Neuroprotective effects of probiotics bacteria on animal model of Parkinson's disease induced by 6-hydroxydopamine: A behavioral, biochemical, and histological study.

Parkinson's disease (PD) is an age-associated, progressive, and common neurodegenerative disorder. It is characterized by dopaminergic neuron degeneration in the substantia nigra pars compacta. The involvement of oxidative stress, inflammation, and dysbiosis in PD has been confirmed and probiotics also have the ability to regulate the mentioned mechanisms. Here, we assessed probiotics supplementation effects on experimental model of PD. Thirty Male Wistar rats were divided into three groups for a 14-day treatment. It was shown that a mixture of probiotics containing Lactobacillus acidophilus, Bifidobacterium bifidum, Lactobacillus reuteri, and Lactobacillus fermentum could improve rotational behavior, cognitive function, lipid peroxidation, and neuronal damage in the group received probiotic supplementation compared to the other groups (P < 0001, P < .001, and P = .026, respectively). Taken together, these findings revealed that probiotics supplementation could be an appropriate complementary treatment for PD.

Selection, Characterization and Interaction Studies of a DNA Aptamer for the Detection of Bifidobacterium bifidum.

A whole-bacterium-based SELEX (Systematic Evolution of Ligands by Exponential Enrichment) procedure was adopted in this study for the selection of an ssDNA aptamer that binds to Bifidobacterium bifidum. After 12 rounds of selection targeted against B. bifidum, 30 sequences were obtained and divided into seven families according to primary sequence homology and similarity of secondary structure. Four FAM (fluorescein amidite) labeled aptamer sequences from different families were selected for further characterization by flow cytometric analysis. The results reveal that the aptamer sequence CCFM641-5 demonstrated high-affinity and specificity for B. bifidum compared with the other sequences tested, and the estimated Kd value was 10.69 ± 0.89 nM. Additionally, sequence truncation experiments of the aptamer CCFM641-5 led to the conclusion that the 5'-primer and 3'-primer binding sites were essential for aptamer-target binding. In addition, the possible component of the target B. bifidum, bound by the aptamer CCFM641-5, was identified as a membrane protein by treatment with proteinase. Furthermore, to prove the potential application of the aptamer CCFM641-5, a colorimetric bioassay of the sandwich-type structure was used to detect B. bifidum. The assay had a linear range of 104 to 107 cfu/mL (R² = 0.9834). Therefore, the colorimetric bioassay appears to be a promising method for the detection of B. bifidum based on the aptamer CCFM641-5.

Improvement of loperamide-induced slow transit constipation by Bifidobacterium bifidum G9-1 is mediated by the correction of butyrate production and neurotransmitter profile due to improvement in dysbiosis.

A treatment option for constipation that improves the quality of life is needed since available laxatives do not effectively improve the quality of life in patients with constipation. A significant association between gut dysbiosis and constipation is recognized, suggesting that probiotics may be an important option for management of constipation. The underlying mechanism by which probiotics improve constipation remains unclear. In this study, we aimed to evaluate the effects of the probiotic Bifidobacterium bifidum G9-1 (BBG9-1) on loperamide-induced delayed colonic transit constipation and to elucidate its mechanism of action. First, the effect of BBG9-1 was evaluated in a rat model of constipation induced by subcutaneous administration of loperamide. BBG9-1 improved constipation parameters (number of feces, fecal water content, and fecal hardness) in constipated rats. Next, the relationship of organic acids and neurotransmitters to gut microbiota was investigated. BBG9-1 improved dysbiosis and prevented a decrease in butyric acid concentration in the gut, increased serum serotonin, and suppressed an increase in dopamine and a decrease in acetylcholine in serum. Further, an increase in the expression level of tryptophan hydroxylase 1, a 5-HT-synthetizing enzyme, was observed. These results suggest that BBG9-1 improves dysbiosis, which results in an increase in organic acids and improvement of neurotransmission. These actions may increase intestinal mobility, finally leading to alleviating constipation. The probiotic BBG9-1 may, therefore, be a potential option for the treatment of constipation.

Selective Synthesis of Galactooligosaccharides Containing ß(1→3) Linkages with ß-Galactosidase from Bifidobacterium bifidum (Saphera).

The transglycosylation activity of a novel commercial ß-galactosidase from Bifidobacterium bifidum (Saphera) was evaluated. The optimal conditions for the operation of this enzyme, measured with o-nitrophenyl-ß-d-galactopyranoside, were 40 °C and pH around 6.0. Although at low lactose concentrations the property of this enzyme was basically hydrolytic, an increase of lactose concentration to 400 g/L resulted in a significant formation (107.2 g/L, 27% yield) of prebiotic galactooligosaccharides (GOS). The maximum amount of GOS was obtained at a lactose conversion of approximately 90%, which contrasts with other ß-galactosidases, for which the highest GOS yield is achieved at 40-50% lactose conversion. Using high-performance anion-exchange chromatography with pulsed amperometric detection, semipreparative high-performance liquid chromatography-hydrophilic interaction liquid chromatography, mass spectrometry, and 1D and 2D NMR, we determined the structure of most of the GOS synthesized by this enzyme. The main identified products were Gal-ß(1→3)-Gal-ß(1→4)-Glc (3'-O-ß-galactosyl-lactose), Gal-ß(1→6)-Glc (allolactose), Gal-ß(1→3)-Glc (3-galactosyl-glucose), Gal-ß(1→3)-Gal (3-galactobiose), and the tetrasaccharide Gal-ß(1→3)-Gal-ß(1→3)-Gal-ß(1→4)-Glc. In general, B. bifidum ß-galactosidase showed a tendency to form ß(1→3) linkages followed by ß(1→6) and more scarcely ß(1→4).

The Use of Probiotic-Loaded Single- and Double-Layered Microcapsules in Cake Production.

To date, the probiotic product development studies have mostly focused on dairy-based foods. However, endowing bakery products with probiotic properties not only provides a variety in food selection but would also potentially improve public health when the consumption rates are taken into consideration. This study aimed to incorporate single- and double-layered microcapsules containing Saccharomyces boulardii, Lactobacillus acidophilus, and Bifidobacterium bifidum, produced by spray drying and chilling, in cake production. Microcapsules were added after baking to the three different types of cakes (cream-filled, marmalade-filled, and chocolate-coated). Additionally, the microcapsules were injected into the center of the cake mix and baked at 200 °C for 20 min, for plain cake only. After baking of plain cakes, the count of S. boulardii and L. acidophilus as determined in the double-layered microcapsules produced by spray chilling was 2.9 log cfu/g. The survivability rates of S. boulardii and L. acidophilus were also determined as 67.4 and 70.7% in this microcapsule, respectively. However, there were no viable B. bifidum detected after baking. The free forms of these probiotics did not survive in any plain cake experiments. Single-layered microcapsules produced by spray chilling provided a better protective effect on the probiotics in cream-filled and marmalade-filled cake samples during storage, particularly the cream-filled cakes. This study showed that combined spray chilling and spray drying microencapsulation techniques (double-layered microcapsules) could increase the survivability of probiotic microorganisms during the cake baking process. During storage, the cake samples had a near neutral pH value, and the textural properties deteriorated due to staling. However, cake staling had a limited effect on the sensorial attributes of the cakes and the samples could be readily consumed after storage for 90 days.

In-vitro GIT Tolerance of Microencapsulated Bifidobacterium bifidum ATCC 35914 Using Polysaccharide-Protein Matrix.

Longevity of probiotic is the main concern for getting maximum benefits when added in food product. Bifidobacterium, a probiotic, tends to lose its viability during gastrointestinal track (GIT) transit and storage of food. Their viability can be enhanced through microencapsulation technology. In this study, Bifidobacterium bifidum (B. bifidum) ATCC 35914 was encapsulated by using two experimental plans. In the first plan, chitosan (CH) at 0.6, 0.8, and 1.0% and sodium alginate (SA) at 4, 5, and 6% were used. Based on encapsulation efficiency, 6% sodium alginate and 0.8% chitosan were selected for single coating of the bacteria, and the resulting micro beads were double coated with different concentrations (5, 7.5, and 10%) of whey protein concentrate (WPC) in the second plan. Encapsulation efficiency and GIT tolerance were determined by incubating the micro beads in simulated gastrointestinal juices (SIJ) at variable pH and exposure times, and their release (liberation of bacterial cells) profile was also observed in SIJ. The microencapsulated bacterial cells showed significantly (P < 0.01) higher viability as compared to the unencapsulated (free) cells during GIT assay. The double-coated micro beads SA 6%-WPC 5% and CH 0.8%-WPC 5% were proven to have the higher survival at pH 3.0 after 90 min of incubation time and at pH 7.0 after 3-h exposure in comparison to free cells in simulated conditions of the stomach and intestine, respectively. Moreover, double coating with whey protein concentrate played a significant role in the targeted (106-9 CFU/mL) delivery under simulated intestinal conditions.

Bifidobacterium bifidum presents on the cell surface a complex mixture of glucans and galactans with different immunological properties.

The chemical structure of cell surface polysaccharides isolated from Bifidobacterium bifidum strain PRI1, an important member of the gut microbiota of breast-fed infants, has been elucidated by chemical and NMR spectroscopy analysis. Results demonstrated that the bacterium produces a complex mixture of polysaccharides that could be classified in two main groups: a phospho-glycero-ß-galactofuranan, PGßG, and a mixture composed of four neutral polysaccharides named as (CSGG), composed of ß-(1 → 6)-glucan, ß-(1 → 4)-galactan, ß-(1 → 6)-galactan, ß-galactofuranan and starch. These two fractions exerted different immune responses when assayed on dendritic cells: PGßG enhanced pro-inflammatory immune responses by increasing interferon-γ levels while CSGG induced immunosuppressive regulatory T cells and interleukin-10. These findings demonstrate that bacterial polysaccharides have a distinct role depending on their chemical structure in regulation of the host/bacterium interaction. Our findings suggest that polysaccharides may differentially regulate the host immunity depending on the composition of this complex mixture, either enhancing immunity or inducing immune tolerance.

Microencapsulation of Bifidobacterium bifidum BB01 by xanthan-chitosan: preparation and its stability in pure milk.

Xanthan-chitosan (XC) and xanthan-chitosan-xanthan (XCX) were employed for microencapsulation of Bifidobacterium bifidum BB01 using extrusion technique. To optimize the process of B. bifidum BB01 microcapsules based on XC hydrogels, response surface methodology was employed to obtain the best possible combination of chitosan concentration, xanthan-B. bifidum BB01 mixture (XBM)/chitosan, stirring time for the maximum viable count and encapsulation yield. The optimum conditions were: chitosan concentration of 0.84 g·mL-1, XBM/chitosan of 1:9.0, stirring time of 60 min with high viable count and encapsulation yield of 1.52 ± 0.15 × 1010 CFU·g-1, 90 ± 0.65%, respectively. In addition, the effective encapsulation system (XC and XCX) resulted in improvement in survival of B. bifidum BB01 compared to non-encapsulated cells during 3 weeks storage at 4 and 25 °C in pure milk.

Linear α-(1 → 6)-d-glucan from Bifidobacterium bifidum BIM В-733D is low molecular mass biopolymer with unique immunochemical properties.

Role of microorganisms in induction of/protection from autoimmune diseases is proven though molecular mechanisms and bacterial/viral/yeast biopolymers responsible for these effects are in the research stage. Autoantobodies (AAbs) to thyroid peroxidase (anti-TPO) and thyroglobulin (anti-Tg) as well as AAbs to transglutaminase 2 (anti-TG2) and antibodies to gliadins (anti-gliadins) are serological markers of autoimmune thyroid disease and celiac disease, respectively, and players in pathogenesis of these autoimmune diseases. In current study, biopolymer of Bifidobacterium bifidum BIM В-733D that interacts selectively with anti-gliadins (Bb-Ganti-gliadins) was isolated by affinity chromatography with anti-gliadins, purified by size exclusion chromatography on TSK 40 gel and identified by NMR as linear α-(1 → 6)-d-glucan with molecular mass about 5000 Da. It was proven that compounds Bb-Ganti-gliadins and Bb-Ganti-TPO/Bb-Ganti-Tg isolated early from the same strain [Kiseleva, E. P. et al., Benef Microbes.2013, 4, 375 -391] are the same substance designated GBb. Its unique immunochemical property is the ability to interact selectively with anti-TPO, anti-Tg, anti-TG2 and anti-gliadins in presence of no less than 10-fold excess of total immunoglobulins of class G (tIgG), as it was proven by ELISA. Synthesis of GBb-bovine serum albumin (GBb-BSA) conjugate is an example of increasing the reliability and reproducibility of ELISA results by mediated immobilization of a polysaccharide covalently attached to a well-adsorbed protein. Taking into account that there are population of bispecific anti-gliadins (anti-gliadins and anti-TG2 simultaneously) we regard our data as first argument in favor of hypothesis that GBb differentiates between human AAbs per se and other human Ig (e.g. antibodies to antigens of infectious agents) due to its binding with a yet unidentified site which is present in the molecules of all AAbs (independently on their specificity) and absent in other human Igs.

Immunomodulatory effects of Bifidobacterium bifidum 231 on trinitrobenzenesulfonic acid-induced ulcerative colitis in rats.

AIM: Ulcerative colitis and Crohn's disease are two important chronic Inflammatory bowel diseases (IBD) characterized by prominent intestinal inflammation. Probiotics are the bacteria that promote the host health by its immunomodulatory activity. The present study investigated the correlation between in vitro adhesion and immunomodulatory properties, and to assess the therapeutic potential of Bifidobacterium bifidum 231 (BIF 231), a new strain of probiotic in ulcerative colitis in rats. METHODS: In vitro adhesion assays and immunomodulatory effect of BIF 231 on interleukins (IL-1ß and IL-10) in IEC-6 cell lines were quantified by gram staining, scanning electron microscopy and q-PCR respectively. Colitis was induced by intra-rectal instillation of trinitrobenzenesulfonic acid. Colitis was evaluated by alterations in colon gross morphology, histologically and biochemically. Colonic interleukin-1ß (IL-1ß) and interleukin-10 (IL-10) mRNA and protein expression were assessed by q-PCR, ELISA and western blot. RESULTS: BIF 231 showed better adhesion and immunomodulation by up-regulating IL-10 levels in IEC-6 cell lines. In vivo studies with treatment of BIF 231 (1.4×1011 CFU/rat/day) revealed anti-inflammatory effects both macroscopically and histologically. BIF 231 lowered TBARS, nitric oxide and augmented GSH levels. BIF 231 treatment to colitic rats down regulated IL-1ß levels with concurrent increase in IL-10 levels. CONCLUSIONS: BIF 231 exerted beneficial in vitro adhesion and immunomodulatory properties which facilitated the recovery of the damaged tissue in TNBS-induced colitis.