Boulardii Strains Research Articles

Effect of Mutant and Engineered High-Acetate-Producing Saccharomyces cerevisiae var. boulardii Strains in Dextran Sodium Sulphate-Induced Colitis.

Acetate-producing Saccharomyces cerevisiae var. boulardii strains could exert improved effects on ulcerative colitis, which here, was preclinically evaluated in an acute dextran sodium sulphate induced model of colitis. Nine-week-old female mice were divided into 12 groups, receiving either drinking water or 2.75% dextran sodium sulphate for 7 days, combined with a daily gavage of various treatments with different levels of acetate accumulation: sham control (phosphate buffered saline, no acetate), non-probiotic control (Baker's yeast, no acetate), probiotic control (Enterol®, transient acetate), and additionally several Saccharomyces cerevisiae var. boulardii strains with respectively no, high, and extra-high acetate accumulation. Disease activity was monitored daily, and feces samples were collected at different timepoints. On day 14, the mice were sacrificed, upon which blood and colonic tissue were collected for analysis. Disease activity in inflamed mice was lower when treated with the high-acetate-producing strain compared to sham and non-probiotic controls. The non-acetate-producing strain showed higher disease activity compared to the acetate-producing strains. Accordingly, higher histologic inflammation was observed in non- or transient-acetate-producing strains compared to the sham control, whereas this increase was not observed for high- and extra-high-acetate-producing strains upon induction of inflammation. These anti-inflammatory findings were confirmed by transcriptomic analysis of differentially expressed genes. Moreover, only the strain with the highest acetate production was superior in maintaining a stable gut microbial alpha-diversity upon inflammation. These findings support new possibilities for acetate-mediated management of inflammation in inflammatory bowel disease by administrating high-acetate-producing Saccharomyces cerevisae var. boulardii strains.

Differentiation of Saccharomyces boulardii from Saccharomyces cerevisiae Strains Using the qPCR-HRM Technique Targeting AAD15 Gene

ABSTRACT S. boulardii, the probiotic variety of S. cerevisiae, is known for its different characteristics, even though the two strains share more than 99% genomic relatedness. Therefore, S. boulardii has been examined and evaluated for differential characteristics in many studies. This study aims to explain the differentiation of S. boulardii from well-identified S. cerevisiae strains by biochemical characteristics and the qPCR-HRM. Initially, S. cerevisiae isolates from the HUF Culture Collection, were screened for basic biochemical characteristics such as growth temperature, glucose, and galactose utilization tests. Out of all selected cultures, the viability at gastric pH was investigated by spectrophotometric readings. The results showed that one isolate was more resistant than other strains when exposed to gastric acids. After sequencing and phylogeny steps using LR0R universal primer, there was no significant differentiation for S. boulardii strains and therefore HRM analysis was performed to interrogate sequence variation and differentiate S. boulardii strains. HRM analysis has successfully distinguished even highly closely related yeast strains.

YEASTRACT+: a portal for the exploitation of global transcription regulation and metabolic model data in yeast biotechnology and pathogenesis.

YEASTRACT+ (http://yeastract-plus.org/) is a tool for the analysis, prediction and modelling of transcription regulatory data at the gene and genomic levels in yeasts. It incorporates three integrated databases: YEASTRACT (http://yeastract-plus.org/yeastract/), PathoYeastract (http://yeastract-plus.org/pathoyeastract/) and NCYeastract (http://yeastract-plus.org/ncyeastract/), focused on Saccharomyces cerevisiae, pathogenic yeasts of the Candida genus, and non-conventional yeasts of biotechnological relevance. In this release, YEASTRACT+ offers upgraded information on transcription regulation for the ten previously incorporated yeast species, while extending the database to another pathogenic yeast, Candida auris. Since the last release of YEASTRACT+ (January 2020), a fourth database has been integrated. CommunityYeastract (http://yeastract-plus.org/community/) offers a platform for the creation, use, and future update of YEASTRACT-like databases for any yeast of the users' choice. CommunityYeastract currently provides information for two Saccharomyces boulardii strains, Rhodotorula toruloides NP11 oleaginous yeast, and Schizosaccharomyces pombe 972h-. In addition, YEASTRACT+ portal currently gathers 304 547 documented regulatory associations between transcription factors (TF) and target genes and 480 DNA binding sites, considering 2771 TFs from 11 yeast species. A new set of tools, currently implemented for S. cerevisiae and C. albicans, is further offered, combining regulatory information with genome-scale metabolic models to provide predictions on the most promising transcription factors to be exploited in cell factory optimisation or to be used as novel drug targets. The expansion of these new tools to the remaining YEASTRACT+ species is ongoing.

Strain engineering and metabolic flux analysis of a probiotic yeast Saccharomyces boulardii for metabolizing l-fucose, a mammalian mucin component

BackgroundSaccharomyces boulardii is a probiotic yeast that exhibits antimicrobial and anti-toxin activities. Although S. boulardii has been clinically used for decades to treat gastrointestinal disorders, several studies have reported weak or no beneficial effects of S. boulardii administration in some cases. These conflicting results of S. boulardii efficacity may be due to nutrient deficiencies in the intestine that make it difficult for S. boulardii to maintain its metabolic activity.ResultsTo enable S. boulardii to overcome any nutritional deficiencies in the intestine, we constructed a S. boulardii strain that could metabolize l-fucose, a major component of mucin in the gut epithelium. The fucU, fucI, fucK, and fucA from Escherichia coli and HXT4 from S. cerevisiae were overexpressed in S. boulardii. The engineered S. boulardii metabolized l-fucose and produced 1,2-propanediol under aerobic and anaerobic conditions. It also produced large amounts of 1,2-propanediol under strict anaerobic conditions. An in silico genome-scale metabolic model analysis was performed to simulate the growth of S. boulardii on l-fucose, and elementary flux modes were calculated to identify critical metabolic reactions for assimilating l-fucose. As a result, we found that the engineered S. boulardii consumes l-fucose via (S)-lactaldehyde-(S)-lactate-pyruvate pathway, which is highly oxygen dependent.ConclusionTo the best of our knowledge, this is the first study in which S. cerevisiae and S. boulardii strains capable of metabolizing l-fucose have been constructed. This strategy could be used to enhance the metabolic activity of S. boulardii and other probiotic microorganisms in the gut.

Anti-salmonella properties of kefir yeast isolates: An in vitro screening for potential infection control

The rise of antibiotic resistance has increased the need for alternative ways of preventing and treating enteropathogenic bacterial infection. Various probiotic bacteria have been used in animal and human. However, Saccharomyces boulardii is the only yeast currently used in humans as probiotic. There is scarce research conducted on yeast species commonly found in kefir despite its claimed potential preventative and curative effects. This work focused on adhesion properties, and antibacterial metabolites produced by Kluyveromyces lactis and Saccharomyces unisporus isolated from traditional kefir grains compared to Saccharomyces boulardii strains. Adhesion and sedimentation assay, slide agglutination, microscopy and turbidimetry assay were used to analyze adhesion of Salmonella Arizonae and Salmonella Typhimurium onto yeast cells. Salmonella growth inhibition due to the antimicrobial metabolites produced by yeasts in killer toxin medium was analyzed by slab on the lawn, turbidimetry, tube dilution and solid agar plating assays. Alcohol and antimicrobial proteins production by yeasts in killer toxin medium were analyzed using gas chromatography and shotgun proteomics, respectively. Salmonella adhered onto viable and non-viable yeast isolates cell wall. Adhesion was visualized using scanning electron microscope. Yeasts-fermented killer toxin medium showed Salmonella growth inhibition. The highest alcohol concentration detected was 1.55%, and proteins with known antimicrobial properties including cathelicidin, xanthine dehydrogenase, mucin-1, lactadherin, lactoperoxidase, serum amyloid A protein and lactotransferrin were detected in yeasts fermented killer medium. These proteins are suggested to be responsible for the observed growth inhibition effect of yeasts-fermented killer toxin medium. Kluyveromyces lactis and Saccharomyces unisporus have anti-salmonella effect comparable to Saccharomyces boulardii strains, and therefore have potential to control Salmonella infection.

Production of lactic and acetic acids during fermentation of milk fortified with kiwi juice using Saccharomyces boulardii and lactobacilli

Purpose: To investigate the synergistic effect of Saccharomyces boulardii and lactobacilli on lactic and acetic acids produced during fermentation of milk fortified with kiwi juice, relative to fermentation of unfortified milk.
 Methods: Skimmed milk was fortified with kiwi juice (4 % v/v) and fermented for 12 h at 37 °C by a combination of S. boulardii and lactobacilli strains. Lactic and acetic acids were determined using gas chromatography-mass spectrometry (GS-MS).
 Results: The presence of kiwi juice in the milk stimulated the production of lactic (1.35 g/100g) and acetic (0.29 mg/g) by S. boulardii in the absence of lactobacilli. When S. boulardii was inoculated with Lb. casei 20975, the production of lactic acid and acetic acid increased to 2.36 g/100 g and 0.71 mg/g, respectively. Furthermore, acid production increased when Lb. plantarum RS (35-11), Lb. casei LCS, and Lb. plantarum JXJ (6 - 12) were inoculated into milk free of kiwi juice in which S. boulardii was grown. Saccharomyces boulardii resulted in marginal production of acids by Lb. fermentum F9.
 Conclusion: These results show that acid production is positively affected by some lactobacilli strains in the milk whether fortified with kiwi juice or free of this juice. However, fermentation of these formulations for a period longer than 6 h may result in losses in acid yield.

Comparison of functional characteristics of distinct Saccharomyces boulardii strains isolated from commercial food supplements

Saccharomyces boulardii is a probiotic yeast with several health benefits. An important number of products are available in the market claiming to contain probiotic S. boulardii strains and these numbers are increasing. In this study, S. boulardii strains were isolated from commercial products, identified and characterised in terms of several probiotic features. Eleven distinct S. boulardii strains were identified by genotypic tests followed by FT-IR spectroscopy and biochemical analysis which were used to differentiate S. boulardii strains from other yeasts including Saccharomyces cerevisiae. All tested strains showed high level of survival under harsh conditions and auto-aggregation characteristics. The hydrophobicity of S. boulardii strains revealed strain specific features and only two strains survived at pH of 2.5. A high level of survival under bile salts conditions were observed for all tested strains. Distinct S. boulardii strains were found to be resistant to bacterial antibiotics whereas they were sensitive to fungal antibiotics. Finally, S. boulardii strains showed high level of antibacterial and antifungal activities particularly against Salmonella Typhimurium and Aspergillus niger, respectively. This study revealed the differences in the functional characteristics of S. boulardii strains available in the market.

Чи можливо використання нанокристалічних сполук у середовищах для гіпотермічного зберігання дріжджів штаму Saccharomyces cerevisiae var. boulardii?

Чи можливо використання нанокристалічних сполук у середовищах для гіпотермічного зберігання дріжджів штаму Saccharomyces cerevisiae var. boulardii?

In vitro investigation of Debaryomyces hansenii strains for potential probiotic properties.

In this study, 23 Debaryomyces hansenii strains, isolated from cheese and fish gut, were investigated in vitro for potential probiotic properties i.e. (1) survival under in vitro GI (gastrointestinal) conditions with different oxygen levels, (2) adhesion to Caco-2 intestinal epithelial cells and mucin, and (3) modulation of pro- and anti-inflammatory cytokine secretion by human monocyte-derived dendritic cells. As references two commercially available probiotic Saccharomyces cerevisiae var. boulardii (S. boulardii) strains were included in the study. Our results demonstrate that the different D. hansenii yeast strains had very diverse properties which could potentially lead to different probiotic effects. One strain of D. hansenii (DI 09) was capable of surviving GI stress conditions, although not to the same degree as the S. boulardii strains. This DI 09 strain, however, adhered more strongly to Caco-2 cells and mucin than the S. boulardii strains. Additionally, two D. hansenii strains (DI 10 and DI 15) elicited a higher IL-10/IL-12 ratio than the S. boulardii strains, indicating a higher anti-inflammatory effects on human dendritic cells. Finally, one strain of D. hansenii (DI 02) was evaluated as the best probiotic candidate because of its outstanding ability to survive the GI stresses, to adhere to Caco-2 cells and mucin and to induce a high IL-10/IL-12 ratio. In conclusion, this study shows that strains of D. hansenii may offer promising probiotic traits relevant for further study.

Twenty-Five Years of Research on Saccharomyces boulardii Trophic Effects: Updates and Perspectives

Saccharomyces boulardii (S. boulardii) was discovered by Henri Boulard in 1920 in Indochina. Since then, lyophilized preparations of S. boulardii have increasingly been used throughout the world, providing empirical evidence of its efficacy as an adjuvant agent to treat diarrhea and prevent antibiotic-associated complications. Since 1982, the year of the first publication [1], increasing numbers of studies have been conducted each year to determine the mechanism(s) of action of S. boulardii and to evaluate whether it has beneficial properties for the host organism. More recently, placebo-controlled, double-blind studies have demonstrated the efficacy of S. boulardii as a probiotic medication and as a biotherapeutic agent in some intestinal disorders in both children and adults. The target intestinal disorders are listen in Table 1 along with the corresponding references [2–28]. The references listed here were selected from the MEDLINE database (NIH, National Library) from among 247 publications, including both clinical and experimental studies. The number of publications on S. boulardii has increased since the mid-1990s, now reaching 15 publications per year, while very recently meta-analyses have demonstrated the beneficial effects of S. boulardii treatment, mainly in Clostridium difficileassociated disease, antibiotic-associated diarrheas, and acute infectious diarrheas. Saccharomyces boulardii is a nonpathogenic yeast widely prescribed in a lyophilized form in many countries of the world and used as a biotherapeutical agent [30–32]. The objective of the present report is to provide an update on the intestinal trophic properties of S. boulardii in endoluminal fluid and in intestinal cells at a molecular level. Several authors, using different methods of molecular biology, have shown [33–35] that S. boulardii strains are clustered within the species of S. cerevisiae, the latter having no probiotic effect. The taxonomic position of S. boulardii was evaluated by sequence analysis of the D1/D2 domain of 26S rDNA, the ITS1-5.8S rDNA-ITS2 region, and the mitochondrial cytochrome-c oxidase II gene [36]. The trophic effects of S. boulardii on human and on rat small-intestinal cells have been studied since 1986 [37] and show that oral administration of the lyophilized preparation of the yeast results in a stimulation of human and rat brush border membrane (BBM) enzymes including lactase, sucrase-isomaltase, maltase glucoamylase [37], and a,atrehalase. Incorporation of C-glucosamine into lactase precursor was stimulated by S. boulardii cells whether cells were viable or killed by heating [37]. In addition, oral therapy enhanced the intestinal synthesis of the receptor for polymeric immunoglobulins and the secretion of s-IgA [38] and transporters such as the sodium–glucose cotransporter SGLT1 with increase in glucose uptake by BBM vesicles [39]. Although the precise mechanisms by which yeast cells exert these trophic effects remain unknown there are at least three potential explanations. First, orally given yeast cells secrete directly into the endoluminal fluid enzymes such as sucrase, a,a-trehalase, aminopeptidase [40], and a phosphatase that dephosphorylates very actively the endotoxin of E. coli 55B5, resulting in an inactivation of the toxins by more than 60% as attested by the marked reduction in tissue TNF-a levels and in the absence of liver and cardiac tissue lesions [41]. J.-P. Buts (&) Laboratory of Pediatric Gastroenterology and Nutrition, Faculty of Medicine, Universite Catholique de Louvain, Tour Pasteur +3, 53, Avenue Mounier, 1200 Brussels, Belgium e-mail: buts@gype.ucl.ac.be

Assimilation of omega 3 and omega 6 fatty acids and removing of cholesterol from environment by Saccharomyces cerevisiae and Saccharomyces boulardii strains

Assimilation of omega 3 and omega 6 fatty acids and removing of cholesterol from environment by Saccharomyces cerevisiae and Saccharomyces boulardii strains

Molecular tools for differentiating probiotic and clinical strains of Saccharomyces cerevisiae

The subtype of the Saccharomyces cerevisiae yeast species known as S. cerevisiae Hansen CBS 5926 was formerly believed to be a separate species, Saccharomyces boulardii. It is widely considered non-pathogenic and is used as a probiotic agent for treatment and prevention of diarrhea. The biological properties of Saccharomyces spp. show considerable intraspecies variability and the beneficial properties of probiotic yeasts are considered strain-specific. Septicemia and fungemia caused by S. boulardii have recently been described in both immunocompromised and immunocompetent patients receiving biotherapy with this yeast. It cannot be distinguished from other S. cerevisiae strains by phenotypic criteria, so identification of these infections requires molecular typing. To identify the most effective approach for distinguishing S. boulardii, we typed 35 isolates of S. cerevisiae, of which 27 were from various clinical specimens and 8 were isolates of S. boulardii (6 obtained from probiotic preparations and 2 from clinical specimens) using four different molecular methods, two based on PCR-restriction enzyme analysis or sequencing of rDNA spacer regions, the third based on microsatellite polymorphism analysis of the S. cerevisiae genes YKL139w and YLR177w, and the last based on hybridization analysis with retrotransposon Ty917. Several clinical isolates appeared to be identical to one or more other isolates with the first three methods used, whereas with the Ty917 hybridization method all of the isolates tested appeared to be very heterogeneous. The eight S. boulardii isolates were clearly distinguishable from the clinical S. cerevisiae isolates only with Ty917 hybridization and microsatellite DNA analyses. In the latter method, the eight S. boulardii isolates exhibited an allelic variant at one of loci tested that was not shared with any other strain. Our results suggest that microsatellite polymorphism analysis of the YKL139w and YLR177w genes, as well as the analysis by Ty917 hybridization, are the most useful tools for a correct identification of S. boulardii strains.

In vitro screening of probiotic properties of Saccharomyces cerevisiae var. boulardii and food-borne Saccharomyces cerevisiae strains

The probiotic potential of 18 Saccharomyces cerevisiae strains used for production of foods or beverages or isolated from such, and eight strains of Saccharomyces cerevisiae var. boulardii, was investigated. All strains included were able to withstand pH 2.5 and 0.3% Oxgall. Adhesion to the nontumorigenic porcine jejunal epithelial cell line (IPEC-J2) was investigated by incorporation of 3H-methionine into the yeast cells and use of liquid scintillation counting. Only few of the food-borne S. cerevisiae strains exhibited noteworthy adhesiveness with the strongest levels of adhesion (13.6–16.8%) recorded for two isolates from blue veined cheeses. Merely 25% of the S. cerevisiae var. boulardii strains displayed good adhesive properties (16.2–28.0%). The expression of the proinflammatory cytokine IL-1α decreased strikingly in IPEC-J2 cells exposed to a Shiga-like toxin 2e producing Escherichia coli strain when the cells were pre- and coincubated with S. cerevisiae var. boulardii even though this yeast strain was low adhesive (5.4%), suggesting that adhesion is not a mandatory prerequisite for such a probiotic effect. A strain of S. cerevisiae isolated from West African sorghum beer exerted similar effects hence indicating that food-borne strains of S. cerevisiae may possess probiotic properties in spite of low adhesiveness.

The Taxonomic Position of Saccharomyces boulardii as Evaluated by Sequence Analysis of the D1/D2 Domain of 26S rDNA, the ITS1-5.8S rDNA-ITS2 Region and the Mitochondrial Cytochrome- c Oxidase II Gene

A taxonomic study was carried out on eight strains of Saccharomyces boulardii. Morphological and physiological characteristics were consistent with those of Saccharomyces cerevisiae. Sequences of the D1/D2 domain of the 26S rDNA were identical for all strains examined and had a similarity value of 100% compared to sequences of the type strain of S. cerevisiae (CBS 1171T) and strain S288c. For all S. boulardii isolates was found the exact same ITS1-5.8S rDNA-ITS2 sequence, which displayed a close resemblance with the sequences published for S288c (99.9%), CBS 1171(T) (99.3%) and other S. cerevisiae strains. Sequence analysis of the mitochondrial cytochrome-c oxidase II gene (COX2) also resulted in identical sequences for the S. boulardii isolates and comparisons with available nucleotide sequences revealed close relatedness to strains of S. cerevisiae including S288c (99.5%) and CBS 1171(T) (96.6%). The electrophoretic karyotypes of the S. boulardii strains appeared quite uniform and although very typical of S. cerevisiae, they formed a cluster separate from strains of this species. The results of the present study strongly indicate a close relatedness of S. boulardii to S. cerevisiae and thereby support the recognition of S. boulardii as a member of S. cerevisiae and not as a separate species.

Protein fingerprinting of Saccharomyces isolates with therapeutic relevance using one- and two-dimensional electrophoresis.

In therapeutic products and preparations Saccharomyces cerevisiae is used because of its nutritive properties. Moreover, so called Saccharomyces boulardii yeasts are used in the prevention and treatment of several types of diarrhea. Taxonomically however, S. boulardii is not accepted as a distinct species. The protein fingerprint obtained after sodium dodecyl sulfate-polyacrylamide gel electrophoresis was identical for all isolates and therefore confirmed the designation of S. boulardii to the species S. cerevisiae. In contrast, using native polyacrylamide gel electrophoresis, 12 different protein fingerprints were detected, and allowed grouping of the product isolates. The spot patterns obtained by two-dimensional electrophoresis revealed a large degree of resemblance, however, small qualitative expression differences could be detected as well. Firstly, a spot having an isoelectric point of approximately 6 and 30 kDa could not be detected in S. boulardii yeasts. Secondly, nine different formations of spots occurred in the region around 16 kDa and pH 6. Therefore, on the one hand, it could be demonstrated that all of the product isolates belong to the same species, and on the other hand, it was possible to extensively subdivide the strains. In particular, two-dimensional electrophoresis allowed clustering of so called S. boulardii strains within the species S. cerevisiae.

Clustering of Saccharomyces boulardii strains within the species S. cerevisiae using molecular typing techniques

This study was undertaken to characterize and differentiate therapeutically relevant Saccharomyces yeasts. Among the isolates were so-called Saccharomyces boulardii strains, which are considered as probiotic agents, but whose taxonomic assignment is controversial. Moreover, the discriminative power of the applied molecular typing techniques should be evaluated. Genotyping was performed using species-specific polymerase chain reaction (PCR), randomly amplified polymorphic DNA-PCR, restriction fragment length polymorphism analysis of rDNA spacer regions and pulsed-field gel electrophoresis. Species-specific PCR assigned all of the product isolates to the species S. cerevisiae. By combining the other techniques, all isolates could be discriminated. Moreover, it could be demonstrated that probiotic S. boulardii strains form a separate cluster located within the species. With the exception of species-specific PCR, all of the applied methodologies were suitable for subspecies typing and indicated a close relationship between the probiotic strains. The methods applied in this study are considered powerful tools for quality control of therapeutically relevant yeasts. It is of crucial importance, especially regarding S. boulardii yeasts, to verify the identity of the correct strain, since the beneficial properties are considered to be strain-specific.

Utilization of prebiotic carbohydrates by yeasts of therapeutic relevance.

To investigate 17 strains of therapeutically relevant strains of Saccharomyces cerevisiae (including 10 strains of so-called S. boulardii) isolated from various pharmaceutical products, feed supplements and brewer's yeast for their capability of utilizing selected carbohydrates of prebiotic importance. Automated turbidimetric measurements and conventional test combinations were used to examine the basic sugar assimilation profiles of the test strains. It was shown that none of the so-called S. boulardii strains utilized galactose and palatinose. Among the prebiotic substrates, the yeasts indicated a pronounced preference for metabolizing the fructo-oligosaccharides. Yeast strains of therapeutic relevance can be successfully combined with certain prebiotics in symbiotic formulations. The results of this study may serve as a basis for the development of new pharmaceutical preparations for medical therapy and a better understanding of intestinal micro-ecology.