BdPUL12 depolymerizes ß-mannan-like glycans into mannooligosaccharides and mannose, which serve as carbon sources for Bacteroides dorei and gut probiotics.

Symbiotic bacteria, including members of the Bacteroides genus, are known to digest dietary fibers in the gastrointestinal tract. The metabolism of complex carbohydrates is restricted to a specified subset of species and is likely orchestrated by polysaccharide utilization loci (PULs) in these microorganisms. ß-Mannans are plant cell wall polysaccharides that are commonly found in human nutrients. Here, we report the structural basis of a PUL cluster, BdPUL12, which controls ß-mannan-like glycan catabolism in Bacteroides dorei. Detailed biochemical characterization and targeted gene disruption studies demonstrated that a key glycoside hydrolase, BdP12GH26, performs the initial attack on galactomannan or glucomannan likely via an endo-acting mode, generating mannooligosaccharides and mannose. Importantly, coculture assays showed that the B. dorei promoted the proliferation of Lactobacillus helveticus and Bifidobacterium adolescentis, likely by sharing mannooligosaccharides and mannose with these gut probiotics. Our findings provide new insights into carbohydrate metabolism in gut-inhabiting bacteria and lay a foundation for novel probiotic development.

Higher microbial diversity in raw than in pasteurized milk Raclette-type cheese enhances peptide and metabolite diversity after in vitro digestion.

Numerous bacteria are responsible for hydrolysis of proteins during cheese ripening. The raw milk flora is a major source of bacterial variety, starter cultures are needed for successful acidification of the cheese and proteolytic strains like Lactobacillus helveticus, are added for flavor improvement or acceleration of ripening processes. To study the impact of higher bacterial diversity in cheese on protein hydrolysis during simulated human digestion, Raclette-type cheeses were produced from raw or heat treated milk, with or without proteolytic L. helveticus and ripened for 120 days. Kinetic processes were studied with a dynamic (DIDGI®) in vitro protocol and endpoints with the static INFOGEST in vitro digestion protocol, allowing a comparison of the two in vitro protocols at the level of gastric and intestinal endpoints. Both digestion protocols resulted in comparable peptide patterns after intestinal digestion and higher microbial diversity in cheeses led to a more diverse peptidome after simulated digestion.

Proteolytic activity of selected commercial Lactobacillus helveticus strains on soy protein isolates.

Soy protein isolates were fermented by three commercial Lactobacillus helveticus strains for a maximum of seven days to investigate the resulting proteolysis. The proteolytic activity of the most active strain (LH88) was further analyzed (LC-MS/MS and GC-MS) and it was shown that the ß-conglycinin α subunit 1, ß-conglycinin α' subunit, glycinin G1, and 2S albumin were specifically degraded. Peptigram analysis and visualization of the crystal structure showed that the hydrolysis sites of ß-conglycinin α subunit, α' subunit, and the glycinin G1 were located on the surface of the molecule or at the mobile disordered region, hence being highly accessible for the proteinase of LH88. The proteins were partially further degraded to free amino acids, and subsequently catabolized to volatile compounds. However, most of the proteins remained native, even after seven days of fermentation, thus additional modification of protein structure or adjustment of fermentation conditions are required for effective generation of flavor compounds.

ß-Galactosidase from Lactobacillus helveticus DSM 20075: Biochemical Characterization and Recombinant Expression for Applications in Dairy Industry.

ß-Galactosidase encoding genes lacLM from Lactobacillus helveticus DSM 20075 were cloned and successfully overexpressed in Escherichia coli and Lactobacillus plantarum using different expression systems. The highest recombinant ß-galactosidase activity of ∼26 kU per L of medium was obtained when using an expression system based on the T7 RNA polymerase promoter in E. coli, which is more than 1000-fold or 28-fold higher than the production of native ß-galactosidase from L. helveticus DSM 20075 when grown on glucose or lactose, respectively. The overexpression in L. plantarum using lactobacillal food-grade gene expression system resulted in ∼2.3 kU per L of medium, which is approximately 10-fold lower compared to the expression in E. coli. The recombinant ß-galactosidase from L. helveticus overexpressed in E. coli was purified to apparent homogeneity and subsequently characterized. The Km and vmax values for lactose and o-nitrophenyl-ß-d-galactopyranoside (oNPG) were 15.7 ± 1.3 mM, 11.1 ± 0.2 µmol D-glucose released per min per mg protein, and 1.4 ± 0.3 mM, 476 ± 66 µmol o-nitrophenol released per min per mg protein, respectively. The enzyme was inhibited by high concentrations of oNPG with Ki,s = 3.6 ± 0.8 mM. The optimum pH for hydrolysis of both substrates, lactose and oNPG, is pH 6.5 and optimum temperatures for these reactions are 60 and 55 °C, respectively. The formation of galacto-oligosaccharides (GOS) in discontinuous mode using both crude recombinant enzyme from L. plantarum and purified recombinant enzyme from E. coli revealed high transgalactosylation activity of ß-galactosidases from L. helveticus; hence, this enzyme is an interesting candidate for applications in lactose conversion and GOS formation processes.

Influence of the culture preparation and the addition of an adjunct culture on the ripening profiles of hard cheese.

The aim of this work was to evaluate the impact of two factors on the ripening profiles of hard cooked cheeses: (F1) the growth medium for the primary and adjunct cultures, constituted by autochthonous strains: Lactobacillus helveticus 209 (Lh209) and Lactobacillus paracasei 90 (Lp90), respectively, and (F2) the addition of L. paracasei Lp90 as adjunct culture. Four types of cheeses were made: W and M cheeses in which only Lh209 was added after its growth in whey and MRS, respectively; Wa and Ma cheeses in which both strains (Lh209 and Lp90) were added after their growth in whey and MRS, respectively. Physicochemical and microbial composition, proteolysis and profiles of organic acids and volatile compounds were analyzed. According to the methodology of the cultures preparation, W and Wa cheeses showed a higher level of secondary proteolysis and lower level of primary proteolysis (P < 0·05), lower content of citric and acetic acids and higher amount of propionic acid (P < 0·05), in comparison with M and Ma cheeses. The incorporation of Lp90 increased the secondary proteolysis (P < 0·05), decreased the citric acid (P < 0·05), and increased the propionic acid only when was added after their growth in whey (P < 0·05). Both factors significantly modified the percentages of the volatile compounds grouped in chemical families; in addition, for the half of the compounds detected, significant differences were found. Based on the obtained results, the use of Lp90 as an adjunct in hard cooked cheeses, and the preincubation of the cultures in whey are strategies to accelerate the cheese ripening and to enhance the production of some characteristic compounds of this type of cheeses, such as propan-2-one, hexan-2-one, 2- and 3-methyl butanal, heptan-2-ol, acetic and 3-methylbutanoic acids and 3-hydroxy butan-2-one.

Probiotic mixture of Lactobacillus helveticus R0052 and Bifidobacterium longum R0175 attenuates hippocampal apoptosis induced by lipopolysaccharide in rats.

In recent years, the beneficial impact of targeted gut microbiota manipulation in various neurological disorders has become more evident. Therefore, probiotics have been considered as a promising approach to modulate brain gene expression and neuronal pathways even in some neurodegenerative diseases. The purpose of this study was to determine the effect of probiotic biotherapy with combination of Lactobacillus helveticus R0052 and Bifidobacterium longum R0175 on the expression levels of proteins critical to neuronal apoptosis in hippocampus of lipopolysaccharide (LPS)-exposed rats. Four groups of animals (Control, LPS, Probiotic + LPS, and Probiotic) were treated with maltodextrin (placebo) or probiotic (109 CFU/ml/rat) for 2 weeks by gavage. On the 15th day, a single intraperitoneal dose of saline or LPS (1 mg/kg) was injected and 4 h later, protein assessment was performed by western blotting in hippocampal tissues. LPS significantly increased the Bax, Bax/Bcl-2 ratio, and cleaved caspase-3 expression along with decreased the Bcl-2 and procaspase-3 protein levels. However, probiotic pretreatment (L. helveticus R0052 + B. longum R0175) significantly downregulated the Bax and Bax/Bcl-2 ratio accompanied with upregulated Bcl-2 expression. Prophylactic treatment with these bacteria also attenuated LPS-induced caspase-3 activation by remarkably increasing the expression of procaspase-3 while reducing the level of cleaved caspase-3 in target tissues. Our data indicate that probiotic formulation (L. helveticus R0052 + B. longum R0175) alleviated hippocampal apoptosis induced by LPS in rats via the gut-brain axis and suggest that this probiotic could play a beneficial role in some neurodegenerative conditions.

Single probiotic supplement suppresses colitis-associated colorectal tumorigenesis by modulating inflammatory development and microbial homeostasis.

BACKGROUND AND AIM: Chronic inflammation is a major contributor to the initiation and progression of cancers. Lactobacillus helveticus NS8, which was originally separated from fermented koumiss, exhibited anti-inflammatory functions in our prior studies. In this study, NS8 was investigated for its potential to prevent colitis-associated colorectal cancer (CAC). METHODS: The protective effects of NS8 against CAC was explored by employing the azoxymethane plus dextran sodium sulfate-induced carcinogenesis mouse model. The prevalences of T cells expressing specific inflammatory cytokines were measured by flow cytometry at the early stage of CAC. Inflammatory modulation by NS8 was also tested in the Caco2-Raw264.7 cell co-culture system. The alternations in the intestinal microbiota following the health-inflammation-cancer sequence were analyzed by 16S rDNA sequencing. RESULTS: Oral intake of NS8 lactobacilli clearly reduced tumor number and the degree of hyperplasia. The increased proliferation of enterocytes at the early stage of CAC was significantly suppressed by NS8, while the level of apoptosis was elevated. The anticancer effects of NS8 were associated with its anti-colitis outcomes before tumor formation. NS8 significantly suppressed the activation of NF-κB and upregulated the anti-inflammatory cytokine IL-10. Further analysis revealed the marked downregulation of IL-17-producing T cells by NS8. Furthermore, NS8 modulated intestinal dysbiosis by promoting beneficial commensal microbes while suppressing cancer-associated microbes. Notably, Bacteroides acidifaciens was the most sensitive commensal bacteria to NS8 intervention. CONCLUSION: These results provide insight into the protective effects of L. helveticus NS8 against colorectal cancer.

Probiotics Affect One-Carbon Metabolites and Catecholamines in a Genetic Rat Model of Depression.

SCOPE: Probiotics may influence one-carbon (C1) metabolism, neurotransmitters, liver function markers, or behavior. METHODS AND RESULTS: Male adult Flinders Sensitive Line rats (model of depression, FSL; n = 22) received Lactobacillus helveticus R0052 and Bifidobacterium longum R0175 (109 or 1010 colony-forming units per day) or vehicle for 10 weeks. The controls, Flinders Resistant Line rats (FRL, n = 8), only received vehicle. C1-related metabolites were measured in plasma, urine, and different tissues. Monoamine concentrations were measured in plasma, hippocampus, and prefrontal cortex. Vehicle-treated FSL rats had higher plasma concentrations of betaine, choline, and dimethylglycine, but lower plasma homocysteine and liver S-adenosylmethionine (SAM) than FRLs. FSL rats receiving high-dose probiotics had lower plasma betaine and higher liver SAM compared to vehicle-treated FSL rats. FSLs had higher concentrations of norepinephrine, dopamine, and serotonin than FRLs across various brain regions. Probiotics decreased plasma dopamine in FSLs in a dose-dependent manner. There were no detectable changes in liver function markers or behavior. CONCLUSIONS: Probiotics reduced the flow of methyl groups via betaine, increased liver SAM, and decreased plasma dopamine and norepinephrine. Since these changes in methylation and catecholamine pathways are known to be involved in several diseases, future investigation of the effect of probiotics is warranted.

Effect of ultrasound on lactic acid production by Lactobacillus strains in date (Phoenix dactylifera var. Kabkab) syrup.

Date syrup is rich in fermentable sugars and may be used as a substrate for different microbial fermentations, including lactic acid fermentation processes. The beneficial effects of ultrasounds (US) on bioprocesses have been reported for several microorganisms, due to the enhancement of cell growth, as well as improvements in yields and productivities. Therefore, US treatments (30 kHz, 100 W, 10-30 min) were applied to two lactobacilli (Lactobacillus helveticus PTCC 1332 and Lactobacillus acidophilus PTCC 1643), during fermentation using date syrup as substrate. The effects on lactic acid fermentation were evaluated by analyzing cell growth (dry cell weight and viable cell count), substrate consumption (quantification of glucose and fructose), and product formation (quantification of lactic acid) over time. The effects of US were also evaluated on cell membrane permeability. Both lactobacilli were able to grow well on date syrup without the need for addition of further ingredients. The US effects were highly dependent on treatment duration: treatments of 10- and 20-min stimulated lactobacilli growth, while the treatment extension to 30 min negatively affected cell growth. Similarly, the 10- and 20-min treatments increased sugar consumption and lactic acid production, contrarily to the 30-min treatment. All US treatments increased cell membrane permeability, with a more pronounced effect at more extended treatments. The results of this work showed that application of appropriate US treatments could be a useful tool for stimulation of lactic acid production from date syrup, as well as for other fermentative processes that use date syrup as substrate.

Redefining the effect of salt on thermophilic starter cell viability, culturability and metabolic activity in cheese.

This study investigated the differential effect of salt concentration in the outside and inside layers of brine salted cheeses on viability, culturability and enzyme activity of starter bacteria. The high-salt environment of the outside layer caused a sharp decrease in L. helveticus viability as measured by traditional plate counts. Remarkably, this was associated with lower release of intracellular enzymes (LDH), reduced levels of proteolysis and larger membrane integrity as measured by flow cytometry (FC) following classical Live/Dead staining. FC analysis of light scattering properties highlighted a significant reduction in size and granularity of the microbiota located in the cheese surface, suggestive of cell shrinkage and condensation of internal macromolecules probably due to hyperosmotic stress. The microbiota of the cheese surface were found to experience greater oxidative stress, as measured by FC analysis of the total levels of reactive oxygen species, compared to that of the interior layer. These results lead us to postulate that the physiology and health status of the microbiota were significantly different in the outer and inner layers of the cheese. The hyperosmotic environment of the outer layer resulted in reduced cell lysis, as measurable by assays based upon membrane integrity, but rather triggered cell death via mechanisms involving cell shrinkage and ROS-mediated damage of vital intracellular components. This study challenges the current thinking on how salt controls microbial activity in ripening cheese, especially in cheeses which are brine salted as local variations in biochemical ripening indices can differ significantly from the outside to the inside of a ripening cheese.

Flow cytometry: a versatile technology for specific quantification and viability assessment of micro-organisms in multistrain probiotic products.

AIMS: Classical microbiology techniques are the gold standard for probiotic enumeration. However, these techniques are limited by parameters of time, specificity and incapacity to detect viable but nonculturable (VBNC) micro-organisms and nonviable cells. The aim of the study was to evaluate flow cytometry as a novel method for the specific quantification of viable and nonviable probiotics in multistrain products. METHODS AND RESULTS: Custom polyclonal antibodies were produced against five probiotic strains from different species (Bifidobacterium bifidum R0071, Bifidobacterium longum ssp. infantis R0033, Bifidobacterium longum ssp. longum R0175, Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011). Evaluation of specificity confirmed that all antibodies were specific at least at the subspecies level. A flow cytometry method combining specific antibodies and viability assessment with SYTO® 24 and propidium iodide was applied to quantify these strains in three commercial products. Analyses were conducted on two flow cytometry instruments by two operators and compared with classical microbiology using selective media. Results indicated that flow cytometry provides higher cell counts than classical microbiology (P < 0·05) in 73% of cases highlighting the possible presence of VBNC. Equivalent performances (repeatability and reproducibility) were obtained for both methods. CONCLUSIONS: This study showed that flow cytometry methods can be applied to probiotic enumeration and viability assessment. Combination with polyclonal antibodies can achieve sufficient specificity to differentiate closely related strains. SIGNIFICANCE AND IMPACT OF THE STUDY: Flow cytometry provides absolute and specific quantification of viable and nonviable probiotic strains in a very short time (<2 h) compared with classical techniques (>48 h), bringing efficient tools for research and development and quality control.

In vitro analysis of 4-methylumbelliferone as a sole carbon source for Lactobacillus helveticus 2126.

In the recent years, 4-methylumbelliferone (4-MU) has been gaining importance, both as an anti-cancer agent and as a dietary supplement. The aim of this study was to determine the effectiveness of 4-MU as a carbon source for potential probiotic bacteria Lactobacillus helveticus 2126. For this purpose, a series of plate assays and infrared spectroscopy (FTIR) were used for 4-MU before and after the treatment with L. helveticus 2126. The plate assays indicated an initial inhibition followed by utilization of 4-MU that stimulated bacterial growth. A significant shift was observed in the FTIR peaks, which also have suggested possible extracellular activity of the bacteria for 4-MU utilization. SIGNIFICANCE AND IMPACT OF THE STUDY: 4-Methylumbelliferone (4-MU) is a widely used chloretic and is currently under research for treating colon cancer. Preliminary studies suggest that it has the potential to be used as an effective and sustainable prebiotic for the human microbiome, as it can be naturally obtained from plants. This manuscript describes the effectiveness of 4-MU as a carbon source for the probiotic bacteria Lactobacillus helveticus. Our study also suggests the role of bacterial superoxide dismutase in transforming 4-MU as a possible prebiotic for the human microbiome.

Screening for proteolytically active lactic acid bacteria and bioactivity of peptide hydrolysates obtained with selected strains.

In a screening for proteolytically active lactic acid bacteria, three strains, Lactobacillus delbrueckii ssp. lactis 92202, Lactobacillus helveticus 92201, and Lactobacillus delbrueckii ssp. bulgaricus 92059, showed the highest activities following growth in milk. All three strains degraded α- and ß-casein, but did not hydrolyse κ-casein. HPLC analysis of skim milk fermentation revealed increasing amounts of peptides after 5 and 10 h with Lb. d. ssp. bulgaricus 92059. Hydrolysates obtained with Lb. d. ssp. lactis 92202 and Lb. d. ssp. bulgaricus 92059 revealed the highest angiotensin-converting enzyme-inhibitory effect. The effect was dose dependent. Almost no effect (<10%) was seen for Lb. helveticus 92201. For Lb. d. ssp. bulgaricus 92059, maximal inhibition of approx. 65% was reached after 25 h of fermentation. In an in vitro assay measuring potential immunomodulation, hydrolysates of the three strains yielded anti-inflammatory activities in the presence of TNF-α. However, the effects were more pronounced at lower hydrolysate concentrations. In the absence of TNF-α, slight pro-inflammatory effects were observed. The hydrolysate of Lb. d. ssp. bulgaricus 92059, when purified by means of solid-phase extraction, exhibited pro-inflammatory activity. Sour whey containing Lb. d. ssp. bulgaricus 92059 cells showed pro-inflammatory activity while cell-free sour whey was clearly anti-inflammatory. In the purified hydrolysate, 20 different α- and ß-casein (CN)-derived peptides could be identified by LC-MS. Most peptides originated from the central and C-terminal regions of ß-casein. Peptide length was between 9 (ß-CN(f 59-67)) and 22 amino acids (ß-CN(f 117-138)).

Proteome analysis of Lactobacillus helveticus H9 during growth in skim milk.

Lactobacillus helveticus H9 was isolated from traditionally fermented yak milk in Tibet (China) with the ability to produce the antihypertensive peptides Val-Pro-Pro (VPP) and Ile-Pro-Pro (IPP) during milk fermentation. To understand the changes in the protein expression of L. helveticus H9, proteome analysis was performed at 3 different growth stages, lag phase (pH 6.1), log phase (pH 5.1), and stationary phase (pH 4.5) using 2-dimensional electrophoresis (2-DE). Further analysis showed that 257 differential protein spots were found and 214 protein spots were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS). The cellular functions of the differentially expressed proteins were complex. Interestingly, the proteolytic system-related proteins aminopeptidase N (PepN), aminopeptidase E (PepE), endopeptidase O2 (PepO2), and oligopeptide transport system permease protein (OppC) were observed only on the maps of pH 5.1 and pH 4.5, which was consistent with the presence of angiotensin I-converting enzyme (ACE)-inhibitory peptides VPP and IPP during these 2 growth stages (log phase and stationary phase). These results, combined with a previous study of gene expression of the proteolytic system, led us to conclude that the Opp transport system, pepE, and pepO2 are likely related to the production of ACE-inhibitory peptides.

Cell growth and proteolytic activity of Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus delbrueckii ssp. bulgaricus, and Streptococcus thermophilus in milk as affected by supplementation with peptide fractions.

The present investigation examined the effects of supplementation of milk peptide fractions produced by enzymatic hydrolysis on the fermentation of reconstituted skim milk (RSM). Changes in pH, cell growth, proteolytic activity, and angiotensin-converting enzyme (ACE)-inhibitory activity were monitored during fermentation of RSM by pure cultures of Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus delbrueckii ssp. bulgaricus, and Streptococcus thermophilus. The study showed that supplementation with peptide fractions of different molecular weights did not significantly affect the bacterial growth in RSM. All bacteria showed an increased proteolytic activity in RSM supplemented with large peptides (>10 kDa), and L. helveticus in general exhibited the highest proteolytic activity among the bacteria studied. The ACE-inhibitory activity was observed to be the maximum in RSM supplemented with larger peptides (>10 kDa) for all bacteria. The results suggest that proteolysis by bacteria leads to increased production of ACE-inhibitory peptides compared to the supplemented peptides produced by enzymatic hydrolysis.

Structural elucidation and antioxidant activities of exopolysaccharides from Lactobacillus helveticus MB2-1.

In the present study, the crude polysaccharides (EPS) from Lactobacillus helveticus MB2-1 were fermented with reconstituted whey medium and further fractionated on DEAE-52 cellulose and Sephadex G-100 chromatography to afford three purified fractions of EPS-1, EPS-2 and EPS-3. Then, their preliminary structures and antioxidant activities in vitro were investigated. Three purified EPS fractions (EPS-1, EPS-2 and EPS-3) had similar molecular weight distributions (about 2 × 10(5)Da), and it was composed of galactose, glucose and mannose with a molar ratio of 1.33:2.75:1.00, 1.00:1.43:9.34 and 1.17:1.00:2.96, respectively. In addition, the antioxidant activity of EPS-1, EPS-2 and EPS-3 was evaluated with the in vitro scavenging abilities. The results indicated that both crude and purified EPS showed strong scavenging activities on three kinds of radical and chelating activities on ferrous ion, and their antioxidant activities decreased in the order of crude EPS>EPS-3>EPS-2>EPS-1.

A comprehensive post-market review of studies on a probiotic product containing Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011.

The probiotic preparation Lacidofil® has been commercially available in Europe, Asia and North America since 1995. This product is a combination of two strains, Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011. The strains have been evaluated for safety, identity and mechanisms of probiotic action in vitro, in animal models and human clinical trials. The strains adhered to human epithelial cells, helped to maintain the barrier function and blocked the adhesion of a number of pathogens, allowing them to be cleared from the intestine. The strains also elicited an anti-inflammatory response by down-regulating IL-1ß, IL-8 and TNF-α. In various stress models, the probiotic combination facilitated better coping and outcomes which may be through the maintenance of barrier function and suppressing inflammation. Overall, pre-clinical studies suggest a potential anti-infectious role for the strains and the combination. Clinical studies, primarily in children, have identified Lacidofil as an effective supplement for various gastrointestinal diseases such as antibiotic-associated diarrhoea and acute gastroenteritis. Recent research has also indicated that Lacidofil may be beneficial for individuals with atopic dermatitis or vaginal dysbacteriosis.

Improved growth model for two-stage continuous cultures of Lactobacillus helveticus

An unstructured model for growth and lactic acid production during two-stage continuous cultures of Lactobacillus helveticus was previously developed. The Verlhust model was considered to describe growth kinetics. Production models was based on modified Luedeking-Piret expressions involving an inhibitory effect for the first stage (seed culture) and a nutritional limitation effect for the second stage (culture). To account for the decrease of the biomass concentration observed in the second stage, the dilution rate Dc was replaced by an exponential term of the dilution rate (alpha exp (Dc/beta)) in the growth and product relations. Contrarily to the previous model, the important decrease of the biomass concentration observed at steady state in the second stage at high dilution rates, namely close to wash out, was correctly described by the new model. It also proved to satisfactory describes production data and volumetric productivity.

Effect of fermentation by pure and mixed cultures of Streptococcus thermophilus and Lactobacillus helveticus on isoflavone and B-vitamin content of a fermented soy beverage.

Enhancement of nutritional or vitamin content of foods is commonly touted as a major benefit of probiotics. In this paper, we examined the ability of three probiotic bacteria either alone or in combination to enhance nutritional content. Pure and mixed cultures of Streptococcus thermophilus ST5 and Lactobacillus helveticus R0052 or Bifidobacterium longum R0175 were used to prepare fermented soy beverages. The effects of strain, acidification and mixed cultures on the deconjugation of soy isoflavones were examined. Acidification to pH 4.7 alone resulted in a 7% drop in isoflavone levels. Deconjugation levels varied between the different glucosides. Fermentation by L. helveticus R0052 resulted in the 50% reduction in total glucosides with O-malonyl glucosides being reduced 64%. Fermentation by S. thermophilus ST5 or B. longum R0175 had no significant effect on isoflavone levels. Combining a S. thermophilus strain with a L. helveticus culture reduced the effectiveness of the latter. Fermentation did not significantly modify vitamin B1 or B6 levels.

A dairy bacterium displays in vitro probiotic properties for the pharyngeal mucosa by antagonizing group A streptococci and modulating the immune response.

The probiotic approach represents an alternative strategy in the prevention and treatment of infectious diseases, not only at the intestinal level but also at other sites of the body where the microbiota plays a role in the maintenance of physiological homeostasis. In this context, we evaluated in vitro the potential abilities of probiotic and dairy bacteria in controlling Streptococcus pyogenes infections at the pharyngeal level. Initially, we analyzed bacterial adhesion to FaDu hypopharyngeal carcinoma cells and the ability to antagonize S. pyogenes on FaDu cell layers and HaCat keratinocytes. Due to its promising adhesive and antagonistic features, we studied the dairy strain Lactobacillus helveticus MIMLh5, also through in vitro immunological experiments. First, we performed quantification of several cytokines and measurement of NF-κB activation in FaDu cells. MIMLh5 efficiently reduced the induction of interleukin-6 (IL-6), IL-8, and tumor necrosis factor alpha (TNF-α), in a dose-dependent manner. After stimulation of cells with IL-1ß, active NF-κB was still markedly lowered. Nevertheless, we observed an increased secretion of IL-6, gamma interferon (IFN-γ), and granulocyte-macrophage colony-stimulating factor (GM-CSF) under these conditions. These effects were associated with the ability of MIMLh5 to enhance the expression of the heat shock protein coding gene hsp70. In addition, MIMLh5 increased the GM-CSF/G-CSF ratio. This is compatible with a switch of the immune response toward a TH1 pathway, as supported by our observation that MIMLh5, once in contact with bone marrow-derived dendritic cells, triggered the secretion of TNF-α and IL-2. In conclusion, we propose MIMLh5 as a potential probiotic bacterium for the human pharynx, with promising antagonistic and immunomodulatory properties.