Intestinal Isolate Research Articles

Development of a time-dependent oral colon delivery system of anaerobic Odoribacter splanchnicus for bacteriotherapy

Odoribacter (O.) splanchnicus is an anaerobic member of the human intestinal microbiota. Its decrease in abundance has been associated with inflammatory bowel disease (IBD), non-alcoholic fatty liver, and cystic fibrosis. Considering the anti-inflammatory properties of O. splanchnicus and its possible use for IBD, intestinal isolate O. splanchnicus 57 was here formulated for oral colonic release based on a time-dependent strategy. Freeze-drying protocol was determined to ensure O. splanchnicus 57 viability during the process. Disintegrating tablets, containing the freeze-dried O. splanchnicus 57, were manufactured by direct compression and coated by powder-layering technique with hydroxypropyl methylcellulose (Methocel™ E50) in a tangential-spray fluid bed. Eudragit® L was then applied by spray-coating in a top-spray fluid bed. Double-coated tablets were tested for release, showing gastric resistance properties and, as desired, lag phases of reproducible duration prior to release in phosphate buffer pH 6.8. The cell viability and anti-inflammatory activity of the strain were assessed after the main manufacturing steps. While freeze-drying did not affect bacterial viability, the tableting and coating processes were more stressful. Nonetheless, O. splanchnicus 57 cells survived manufacturing and the final formulations had 106-107 CFU/g of viable cells. The strain kept its anti-inflammatory properties after tableting and coating, reducing Escherichia coli lipopolysaccharide-induced interleukin-8 cytokine release from HT-29 cells. Overall, O. splanchnicus 57 strain was formulated successfully for oral colon delivery, opening new ways to formulate pure cultures of single anaerobic strains or mixtures for oral delivery.

Sporulation efficiency and spore quality in a human intestinal isolate of Bacillus cereus

Bacteria classified as Bacillus cereus sensu stricto cause two different type of gastrointestinal diseases associated with food poisoning. Outbreaks of this opportunistic pathogen are generally due to the resistance of its spores to heat, pH and desiccation that makes hard their complete inactivation from food products. B. cereus is commonly isolated from a variety of environments, including intestinal samples of infected and healthy people. We report the genomic and physiological characterization of MV19, a human intestinal strain closely related (ANI value of 98.81%) to the reference strain B. cereus ATCC 14579. MV19 cells were able to grow in a range of temperatures between 20 and 44 °C. At the optimal temperature the sporulation process was rapidly induced and mature spores efficiently released, however these appeared structurally and morphologically defective. At the sub-optimal growth temperature of 25 °C sporulation was slow and less efficient but a high total number of fully functional spores was produced. These results indicate that the reduced rapidity and efficiency of sporulation at 25 °C are compensated by a high quality and quantity of released spores, suggesting the relevance of different performances at different growth conditions for the adaptation of this bacterium to diverse environmental niches.

Towards the isolation of more robust next generation probiotics: The first aerotolerant Bifidobacterium bifidum strain

This work reports on the first described aerotolerant Bifidobacterium bifidum strain, Bifidobacterium bifidum IPLA60003, which has the ability to form colonies on the surface of agar plates under aerobic conditions, a weird phenotype that to our knowledge has never been observed in B. bifidum. The strain IPLA60003 was generated after random UV mutagenesis from an intestinal isolate. It incorporates 26 single nucleotide polymorphisms that activate the expression of native oxidative-defense mechanisms such as the alkyl hydroxyperoxide reductase, the glycolytic pathway and several genes coding for enzymes involved in redox reactions. In the present work, we discuss the molecular mechanisms underlying the aerotolerance phenotype of B. bifidum IPLA60003, which will open new strategies for the selection and inclusion of probiotic gut strains and next generation probiotics into functional foods.

Natural Competence for Transformation in Lactobacili

Natural Competence for Transformation in Lactobacili

Characterisation of EFV12 a bio-active small peptide produced by the human intestinal isolate Lactobacillus gasseri SF1109.

EFV12 is a small bioactive peptide produced by Lactobacillus gasseri SF1109, a human intestinal isolate with probiotic features. In this study, EFV12 antimicrobial and anti-inflammatory properties are characterised. In particular, we propose a possible mechanism of action for EFV12 involving bacterial membranes targeting. Moreover, we show that this small peptide is able to bind lipopolysaccharides (LPS) and to counteract its inflammatory insult preventing LPS action on Toll-like receptor 4, thus interfering with extracellular signal-regulated kinase, p38 and Jun N-terminal kinase, mitogen-activated protein kinases signalling pathways. Altogether these observations suggest that the bioactive peptide EFV12 is a good candidate to promote L. gasseri induced gut homeostasis and counteracting intestinal pathogens.

MtcB, a member of the MttB superfamily from the human gut acetogen Eubacterium limosum, is a cobalamin-dependent carnitine demethylase

The trimethylamine methyltransferase MttB is the first described member of a superfamily comprising thousands of microbial proteins. Most members of the MttB superfamily are encoded by genes that lack the codon for pyrrolysine characteristic of trimethylamine methyltransferases, raising questions about the activities of these proteins. The superfamily member MtcB is found in the human intestinal isolate Eubacterium limosum ATCC 8486, an acetogen that can grow by demethylation of l-carnitine. Here, we demonstrate that MtcB catalyzes l-carnitine demethylation. When growing on l-carnitine, E. limosum excreted the unusual biological product norcarnitine as well as acetate, butyrate, and caproate. Cellular extracts of E. limosum grown on l-carnitine, but not lactate, methylated cob-(I)alamin or tetrahydrofolate using l-carnitine as methyl donor. MtcB, along with the corrinoid protein MtqC and the methylcorrinoid:tetrahydrofolate methyltransferase MtqA, were much more abundant in E. limosum cells grown on l-carnitine than on lactate. Recombinant MtcB methylates either cob(I)alamin or Co(I)-MtqC in the presence of l-carnitine and, to a much lesser extent, γ-butyrobetaine. Other quaternary amines were not substrates. Recombinant MtcB, MtqC, and MtqA methylated tetrahydrofolate via l-carnitine, forming a key intermediate in the acetogenic Wood-Ljungdahl pathway. To our knowledge, MtcB methylation of cobalamin or Co(I)-MtqC represents the first described mechanism of biological l-carnitine demethylation. The conversion of l-carnitine and its derivative γ-butyrobetaine to trimethylamine by the gut microbiome has been linked to cardiovascular disease. The activities of MtcB and related proteins in E. limosum might demethylate proatherogenic quaternary amines and contribute to the perceived health benefits of this human gut symbiont.

MtpB, a member of the MttB superfamily from the human intestinal acetogen Eubacterium limosum, catalyzes proline betaine demethylation

The trimethylamine methyltransferase MttB is the founding member of a widely distributed superfamily of microbial proteins. Genes encoding most members of the MttB superfamily lack the codon for pyrrolysine that distinguishes previously characterized trimethylamine methyltransferases, leaving the function(s) of most of the enzymes in this superfamily unknown. Here, investigating the MttB family member MtpB from the human intestinal isolate Eubacterium limosum ATCC 8486, an acetogen that excretes N-methyl proline during growth on proline betaine, we demonstrate that MtpB catalyzes anoxic demethylation of proline betaine. MtpB along with MtqC (a corrinoid protein) and MtqA (a methylcorrinoid:tetrahydrofolate methyltransferase) was much more abundant in E. limosum cells grown on proline betaine than on lactate. We observed that recombinant MtpB methylates Co(I)-MtqC in the presence of proline betaine and that other quaternary amines are much less preferred substrates. MtpB, MtqC, and MtqA catalyze tetrahydrofolate methylation with proline betaine, thereby forming a key intermediate in the Wood-Ljungdahl acetogenesis pathway. To our knowledge, MtpB methylation of Co(I)-MtqC for the subsequent methylation of tetrahydrofolate represents the first described anoxic mechanism of proline betaine demethylation. The activities of MtpB and associated proteins in acetogens or other anaerobes provide a possible mechanism for the production of N-methyl proline by the gut microbiome. MtpB's activity characterized here strengthens the hypothesis that much of the MttB superfamily comprises quaternary amine-dependent methyltransferases.

Lactobacillus rhamnosus LRB mediated inhibition of oral streptococci.

Lactobacillus rhamnosus is a lactic acid bacterium with a diverse ecological habitat. We recently isolated a L.rhamnosus strain (LRB) from a healthy baby-tooth that had naturally fallen out. We determined the whole genome sequence of LRB and found that the isolate is closely genetically related to an intestinal isolate, L.rhamnosus GG (ATCC 53103). However, the LRB genome had lost about a 75-kb segment and undergone a genomic rearrangement. We assessed LRB's capacity to survive in the gut environment, at least temporarily. We found that LRB, like the intestinal isolate ATCC 53103, showed resistance to low pH but sensitive to bile salt. Surprisingly, we found that this oral isolate LRB showed strong antimicrobial activity against a variety of oral streptococci including Streptococcus mutans. The production of antimicrobial activity is dependent on media composition since some media supported the production while others did not. The production of antimicrobial activity is also dependent on growth temperature, with optimal production at 37°C. The antimicrobial activity was not restricted to streptococci, but effective against a variety of organisms, including ESKAPE pathogens.

The potency of the broad\xa0spectrum bacteriocin, bactofencin A, against staphylococci is highly dependent on primary structure, N-terminal charge and disulphide formation

Bactofencin A is a novel class IId bacteriocin, produced by the intestinal isolate Lactobacillus salivarius DPC6502, which has potent activity against medically significant pathogens including Staphylococcus aureus. This bacteriocin is unusual in that it has a highly cationic N terminus and a single disulfide bond between Cys7 and Cys22, resulting in a large C terminal loop. In this study, a library of synthetic bactofencin A variants were screened against the mastitis isolate, S. aureus DPC5246, to identify key residues responsible for activity. It was apparent that substituting either cysteine of the disulfide bond with either serine or alanine significantly reduced the activity of the bacteriocin, confirming the importance of the C terminal loop. Substituting N terminal amino acids with alanine had no effect on activity, whereas sequential removal of the N terminal positively charged residues resulted in an increasingly inactive peptide. A complete (synthetic) alanine scanning analysis revealed that the residues between Val9 and Gly17 were most affected by substitution suggesting that this area has a major influence on the potency of the bacteriocin. Substituting residues in the loop region between Cys7 and Cys22 for D-amino acid equivalents had a more detrimental effect on activity than L-alanine substitutions. Specifically Y10A, N11A, P15A and T16A are active at 4, 16, 1 and 16 μM respectively while their D equivalents were inactive at 1000 μM, the highest concentration tested. Ultimately, this study identifies the critical features in the primary structure of the bacteriocin which gives it such potent activity against pathogenic staphylococci.

Pharmacological Ascorbate as an Adjuvant for Enhancing Radiation-Chemotherapy Responses in Gastric Adenocarcinoma.

Gastric adenocarcinoma most often presents at an advanced stage and overall five-year survival of ∼30%. Pharmacological ascorbate (high-dose IV ascorbate) has been proposed as a promising nontoxic adjuvant to standard radio-chemotherapies in several cancer types. In the current study, pharmacological ascorbate (0.5-2 m M) caused a dose-dependent decrease (70-85% at 2 m M) in clonogenic survival of gastric adenocarcinoma cells (AGS and MNK-45), but was relatively nontoxic to a small intestinal epithelial nonimmortalized human cell isolate (FHs 74 Int). The addition of pharmacological ascorbate (1 m M) to standard radio-chemotherapies [i.e., 5-FU (5 μ M); cisplatin (0.5 μ M); irinotecan (2.5 μ M); carboplatin (5 μ M); paclitaxel (2-4 n M); and X rays (1.8 Gy)] also potentiated gastric cancer clonogenic cell killing [additional decreases were noted with: ascorbate plus 5-FU/radiation (1%); ascorbate plus cisplatin/irinotecan (9-19%); and ascorbate plus paclitaxel/carboplatin (6-7%)]. The gastric cancer cell toxicity and chemosensitization seen with pharmacological ascorbate was dependent on H2O2 and the presence of catalytic metal ions. In addition, pharmacological ascorbate dosing resulted in a concentration-dependent decrease (64% at 20 m M, P ≤ 0.0001) in cancer cell invasion and migration that was inhibited by catalase. Finally, pharmacological ascorbate significantly increased the overall survival of mice with gastric cancer xenografts when used in combination with paclitaxel, carboplatin and radiation ( P = 0.019). These results demonstrate that pharmacological ascorbate is selectively cytotoxic to gastric adenocarcinoma cells (relative to normal intestinal epithelial cells) by a mechanism involving H2O2 and redox active metal ions. Furthermore, pharmacological ascorbate significantly enhances gastric cancer xenograft responses to radio-chemotherapy as well as inhibiting tumor cell migration and invasiveness. Overall, these results support the hypothesis that pharmacological ascorbate can be used as an adjuvant with standard-of-care radio-chemotherapies for the treatment of gastric adenocarcinomas.

Ex Vivo and In Vivo Mice Models to Study Blastocystis spp. Adhesion, Colonization and Pathology: Closer to Proving Koch's Postulates.

Blastocystis spp. are widely prevalent extra cellular, non-motile anerobic protists that inhabit the gastrointestinal tract. Although Blastocystis spp. have been associated with gastrointestinal symptoms, irritable bowel syndrome and urticaria, their clinical significance has remained controversial. We established an ex vivo mouse explant model to characterize adhesion in the context of tissue architecture and presence of the mucin layer. Using confocal microscopy with tissue whole mounts and two axenic isolates of Blastocystis spp., subtype 7 with notable differences in adhesion to intestinal epithelial cells (IEC), isolate B (ST7-B) and isolate H (more adhesive, ST7-H), we showed that adhesion is both isolate dependent and tissue trophic. The more adhesive isolate, ST7-H was found to bind preferentially to the colon tissue than caecum and terminal ileum. Both isolates were also found to have mucinolytic effects. We then adapted a DSS colitis mouse model as a susceptible model to study colonization and acute infection by intra-caecal inoculation of trophic Blastocystis spp.cells. We found that the more adhesive isolate ST7-H was also a better colonizer with more mice shedding parasites and for a longer duration than ST7-B. Adhesion and colonization was also associated with increased virulence as ST7-H infected mice showed greater tissue damage than ST7-B. Both the ex vivo and in vivo models used in this study showed that Blastocystis spp. remain luminal and predominantly associated with mucin. This was further confirmed using colonic loop experiments. We were also successfully able to re-infect a second batch of mice with ST7-H isolates obtained from fecal cultures and demonstrated similar histopathological findings and tissue damage thereby coming closer to proving Koch’s postulates for this parasite.

Draft Genome Sequence of Lactobacillus crispatus JCM5810, Which Can Reduce Campylobacter jejuni Colonization in Chicken Intestine.

We present the 2.05-Mb draft genome sequence of Lactobacillus crispatus JCM5810, a chicken intestinal isolate with the ability to reduce Campylobacter jejuni colonization in chickens. The genome sequence will provide insights on the probiotic mechanisms of L. crispatus JCM5810.

Characterization of flagellins isolated from a highly motile strain of Lactobacillus agilis.

BackgroundMost lactic acid bacteria are non-motile but some of them are flagellated and exhibit motility. So far, motile lactobacilli have rarely been studied, and characteristics of their flagellins are poorly understood. In this study, a highly motile strain of Lactobacillus agilis was recruited for transcriptional analysis and characterization of its flagellins.ResultsUnlike another motile lactic acid bacteria of intestinal isolate, Lactobacillus ruminis, flagellar filaments of the L. agilis strain probably consist of two homologous but distinct flagellins. Glycosylation of the flagellar filaments and their resistance to heat, acid and SDS were also observed. The immunological activity of the flagellins was evaluated through the stimulation of Caco-2 cells. The results show that TLR5-stimulating activity of the protein is attenuated, likely due to an incomplete TLR5-recognition site.ConclusionsThe flagella filaments of L. agilis BKN88 consist of two homologous glycosylated flagellins, which likely have an incomplete TLR5-recognition site. The characteristics of the flagellin are presumably a consequence of adaptation as a commensal microbe in the gastrointestinal tract.

Physicochemical and biosorption properties of novel exopolysaccharide produced by Enterococcus faecalis

Duck intestinal homogenate isolate Enterococcus faecalis DU10 produced exopolysaccharide (EPS) (840 mg/L) in MRS medium (35 °C, pH 6.5), was characterized for its physicochemical, biosorption, antioxidant, antibiofilm and rheological properties. HPTLC analysis revealed fructose and arabinose are as major, xylose and mannose are as minor constituents of EPS. And further, this EPS DU10 was characterized through FTIR, 1H and 13C NMR. EPS DU10 efficiently adsorbed heavy metals Zn(II) (515.78 ± 0.7 mg/g), Pb(II) (424.42 ± 0.9 mg/g), Cd(II) (334.4 ± 14 mg/g), and Cu(II) (232.9 ± 0.3 mg/g). EPS DU10 exhibits broad spectrum of biofilm inhibition for bacterial pathogens and good in vitro antioxidant properties by scavenging free radicals at very low concentration (0.1 mg/mL). Along with these properties, high viscosity of EPS makes it suitable for pharmaceutical and industrial applications.

The pan-genome of Lactobacillus reuteri strains originating from the pig gastrointestinal tract.

BackgroundLactobacillus reuteri is a gut symbiont of a wide variety of vertebrate species that has diversified into distinct phylogenetic clades which are to a large degree host-specific. Previous work demonstrated host specificity in mice and begun to determine the mechanisms by which gut colonisation and host restriction is achieved. However, how L. reuteri strains colonise the gastrointestinal (GI) tract of pigs is unknown.ResultsTo gain insight into the ecology of L. reuteri in the pig gut, the genome sequence of the porcine small intestinal isolate L. reuteri ATCC 53608 was completed and consisted of a chromosome of 1.94 Mbp and two plasmids of 138.5 kbp and 9.09 kbp, respectively. Furthermore, we generated draft genomes of four additional L. reuteri strains isolated from pig faeces or lower GI tract, lp167-67, pg-3b, 20-2 and 3c6, and subjected all five genomes to a comparative genomic analysis together with the previously completed genome of strain I5007. A phylogenetic analysis based on whole genomes showed that porcine L. reuteri strains fall into two distinct clades, as previously suggested by multi-locus sequence analysis. These six pig L. reuteri genomes contained a core set of 1364 orthologous gene clusters, as determined by OrthoMCL analysis, that contributed to a pan-genome totalling 3373 gene clusters. Genome comparisons of the six pig L. reuteri strains with 14 L. reuteri strains from other host origins gave a total pan-genome of 5225 gene clusters that included a core genome of 851 gene clusters but revealed that there were no pig-specific genes per se. However, genes specific for and conserved among strains of the two pig phylogenetic lineages were detected, some of which encoded cell surface proteins that could contribute to the diversification of the two lineages and their observed host specificity.ConclusionsThis study extends the phylogenetic analysis of L. reuteri strains at a genome-wide level, pointing to distinct evolutionary trajectories of porcine L. reuteri lineages, and providing new insights into the genomic events in L. reuteri that occurred during specialisation to their hosts. The occurrence of two distinct pig-derived clades may reflect differences in host genotype, environmental factors such as dietary components or to evolution from ancestral strains of human and rodent origin following contact with pig populations.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2216-7) contains supplementary material, which is available to authorized users.

Nisin H Is a New Nisin Variant Produced by the Gut-Derived Strain Streptococcus hyointestinalis DPC6484.

Accumulating evidence suggests that bacteriocin production represents a probiotic trait for intestinal strains to promote dominance, fight infection, and even signal the immune system. In this respect, in a previous study, we isolated from the porcine intestine a strain of Streptococcus hyointestinalis DPC6484 that displays antimicrobial activity against a wide range of Gram-positive bacteria and produces a bacteriocin with a mass of 3,453 Da. Interestingly, the strain was also found to be immune to a nisin-producing strain. Genome sequencing revealed the genetic determinants responsible for a novel version of nisin, designated nisin H, consisting of the nshABTCPRKGEF genes, with transposases encoded between nshP and nshR and between nshK and nshG. A similar gene cluster is also found in S. hyointestinalis LMG14581. Notably, the cluster lacks an equivalent of the nisin immunity gene, nisI. Nisin H is proposed to have the same structure as the prototypical nisin A but differs at 5 amino acid positions-Ile1Phe (i.e., at position 1, nisin A has Ile while nisin H has Phe), Leu6Met, Gly18Dhb (threonine dehydrated to dehydrobutyrine), Met21Tyr, and His31Lys--and appears to represent an intermediate between the lactococcal nisin A and the streptococcal nisin U variant of nisin. Purified nisin H inhibits a wide range of Gram-positive bacteria, including staphylococci, streptococci, Listeria spp., bacilli, and enterococci. It represents the first example of a natural nisin variant produced by an intestinal isolate of streptococcal origin.

Purification and Characterization of Halophilic Alkaline Lipase from Halobacillus sp.

An extracellular lipase produced by marine fish intestinal isolate Halobacillus sp. AP-MSU 8 was purified and characterized. The lipase was purified to homogeneity using ammonium sulphate precipitation, DEAE-Sepharose anion exchange chromatography and Sephadex G-75 gel filtration chromatography. The overall purification protocols resulted in 25 % yield of lipase with 10.6-fold. The SDS-PAGE (12 %) analysis of purified lipase resulted in the molecular mass of the purified lipase as 25-kDa. The optimum pH and temperature required for maximum activity of purified lipase was 9.0 and 40 °C respectively. Also, this lipase is halo tolerant and requires 2.5 M NaCl for maximum activity. The activity of the purified lipase was more in the presence of BaCl2 and MgSO4, and in contrast the enzyme activity was totally inhibited in the presence of ZnSO4 and ZnCl2. The surfactants such as polyethylene glycol and Tween 20 enhanced the lipase activity. Likewise 10 % concentration of organic solvents such as benzene and acetone stimulated the lipase activity, whereas at 20 % concentration all the tested solvents inhibited the lipase activity.

Lactobacillus helveticus: importance in food and health.

Lactobacillus helveticus: importance in food and health.

TgaA, a VirB1-like component belonging to a putative type IV secretion system of Bifidobacterium bifidum MIMBb75.

Bifidobacterium bifidum MIMBb75 is a human intestinal isolate demonstrated to be interactive with the host and efficacious as a probiotic. However, the molecular biology of this microorganism is yet largely unknown. For this reason, we undertook whole-genome sequencing of B. bifidum MIMBb75 to identify potential genetic factors that would explain the metabolic and probiotic attributes of this bacterium. Comparative genomic analysis revealed a 45-kb chromosomal region that comprises 19 putative genes coding for a potential type IV secretion system (T4SS). Thus, we undertook the initial characterization of this genetic region by studying the putative virB1-like gene, named tgaA. Gene tgaA encodes a peptidoglycan lytic enzyme containing two active domains: lytic murein transglycosylase (LT, cd00254.3) and cysteine- and histidine-dependent amidohydrolase/peptidase (CHAP, pfam05257.4). By means of several in vitro assays, we experimentally confirmed that protein TgaA, consistent with its computationally assigned role, has peptidoglycan lytic activity, which is principally associated to the LT domain. Furthermore, immunofluorescence and immunogold labeling showed that the protein TgaA is abundantly expressed on the cell surface of B. bifidum MIMBb75. According to the literature, the T4SSs, which have not been characterized before in bifidobacteria, can have important implications for bacterial cell-to-cell communication as well as cross talk with host cells, justifying the interest for further studies aimed at the investigation of this genetic region.

A metallo-β-lactamase is responsible for the degradation of ceftiofur by the bovine intestinal bacterium Bacillus cereus P41

Ceftiofur is a highly effective veterinary cephalosporin, yet it is rapidly degraded by bacteria in the gut. The goal of this work was to directly determine the mechanism of ceftiofur degradation by the bovine intestinal isolate Bacillus cereus P41. B. cereus P41 was isolated from the feces of a cow that had not been treated with cephalosporins, and was found to rapidly degrade ceftiofur in culture. Analysis of spent culture media by HPLC/UV and HPLC/MS revealed one major metabolite of ceftiofur, with a negative ion m/z of 127. Comparison of ceftiofur, ceftriaxone, and cefpodoxime degradation suggested that the major stable ceftiofur metabolite was the thiofuroic acid group eliminated from the C-3 position of the drug after hydrolysis by β-lactamase. Genomic DNA from B. cereus P41 was cloned into Escherichia coli, and the transformants were screened for growth in the presence of ceftiofur. DNA sequencing of the plasmid pHSG299-BC-3 insert revealed the presence of a gene encoding a metallo-β-lactamase. Incubation of ceftiofur with either the E. coli transformant or a commercial B. cereus metallo-β-lactamase showed degradation of the drug and formation of the same major metabolite produced by B. cereus P41. These data demonstrate that a metallo-β-lactamase plays a major role in the degradation of ceftiofur by the bovine intestinal bacterium B. cereus P41.