Safety Assessment of Streptococcus salivarius UBSS-01 in Rats and Double-Blind Placebo-Controlled Study in Healthy Individuals.

Streptococcus salivarius is a common, harmless, and prevalent member of the oral microbiota in humans. In the present study, the safety of S. salivarius UBSS-01 was evaluated using in silico methods and preclinical and clinical studies. In an acute toxicity study, rats were administered with 5 g/kg (500 × 109 CFU) S. salivarius UBSS-01. The changes in phenotypic behaviors and hematological, biochemical, electrolytes, and urine analyses were monitored. No toxicity was observed at 14 days post-treatment. The no observable effects limit (NOEL) of S. salivarius UBSS-01 was >5 g/kg in rats. In a 28-day repeat dose toxicity study, rats were administered S. salivarius UBSS-01 once daily at doses of 0.1, 0.5, and 1 g/kg (10, 50, and 100 billion CFU/kg, respectively) body weight. S. salivarius UBSS-01 did not influence any of the hematology parameters and clinical chemistry parameters in plasma and serum samples after 28-day repeated administration. No structural abnormality was observed in the histological examination of organs. Whole genome analysis revealed the absence of virulence factors or genes that may transmit antibiotic resistance. In the double-blind study with 60 human participants (aged 18-60 years), consumption of S. salivarius UBSS-01 for 30 days was found to be safe and results were comparable with placebo treatment These findings indicate that S. salivarius UBSS-01 may be safe for human consumption.

Toxicological safety evaluation of live Anaerobutyricum soehngenii strain CH106.

The gut commensal Anaerobutyricum soehngenii is an anaerobe that can produce both propionate and butyrate, metabolites that have been shown to have a positive effect on gut and overall health. Murine and human dose finding studies have shown that oral intake of A. soehngenii has a positive influence on peripheral insulin resistance, thereby reducing the risk of type 2 diabetes. A recent human intervention provided support for the mode of action of A. soehngenii as it affected gene expression in the duodenum, stimulated the secretion of GLP-1 and improved insulin sensitivity. For these reasons A. soehngenii has been proposed as a food ingredient. Before introducing this bacterium to the food chain, however, it must be established that oral intake of live A. soehngenii bacteria does not pose any health risk. As part of the safety analysis of A. soehngenii strain CH106, we performed genotoxicity assays to determine its mutagenic potential (bacterial reverse mutation and in vitro mammalian cell micronucleus tests) and a 90-day subchronic toxicity study in rats to determine overall toxicity potential. The results of both genotoxicity studies were negative, showing no genotoxic effects. For the 90-day subchronic toxicity study, no adverse events were registered that could be attributed to the feeding with A. soehngenii strain CH106. Even at the highest dose, which exceeds the expected daily human intake more than 100-fold, no adverse events were observed. These result support the conclusion that the use of A. soehngenii strain CH106 as a food ingredient is safe.

Safety assessment of Streptococcus salivarius DB-B5 as a probiotic candidate for oral health.

Streptococcus salivarius DB-B5 was previously isolated from the supragingival plaque of a healthy female adult and selected for development as a probiotic candidate for oral health. Probiotics are an important emerging therapeutic method for preventing, treating, and maintaining oral health. Although S. salivarius is a predominant member of the commensal oral microbiota and generally regarded as a safe species, it is recognized that each strain needs to be comprehensively assessed for safety. This study describes the in silico, in vitro, and clinical testing that were conducted to evaluate the safety of S. salivarius DB-B5. Both 16S rRNA and multi-gene phylogenetic reconstruction was used to confirm the taxonomic identity of this strain. Bioinformatic analysis of the genome demonstrated the absence of transmissible antibiotic resistance genes or virulence factors. Phenotypic testing further showed S. salivarius DB-B5 to be susceptible to clinically relevant antibiotics. S. salivarius DB-B5 displayed weak alpha-hemolysis, and does not produce biogenic amines. In a randomized, double-blind, placebo-controlled clinical study, consumption of S. salivarius DB-B5 at 10 billion CFU/day for 4 weeks by healthy adults was safe and well-tolerated (ClinicalTrials.gov registry number NCT04492631). This work has indicated that S. salivarius DB-B5 is a safe probiotic candidate.

Genotoxicity and subchronic general toxicity assessments of Lactobacillus curvatus WiKim 38 using Sprague-Dawley rats.

Lactobacillus curvatus WiKim 38 (LCW), isolated from kimchi, has shown novel immunomodulatory and anti-inflammatory properties. In the present study, to obtain data on the safety of LCW, we performed three genotoxicity (bacterial reverse mutation, chromosome aberration, and micronucleus) and two general toxicity (single-dosing and 13-week repeated-dosing) studies. In the genotoxicity assessment, LCW showed no increased reverse mutation for 4 strains of Salmonella typhimurium and a strain of Escherichia coli. In addition, LCW did not induce chromosome aberrations at concentrations up to 5000 µg/mL in cultured Chinese hamster lung (CHL) cells and did not induce an increased frequency of micronuclei in the bone marrow cells of rats at concentrations up to 2000 mg/kg. In the acute toxicity study using Sprague-Dawley (SD) rats, the approximate lethal dose of LCW was determined to be over 5000 mg/kg body weight (b.w.) in both sexes. Finally, in the subchronic toxicity study, no LCW-related adverse effects were observed at concentrations up to 5000 mg/kg b.w./day. Consequently, LCW is considered not to have mutagenic effects, and its no-observed-adverse-effect-level (NOAEL) is 5000 mg/kg b.w., equivalent to approximately 4.71 × 109 CFU/kg b.w., suggesting the LCW could be a potential probiotic for humans based on its safety profile.

Planococcus maritimus ML1206 Isolated from Wild Oysters Enhances the Survival of Caenorhabditis elegans against Vibrio anguillarum.

With the widespread occurrence of aquaculture diseases and the broad application of antibiotics, drug-resistant pathogens have increasingly affected aquatic animals' health. Marine probiotics, which live under high pressure in a saltwater environment, show high potential as a substitute for antibiotics in the field of aquatic disease control. In this study, twenty strains of non-hemolytic bacteria were isolated from the intestine of wild oysters and perch, and a model of Caenorhabditis elegans infected by Vibrio anguillarum was established. Based on the model, ML1206, which showed a 99% similarity of 16S rRNA sequence to Planococcus maritimus, was selected as a potential marine probiotic, with strong antibacterial capabilities and great acid and bile salt tolerance, to protect Caenorhabditis elegans from being damaged by Vibrio anguillarum. Combined with plate counting and transmission electron microscopy, it was found that strain ML1206 could significantly inhibit Vibrio anguillarum colonization in the intestinal tract of Caenorhabditis elegans. Acute oral toxicity tests in mice showed that ML1206 was safe and non-toxic. The real-time qPCR results showed a higher expression level of genes related to the antibacterial peptide (ilys-3) and detoxification (ugt-22, cyp-35A3, and cyp-14A3) in the group of Caenorhabditis elegans protected by ML1206 compared to the control group. It is speculated that ML1206, as a potential probiotic, may inhibit the infection caused by Vibrio anguillarum through stimulating Caenorhabditis elegans to secrete antibacterial effectors and detoxification proteins. This paper provides a new direction for screening marine probiotics and an experimental basis to support the potential application of ML1206 as a marine probiotic in aquaculture.

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.

Lactobacillus rhamnosus MP108: Toxicological evaluation.

A lyophilized powder of Lactobacillus rhamnosus MP108, a strain isolated from infant feces, has been developed for potential as a probiotic. To assess safety, three genetic toxicity studies (Ames, in vivo mouse micronucleus, and in vivo mouse spermatocyte chromosome aberration assays) and one 90-day oral rat toxicity study were conducted. Lactobacillus rhamnosus MP108 had no activity in the Ames assay, with or without metabolic activation, up to the highest dose tested (5000 µg/plate). Also, no evidence of genotoxicity was observed in the two in vivo genotoxicity assays up to the maximum dose tested of 5.6 g/kg body weight. Feeding of L. rhamnosus MP108 to SD rats at doses of 0, 0.25, 0.5, or 1.5 g/kg body weight/day had no statistically or biologically significant effects on any of the parameters measured. An evaluation of Lactobacillus spp. generally, and L. rhamnosus specifically, showed no indication of concern for translocation or pathogenicity. The results of the toxicology studies showed no indications for safety concerns. PRACTICAL APPLICATION: Lactobacillus rhamnosus MP108 is a bacterium that shows potential as a probiotic. While this bacterium has been used in food and food production, a series of studies were conducted to provide further assurance that this particular strain of L. rhamnosus is safe.

Two-weeks repeated-dose oral toxicity study of Pediococcus acidilactici J9 in a mice model.

BACKGROUND: Helicobacter pylori (H. pylori) is an important pathogen that causes chronic gastritis and peptic ulcer, and is related to the development of gastric carcinoma. Several chemicals, including antibiotics, have been used to eradicate H.pylori. However, more studies are yet requred to accomplish a sufficient therapy. Pediococcus acidilactici (P. acidilactici) J9 were studied for inhibition of binding of H.pylori binding to human gastric cell lines. This study was performed in order to investigate the repeated-dose toxicity of P. acidilactici J9 in male and female mice. RESULTS: C57BL/6 male and female Mus musculus were divided into four groups (n = 10 in each group). P. acidilactici J9 was administered daily by oral injection of vehicle control at dosage levels to a low-dose group (500 mg/kg/day), middle-dose group (1000 mg/kg/day), and high-dose group (2000 mg/kg/day) for 2 weeks. After 14 days of exposure, the blood biochemistry and hematology were investigated, along with a histopathology exam. There were no bacterial-related deaths or abnormal clinical signs in either gender of mouse. The data was observed during the period in terms of body weight, food intake, and water consumption. Also, no alterations in organ weights upon administration of P. acidilactici J9 alone were observed. The adhesion and growth of H. pylori were inhibited by a 24 h treatment of H. pylori and P. acidilactici J9 on adenocarcinoma gastric (AGS) cells, which are gastric cancer cells. Compared to the control group (AGS cell and H. pylori), the number of H. pylori analyzed by FACS significantly (p < 0.01) decreased after incubation of AGS cell with P. acidilactici J9 for 24 h. CONCLUSIONS: These results suggest that the oral application of P. acidilactici J9, up to a dosage level of 2000 mg/kg/day, causes no adverse effects in both male and female mice. P. acidilactici J9 inhibits the adhesion of H.pylori to AGS cancer cells. When used as probiotics, P. acidilactici J9 may help decrease the occurrence of gastritis and reduce the risk of H.pylori infection with promising safety issues.

Unexpected Observations: Probiotic Administration Greatly Aggravates the Reproductive Toxicity of Perfluorobutanesulfonate in Zebrafish.

The present study exposed adult zebrafish to 0, 10, and 100 µg/L perfluorobutanesulfonate (PFBS) with or without dietary supplement of probiotic Lactobacillus rhamnosus. Interaction between probiotic and PFBS on sex endocrine and reproduction was investigated. It was striking to find that PFBS and probiotic coexposures almost ceased the fecundity, which was accompanied by disturbances in sex hormones and oocyte maturation in females. In contrast, probiotic additive efficiently antagonized the estrogenic activity of PFBS in males. For the first time, this study reported that probiotic heavily depended on sex to modulate the endocrine disruption and reproductive toxicity of aquatic pollutants.

Safety evaluation of Bacillus coagulans SNZ 1969 in Wistar rats.

Bacillus coagulans SNZ 1969 is a rod-shaped, slightly acidophilic, gram-positive, spore forming and highly resilient bacteria. B. coagulans SNZ 1969 has GRAS (Generally Recognized As Safe) status for use as a probiotic in foods (US FDA number GRN-597). The present study was aimed to assess the safety of a proprietary strain Bacillus coagulans SNZ 1969 by conducting acute and sub-acute 28 days repeated dose oral toxicity studies in Wistar Rats. In the acute toxicity study, the rats were orally fed with 2000 mg/kg body weight (BW) (5 × 1011 CFU/g) of B. coagulans SNZ 1969 as a single dose to determine the LD50 values. In the sub-acute repeated dose toxicity study, six groups of experimental rats received 250, 500, 1000 mg/kgBW/day (5 × 1011 CFU/g) of the test item for 28 consecutive days. The control animals received only water. Four groups of rats were sacrificed after 28 days and the remaining two groups were kept as recovery groups and sacrificed after 42 days. The results of these study indicate that there were no treatment related changes in any of the parameters studied i.e. clinical signs, body weight, food intake, urinalysis, hematological examinations, clinical biochemistry, gross pathology and histopathology after 28 days of repeated administration. Based on the results it was concluded that the LD50 of Bacillus coagulans SNZ 1969 is more than 2000 mg/kg body weight and the NOAEL derived from this study was 1000 mg/kg/day for 28 days, this corresponds to the 5 × 1011 CFU/kg.

Preliminary safety assessment of a new Bacteroides fragilis isolate.

The novel commensal strain of Bacteroides fragilis HCK-B3 isolated from a healthy Chinese donor was discovered beneficial effects of attenuating lipopolysaccharides-induced inflammation. In order to contribute to the development of natural next-generation probiotic strains, the safety assessment was carried out with in vitro investigations of its morphology, potential virulence genes and antimicrobial resistance, and an in vivo acute toxicity study based on both healthy and immunosuppressed mice by cyclophosphamide injection. Consequently, the potential virulence genes in the genome of B. fragilis HCK-B3 have yet been identified as toxicity-associated. The absence of plasmids prevents the possibility of transferring antibiotic resistance features to other intestinal commensals. No intracorporal pathogenic properties were observed according to the body weight, hematological and liver parameters, cytokine secretions and tissue integrity. In addition, B. fragilis HCK-B3 performed alleviations on part of the side effects caused by the cyclophosphamide treatment. Thus, the novel strain of B. fragilis HCK-B3 was confirmed to be non-toxigenic and did not display adverse effects in both healthy and immune-deficient mice at a routinely applicable dose.

Deciphering the interplay between the genotoxic and probiotic activities of Escherichia coli Nissle 1917.

Although Escherichia coli Nissle 1917 (EcN) has been used therapeutically for over a century, the determinants of its probiotic properties remain elusive. EcN produces two siderophore-microcins (Mcc) responsible for an antagonistic activity against other Enterobacteriaceae. EcN also synthesizes the genotoxin colibactin encoded by the pks island. Colibactin is a virulence factor and a putative pro-carcinogenic compound. Therefore, we aimed to decouple the antagonistic activity of EcN from its genotoxic activity. We demonstrated that the pks-encoded ClbP, the peptidase that activates colibactin, is required for the antagonistic activity of EcN. The analysis of a series of ClbP mutants revealed that this activity is linked to the transmembrane helices of ClbP and not the periplasmic peptidase domain, indicating the transmembrane domain is involved in some aspect of Mcc biosynthesis or secretion. A single amino acid substitution in ClbP inactivates the genotoxic activity but maintains the antagonistic activity. In an in vivo salmonellosis model, this point mutant reduced the clinical signs and the fecal shedding of Salmonella similarly to the wild type strain, whereas the clbP deletion mutant could neither protect nor outcompete the pathogen. The ClbP-dependent antibacterial effect was also observed in vitro with other E. coli strains that carry both a truncated form of the Mcc gene cluster and the pks island. In such strains, siderophore-Mcc synthesis also required the glucosyltransferase IroB involved in salmochelin production. This interplay between colibactin, salmochelin, and siderophore-Mcc biosynthetic pathways suggests that these genomic islands were co-selected and played a role in the evolution of E. coli from phylogroup B2. This co-evolution observed in EcN illustrates the fine margin between pathogenicity and probiotic activity, and the need to address both the effectiveness and safety of probiotics. Decoupling the antagonistic from the genotoxic activity by specifically inactivating ClbP peptidase domain opens the way to the safe use of EcN.

Safety assessment of Weissella confusa - A direct-fed microbial candidate.

Weissella confusa is part of the lactic acid bacteria genera and a member of an autochthonous microbiota of humans and livestock. W. confusa has been proposed to have potential as a direct-fed microbial product, however, there is a lack of studies assessing its safety. A toxicological safety assessment of W. confusa was performed using a battery of in vitro, ex vivo and in vivo studies, testing for genotoxicity, skin and eye irritation and general toxicity. The bacterial reverse mutation (Ames) study did not reveal any genotoxicity in the presence and absence of metabolic activation (S9) at concentrations up to 5000 µg/mL. Moreover, an in vivo mammalian erythrocyte micronucleus study did not reveal any biologically relevant or statistically significant increases in the frequency of micronucleated polychromatic erythrocytes in mice, when tested at concentrations up to 2000 mg/kg body weight. W. confusa did not exert any skin irritation potential when tested in reconstructed skin membranes. When tested for eye irritation using an ex vivo model of isolated chicken eyes, mild irritation was observed. The 90-day sub-chronic oral toxicity (gavage) study was performed using Sprague Dawley rats at concentrations up to 92 × 108 cfu/kg body weight/day (nominal). The results showed that W. confusa were well tolerated, and no signs of toxicity were seen. The No Observed Adverse Effect Level (NOAEL) for both female and male animals was the highest concentration administered, 92 × 108 cfu/kg body weight/day (nominal). In conclusion, the toxicological studies performed confirmed W. confusa to be safe, making it a good candidate as a direct-fed microbial product.

Microencapsulated Lactobacillus plantarum LAB12 Showed No Sign of Acute or Sub-chronic Toxicity In Vivo.

Lactic acid bacteria (LAB) with probiotic properties are useful options for prophylactic and therapeutic applications against gastrointestinal diseases. The safety of probiotics should, however, be verified before incorporation into food or drinks. The present study had encapsulated Lactobacillus plantarum LAB12 within microcapsules that could withstand extremely high temperature (up to 100 °C) during pelletisation. The microencapsulated LAB12 were then tested for their acute (single dosing) and sub-chronic (a 90-day feeding) toxicity. For acute toxicity study, six male Sprague-Dawley rats were being administered with a single dose of freeze-dried microencapsulated LAB12 at 11 log CFU/kg BW through oral gavage. No clear treatment-related effects were observed after 14 days. For sub-chronic toxicity study, rodents were randomly divided into four groups (6 rats/sex/group) and treated with 0, 8, 9 and 10 log CFU/kg BW of microencapsulated LAB12 in pellet form. No mortality or treatment-related findings were observed in terms of clinical body weight, water intake, or food consumption. No treatment-related adverse effects were observed in blood and tissue samples. The no-observed-adverse-effect-level (NOAEL) for microencapsulated LAB12 was 2.5 × 1010 CFU/kg BW for both genders. These results imply that LAB12 are likely non-pathogenic and non-toxic.

Assessing safety of Lactobacillus plantarum MTCC 5690 and Lactobacillus fermentum MTCC 5689 using in vitro approaches and an in vivo murine model.

Lactobacilli have a long history of safe use in human nutrition, however, inclusion of any new strain, despite its safe usage evidence, warrants proper analysis of its safety and toxicity under the purview of existing regulations. In the present investigation, Lactobacillus plantarum MTCC 5690 and Lactobacillus fermentum MTCC 5689 were evaluated for their safety and toxicity using both in vitro and in vivo approaches. The in vitro assays included mucin degradation, hemolytic activity, biogenic amine production and platelet aggregation assay. The safety was also assessed using acute, subacute and subchronic assays, bacterial translocation studies, intravenous and intravenous administration and genotoxicity assay in murine model. The outcome of this toxicological safety assessment indicated that both the test strains lacked any harmful metabolic activity or any genotoxic effects. Furthermore, the results of oral toxicity studies in mice revealed that short term administration of high cell mass concentration of 1012 cfu/animal as well as long term feeding of the probiotic strains did not alter any hematological, general health parameters or cause any organ specific disorder. Based upon these scientific assessments and supported by long history of safe use, both MTCC 5690 and MTCC 5689 may be considered safe for human consumption.

Inocuidade micotoxicológica e viabilidade de Aspergillus spp com potencial probiótico provenientes do trato digestório bovino / Mycotoxicological safety and viability of Aspergillus spp with probiotic potential from the bovine digestive tract]

Objetivou-se, com a presente pesquisa, avaliar a viabilidade de Aspergillus spp. com potencial probiótico durante o armazenamento e diante da pressão da microbiota autóctone, bem como a tolerância aos principais ácidos graxos da fermentação ruminal. Verificou-se também a inocuidade micotoxicológica desses isolados fúngicos cultivados em meio de cultivo sólido. Foram avaliados 20 isolados de Aspergillus spp., provenientes do trato gastrointestinal de bovinos criados em Urochloa decumbens lignificada. Esses fungos foram identificados por análise de sequências do DNAr e foram selecionados por apresentarem expressivo potencial celulolítico. O método vapor de amônia foi utilizado para detecção de cepas produtoras de micotoxinas. Os isolados foram avaliados quanto à viabilidade de crescimento em fluido ruminal por até 96 horas e estocagem em condições ambientais. Observou-se que os fungos avaliados não produziram aflatoxinas e que 95% dos isolados apresentaram resistência aos ácidos ruminais. Dois isolados, selecionados a partir das análises anteriores, apresentaram viabilidade sob a pressão da microbiota autóctone e de metabólitos do ecossistema ruminal e permanecem viáveis por, no mínimo, dois anos. Conclui-se que os isolados do gênero Aspergillus selecionados nesta pesquisa apresentam características fisiológicas para serem utilizados com aditivos microbianos ou probióticos para o ambiente ruminal.(AU)

The aims in this study were to evaluate the viability of Aspergillus spp. with probiotic potential during storage, pressure of autochthonous microbiota and tolerance to the main fatty acids of ruminal fermentation. The mycotoxicological safety was also verified. Twenty isolates from the gastrointestinal tract of cattle raised in lignified Urochloa decumbens were identified by rDNA sequence analysis and were previously selected because they showed significant cellulolytic potential. The ammonia vapor method was used to detect the production of mycotoxins. The isolates were evaluated for viability of ruminal fluid growth for up to 96 hours and storage under environmental conditions. The evaluated fungi did not produce aflatoxins, and 95% of them had resistance to ruminal acids. Two isolates, selected according these tests, presented viability on autochthonous microbiota pressure and metabolites from the ruminal ecosystem and remain viable for at least two years. In this research, the selected Aspergillus spp. isolates present physiological characteristics to be used with microbial additives or probiotic.(AU)

Safety evaluations and lipid-lowering activity of an Arthrospira platensis enriched diet: A 1-month study in rats.

Arthrospira platensis (A. platensis) is worldwide consumed as dietary supplement, but its use in the form of whole biomass for food purposes may raise toxicity concerns. The aim of this study was to preliminarily evaluate the safety of an A. platensis F&M-C256-enriched diet (20% (weight/weight) corresponding to 12g/kg body weight/day), administered to rats for 1month. A. platensis F&M-C256-enriched diet was well tolerated: behavior, body weight, food consumption and growth curves were not affected; no discomfort, no deaths and no physical signs related to the treatment were observed during the administration period; food daily consumption and apparent digestibility were comparable to those of the standard laboratory AIN-76 control diet. Daily water consumption and urine excretion were, on the contrary, significantly higher (27.18±1.24 vs 21.53±1.68ml and 12.63±0.99 vs 7.00±1.29ml respectively), probably because of a slight increase in sodium intake in rats fed A. platensis F&M-C256-enriched diet. Biochemical markers of kidney and liver function were not varied but a significant increase in cholesterol-HDL and a decreased plasma triglycerides level was observed in rats fed A. platensis F&M-C256-enriched diet. These last changes were associated with an increased fecal lipids excretion and liver PPAR-α gene expression. These results indicate that A. platensis F&M-C256 is likely safe and well tolerated even at a high dosage in rodents and suggest that it may represent a promising functional food for preventing or even for managing dyslipidemias.

Safety evaluation of HOWARU® Restore (Lactobacillus acidophilus NCFM, Lactobacillus paracasei Lpc-37, Bifidobacterium animalis subsp. lactis Bl-04 and B. lactis Bi-07) for antibiotic resistance, genomic risk factors, and acute toxicity.

Although probiotic lactobacilli and bifidobacteria are generally considered safe by various regulatory agencies, safety properties, such as absence of transferable antibiotic resistance, must still be determined for each strain prior to market introduction as a probiotic. Safety requirements for probiotics vary regionally and evaluation methods are not standardized, therefore methodologies are often adopted from food ingredients or chemicals to assess microbial safety. Four individual probiotic strains, Lactobacillus acidophilus NCFM®, Lactobacillus paracasei Lpc-37®, Bifidobacterium animalis subsp. lactis strains Bl-04®, and Bi-07®, and their combination (HOWARU® Restore) were examined for antibiotic resistance by broth microdilution culture, toxin genes by PCR and genome mining, and acute oral toxicity in rats. Only B. lactis Bl-04 exhibited antibiotic resistance above a regulated threshold due to a tetW gene previously demonstrated to be non-transferable. Genomic mining did not reveal any bacterial toxin genes known to harm mammalian hosts in any of the strains. The rodent studies did not indicate any evidence of acute toxicity following a dose of 1.7-4.1 × 1012 CFU/kg body weight. Considering a 100-fold safety margin, this corresponds to 1.2-2.8 × 1012 CFU for a 70 kg human. Our findings demonstrate a comprehensive approach of in vitro, in silico, and in vivo safety testing for probiotics.

Antimicrobial and antioxidant activities of Saccharomyces cerevisiae IFST062013, a potential probiotic.

BACKGROUND: Probiotic yeast has become a field of interest to scientists in recent years. METHODS: Conventional cultural method was employed to isolate and identify yeast and standard methods were used to determine different probiotic attributes, antimicrobial and antioxidant properties. RESULTS: This study reports potential probiotic properties of a strain of S. cerevisiae IFST 062013 isolated from fruit. The isolate is tolerant to a wide range of temperature and pH, high concentration of bile salt and NaCl, gastric juice, intestinal environment, α-amylase, trypsin and lysozyme. It can produce organic acid and showed resistance against tetracycline, ampicillin, gentamycin, penicillin, polymixin B and nalidixic acid. It can assimilate cholesterol, can produce killer toxin, vitamin B12, glutathione, siderophore and strong biofilm. It showed moderate auto-aggregation ability and cell surface hydrophobicity. The isolate can produce enzymes such as amylase, protease, lipase, cellulose, but unable to produce galactosidase. The isolate can't produce gelatinase and DNase. The isolate showed moderate anti-microbial activity against bacteria and fungi and cell lysate showed better antimicrobial activity than whole cell and culture supernatant. Again, the isolate showed better anti-bacterial activity against gram negative bacteria than gram positive. The isolate showed strong antioxidant activity, reducing power, nitric oxide and hydroxyl radical scavenging activity, significant brine shrimp cytotoxicity and acute toxicity and metal ion chelating activity. The isolate did not induce any detectable change in general health of mice upon oral toxicity testing and found to be safe in mouse model. The isolate improve lymphocyte proliferation and cytokine production in treated mice. CONCLUSIONS: Such isolate could be potential as probiotic to be used therapeutically.

Safety assessment of Bacillus subtilis CU1 for use as a probiotic in humans.

Bacillus subtilis CU1 is a recently described probiotic strain with beneficial effects on immune health in elderly subjects. The following work describes a series of studies supporting the safety of the strain for use as an ingredient in food and supplement preparations. Using a combination of 16S rDNA and gyrB nucleotide analyses, the species was identified as a member of the Bacillus subtilis complex (B. subtilis subsp. spizizenii). Further characterization of the organism at the strain level was achieved using random amplified polymorphic DNA polymerase chain reaction (RAPD PCR) and pulsed field gel electrophoresis (PFGE) analyses. B. subtilis CU1 did not demonstrate antibiotic resistance greater than existing regulatory cutoffs against clinically important antibiotics, did not induce hemolysis or produce surfactant factors, and was absent of toxigenic activity in vitro. Use of B. subtilis CU1 as a probiotic has recently been evaluated in a 16-week randomized, double-blind, placebo-controlled, parallel-arm study, in which 2 × 109 spores per day of B. subtilis CU1 were administered for a total 40 days to healthy elderly subjects (4 consumption periods of 10 days separated by 18-day washouts). This work describes safety related endpoints not previously reported. B. subtilis CU1 was safe and well-tolerated in the clinical subjects without undesirable physiological effects on markers of liver and kidney function, complete blood counts, hemodynamic parameters, and vital signs.