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A well-established rat model of diet-induced metabolic syndrome was used to evaluate the effects of the oral administration of spores or cells of HU16, a carotenoid-producing strain of Bacillus indicus. Symptoms of metabolic syndrome were induced in 90-days old, male Sprague-Dawley rats maintained for eight weeks on a high-fat diet, as previously reported. Parallel groups of animals under the same diet regimen also received a daily dose of 1×1010cells or spores of B. indicus HU16. Cells of strain HU16 were able to reduce symptoms of metabolic syndrome, plasma markers of inflammation and oxidative markers in plasma and liver to levels similar to those observed in rats under a standard diet. HU16 cells did not affect obesity markers or the accumulation of triglycerides in the liver of treated animals. Denaturing gradient gel electrophoresis analysis showed that the oral administration of HU16 cells did not significantly affect the gut microbiota of high fat-fed rats, suggesting that the observed beneficial effects are not due to a reshaping of the gut microbiota but rather to metabolites produced by HU16 cells.

Recent Res. Devel. Bacteriol. 1: 311-330

in Recent Research Developed in Bacteriology

It is now commonly accepted that the intestinal microbiota plays a crucial role in the gut physiology and homeostasis, and that both qualitative and quantitative alterations in the compositions of the gut flora exert profound effects on the host's intestinal cells. In spite of this, the details of the interaction between commensal bacteria and intestinal cells are still largely unknown and only in few cases the molecular mechanisms have been elucidated. Here we analyze the effects of molecules produced and secreted by Lactobacillus gasseri SF1183 on human intestinal HCT116 cells. L. gasseri is a well known species of lactic acid bacteria, commonly associated to the human intestine and SF1183 is a human strain previously isolated from an ileal biopsy of an healthy volunteer. SF1183 produces and secretes, in a growth phase-dependent way, molecule(s) able to drastically interfere with HCT116 cell proliferation. Although several attempts to purify and identify the bioactive molecule(s) have been so far unsuccessful, a partial characterization has indicated that it is smaller than 3 kDa, thermostable and of proteinaceous nature. L. gasseri molecule(s) stimulate a G1-phase arrest of the cell cycle by up-regulation of p21WAF1 rendering cells protected from intrinsic and extrinsic apoptosis. A L. gasseri-mediated reduction of apoptosis and of cell proliferation could be relevant in protecting epithelial barrier integrity and helping in reconstituting tissutal homeostasis.

BACKGROUND: Mucosal infections are a major global health problem and it is generally accepted that mucosal vaccination strategies, able to block infection at their entry site, would be preferable with respect to other prevention approaches. However, there are still relatively few mucosal vaccines available, mainly because of the lack of efficient delivery systems and of mucosal adjuvants. Recombinant bacterial spores displaying a heterologous antigen have been shown to induce protective immune responses and, therefore, proposed as a mucosal delivery system. A non-recombinant approach has been recently developed and tested to display antigens and enzymes. RESULTS: We report that the binding subunit of the heat-labile toxin (LTB) of Escherichia coli efficiently adsorbed on the surface of Bacillus subtilis spores. When nasally administered to groups of mice, spore-adsorbed LTB was able to induce a specific immune response with the production of serum IgG, fecal sIgA and of IFN-gamma in spleen and mesenteric lympho nodes (MLN) of the immunized animals. Dot blotting experiments showed that the non-recombinant approach was more efficient than the recombinant system in displaying LTB and that the efficiency of display could be further increased by using mutant spores with an altered surface. In addition, immunofluorescence microscopy experiments showed that only when displayed on the spore surface by the non-recombinant approach LTB was found in its native, pentameric form. CONCLUSION: Our results indicate that non-recombinant spores displaying LTB pentamers can be administered by the nasal route to induce a Th1-biased, specific immune response. Mutant spores with an altered coat are more efficient than wild type spores in adsorbing the antigen, allowing the use of a reduced number of spores in immunization procedures. Efficiency of display, ability to display the native form of the antigen and to induce a specific immune response propose this non-recombinant delivery system as a powerful mucosal vaccine delivery approach.

Lignocellulosic materials can be used to produce bioethanol. The enzymatic hydrolysis of cellulose is catalyzed by cellulases, a complex system of enzymes that work together synergistically to attack native cellulose releasing glucose and soluble carbohydrates. Generally, these enzymes have low thermal stability and it is quite arduous to recover them from the hydrolyzates for the use in successive batches. Immobilization of cellulases may improve their performances. In the present paper, an epoxy support (Sepabeads® EC­EP403/S) was used as enzymes carrier for the cellulases immobilization. The innovative aspect of this work is the use of this particular support to perform a very easy immobilization­stabilization of commercial cellulase enzymes. The highest immobilization rate was obtained at alkaline conditions (pH 8) and it was promoted by long­term incubation of enzymes in the mixture containing the epoxy support and buffer. The presence of support reduced the chemical deactivation of the enzymes due to the pH of the immobilization procedure. The overall efficiency of the cellulase immobilization was 70%., Proceedings of the 19th European Biomass Conference and Exhibition, 6-10 June 2011, Berlin, Germany, pp. 1756-1758

For several years, international research has been performing the optimization of cellulose enzymatic hydrolysis to produce bioethanol. Enzymes low specific activity and fast deactivation are relevant problems in the development of this process and additional improvements are needed to make bioethanol comparable to traditional fuels. Enzymes immobilization facilitates biocatalyst recovery and reuse. Furthermore, enzymes immobilization avoids autolysis and increase enzyme stability allowing continuous operation for long periods, reducing processes costs and making this process more suitable to industrial scale up. In this paper, commercial cellulase enzymes (Celluclast 1.5 and Novozym 188) immobilization on epoxy Sepabeads® has been investigated. These supports ensure a mild physical adsorption of the enzymes followed by a very fast intramolecular covalent binding with the material epoxy groups. Data show that 60% of loaded Celluclast proteins were adsorbed by the support and that more than 90% of these proteins remained stably linked even after repeated washings. Similar results were obtained with Novozym. Novozym enzyme immobilized was used for ß-glucosidase activity test. The first analyses show that immobilized Novozym activity is comparable with free enzyme. The paper shows preliminary results about immobilized biocatalysts performances., Proceedings of the 18th European Biomass Conference and Exhibition, 3-7 May 2010, Lyon, France, pp. 1440-1442

Introduction: Celiac disease (CD), is the most common food-sensitive enteropathy in humans, caused by the lack of oral tolerance to wheat gluten (1). To study this disease we use a transgenic mouse model expressing the HLA-DQ8 molecule in the absence of endogenous class II genes (2). The adjuvant function Lactobacillus casei is well known (3). In this study we analyzed the effects of oral administration of L. casei in DQ8 transgenic mice following mucosal sensitization with gliadin along with cholera toxin. Methods: DQ8 transgenic mice were administered intragastrically with 500 mg of a chymotryptic digest of gliadin (ct-gliadin) along with 25 ?g of cholera toxin (CT) on days 0, 7 and 14. In someexperiments L. casei (1x1010/dose, four doses/week) was co-administered per os. On day 21 mice were sacrificed. Spleen and MLN were isolated for in vitro assessment of gliadin-specific immunity. Small intestine fragments were collected for RNA analysis and immunomorphometric measurements. Results: Co-administration of L.casei in gliadin-sensitive DQ8 transgenic mice caused a strong increase of the intestinal gliadin-specific cell mediated immune response (SI: 2.75 ± 0.1 vs 4.4 ± 0.03; control vs L. casei; p

Bacillus subtilis spores displaying the tetanus toxin fragment C (TTFC) on their surface have been previously shown to induce the production of specific IgG and secretory IgA in mice immunized through the oral or nasal route. Aim of this study was to analyze whether these spores were also able to induce cellular immunity, and whether such immune response was dependent on spore germination in the animal gastro-intestinal tract (GIT). We first developed a germination defective strain of B. subtilis unable to produce viable cells inside the mouse GIT. Germination-defective and congenic wild-type spores both expressing TTFC on their surface were then used to orally immunize Balb/C mice. Both types of spores induced spleen and mesenteric lymph nodes cell proliferation as well as production of IFNgamma but not of IL-4 and IL-10 in both districts. Our results indicate that recombinant spores preferentially induce a strong cell-mediated immune response with a Th1 phenotype, independently from their ability to germinate in the GIT.

A bacteriocin-producing Bacillus cereus strain was isolated. The bacteriocin, here called cerein, was shown to be active specifically against other B. cereus strains and inactive against all other bacterial species tested. The bacterial activity of cerein was insensitive to organic solvents and nonproteolytic enzymes, partially stable to heat, and active over a wide range of pH values. Direct detection of antimicrobial activity on sodium dodecyl sulfate-polyacrylamide gel suggested an apparent molecular mass of about 9 kDa.

Bacillus subtilis spores displaying the tetanus toxin fragment C (TTFC) on their surface have been previously shown to induce the production of specific IgG and secretory IgA in mice immunized through the oral or nasal route. Aim of this study was to analyze whether these spores were also able to induce cellular immunity, and whether such immune response was dependent on spore germination in the animal gastro-intestinal tract (GIT). We first developed a germination defective strain of B. subtilis unable to produce viable cells inside the mouse GIT. Germination-defective and congenic wild-type spores both expressing TTFC on their surface were then used to orally immunize Balb/C mice. Both types of spores induced spleen and mesenteric lymph nodes cell proliferation as well as production of IFNgamma but not of IL-4 and IL-10 in both districts. Our results indicate that recombinant spores preferentially induce a strong cell-mediated immune response with a Th1 phenotype, independently from their ability to germinate in the GIT.

In a search for bacteria having putative probiotic activity, we screened a collection of food-isolated microorganisms for the ability to survive at low pH in the presence of bile salts and for the production of antimicrobial compounds active against a number of animal pathogens. Among these, we found a strain that we classified as a member of Lactobacillus fermentum sp., and we further investigated its features. This organism was able to adhere to human enterocyte-like (Caco-2) cells with high efficiency as compared to that of a well known indicator strain. Chromatographic analysis indicated that at least two small (less than 3 kDa) factors were involved in mediating the in vitro interaction of L. fermentum with Caco-2 cells. Adhesion activity could be abolished by mild treatment of the bacterial cells in buffer and rescued by incubating them with either the same buffer after its use in the treatment or with chromatographic fractions containing each of the two factors, which indicated that these factors were loosely associated with the cell wall and that each of them was sufficient to warrant the adhesiveness of L. fermentum to Caco-2 cells. These data are suggestive of a novel mechanism of bacterial adhesion to epithelial cells.