 production as well as bile salt hydrolase (BSH activity) indicated that L. reuteri 29B was a promising probiotic candidate. An acute toxicity study in vivo involving the administration of 1 × 108, 1 × 109 and 1 × 1010 cfu/mouse/day demonstrated that L. reuteri 29B did not exert any adverse effects towards the mice which were evident through the absence of pathological changes in the histological examination and blood test. There was also a lack of bacterial translocation as confirmed by Random Amplified Polymorphic DNA (RAPD). The present study highlights the possibility of utilising L. reuteri 29B as a probiotic. It also demonstrates the possibility of isolating potential probiotics from the anogenital region