Phage response to CRISPR-encoded resistance in Streptococcus thermophilus.
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Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated genes are linked to a mechanism of acquired resistance against bacteriophages. Bacteria can integrate short stretches of phage-derived sequences (spacers) within CRISPR loci to become phage resistant. In this study, we further characterized the efficiency of CRISPR1 as a phage resistance mechanism in Streptococcus thermophilus. First, we show that CRISPR1 is distinct from previously known phage defense systems and is effective against the two main groups of S. thermophilus phages. Analyses of 30 bacteriophage-insensitive mutants of S. thermophilus indicate that the addition of one new spacer in CRISPR1 is the most frequent outcome of a phage challenge and that the iterative addition of spacers increases the overall phage resistance of the host. The added new spacers have a size of between 29 to 31 nucleotides, with 30 being by far the most frequent. Comparative analysis of 39 newly acquired spacers with the complete genomic sequences of the wild-type phages 2972, 858, and DT1 demonstrated that the newly added spacer must be identical to a region (named proto-spacer) in the phage genome to confer a phage resistance phenotype. Moreover, we found a CRISPR1-specific sequence (NNAGAAW) located downstream of the proto-spacer region that is important for the phage resistance phenotype. Finally, we show through the analyses of 20 mutant phages that virulent phages are rapidly evolving through single nucleotide mutations as well as deletions, in response to CRISPR1. (+info)
Role of galK and galM in galactose metabolism by Streptococcus thermophilus.
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Streptococcus thermophilus is unable to metabolize the galactose moiety of lactose. In this paper, we show that a transformant of S. thermophilus SMQ-301 expressing Streptococcus salivarius galK and galM was able to grow on galactose and expelled at least twofold less galactose into the medium during growth on lactose. (+info)
The inhibitory spectrum of thermophilin 9 from Streptococcus thermophilus LMD-9 depends on the production of multiple peptides and the activity of BlpG(St), a thiol-disulfide oxidase.
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The blp(St) cluster of Streptococcus thermophilus LMD-9 was recently shown to contain all the genetic information required for the production of bacteriocins active against other S. thermophilus strains. In this study, we further investigated the antimicrobial activity of S. thermophilus LMD-9 by testing the susceptibility of 31 bacterial species (87 strains). We showed that LMD-9 displays an inhibitory spectrum targeted toward related gram-positive bacteria, including pathogens such as Listeria monocytogenes. Using deletion mutants, we investigated the contribution of the three putative bacteriocin-encoding operons blpD(St)-orf2, blpU(St)-orf3, and blpE(St)-blpF(St) (bac(St) operons) and of the blpG(St) gene, which encodes a putative modification protein, to the inhibitory spectrum and immunity of strain LMD-9. Our results present evidence that the blp(St) locus encodes a multipeptide bacteriocin system called thermophilin 9. Among the four class II bacteriocin-like peptides encoded within the bac(St) operons, BlpD(St) alone was sufficient to inhibit the growth of most thermophilin 9-sensitive species. The blpD(St) gene forms an operon with its associated immunity gene(s), and this functional bacteriocin/immunity module could easily be transferred to Lactococcus lactis. The remaining three Bac(St) peptides, BlpU(St), BlpE(St), and BlpF(St), confer poor antimicrobial activity but act as enhancers of the antagonistic activity of thermophilin 9 by an unknown mechanism. The blpG(St) gene was also shown to be specifically required for the antilisteria activity of thermophilin 9, since its deletion abolished the sensitivities of most Listeria species. By complementation of the motility deficiency of Escherichia coli dsbA, we showed that blpG(St) encodes a functional thiol-disulfide oxidase, suggesting an important role for disulfide bridges within thermophilin 9. (+info)
Molecular analysis of yogurt containing Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus in human intestinal microbiota.
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BACKGROUND: Yogurt has traditionally been considered a probiotic-carrier food with health-promoting effects. Despite the universal assumption of this assertion, several researchers have evaluated the real capability of the yogurt bacteria Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus to survive and proliferate in the human intestine and have found contradictory results. OBJECTIVE: This double-blind crossover study assessed the qualitative and quantitative effects of fresh and heat-treated yogurt on bacterial intestinal microbiota from healthy subjects. DESIGN: The subjects were divided into experimental (n=63) and control (n=16) groups. The experimental group consumed fresh and heat-treated yogurt for 15 d according to a crossover design with a washout period of 2 wk. Three different fecal samples per individual were recovered: at baseline, after fresh yogurt intake, and after heat-treated yogurt intake. Qualitative changes in microbiota were studied by denaturing gel gradient electrophoresis (DGGE) with universal and lactic acid bacteria (LAB) 16S-rRNA primers. Quantitative changes in LAB, Clostridium coccoides, Clostridium perfringens, and Bacteroides groups were analyzed by real-time polymerase chain reaction. RESULTS: A particular DGGE stable band pattern was observed in each sample. No significant qualitative differences were detected in any fecal sample. However, a significantly higher density of LAB and C. perfringens and a significant decrease in the density of Bacteroides was observed after consumption of both types of yogurt. Microbiota density was not significantly different between the fresh and heat-treated yogurt groups, except for LAB, which was significantly greater in the fresh yogurt group. CONCLUSION: The main change in human microbiota observed after yogurt consumption was an increase in the density of LAB and C. perfringens to the detriment of Bacteroides. Bacterial changes were not different after the consumption of fresh and heat-treated yogurt. (+info)
Plasmid transfer via transduction from Streptococcus thermophilus to Lactococcus lactis.
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Probiotic Leuconostoc mesenteroides ssp. cremoris and Streptococcus thermophilus induce IL-12 and IFN-gamma production.
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AIM: To investigate the capacity of potentially probiotic strains from six bacterial genera to induce cytokine production alone or in combinations in order to identify potential enhancing or synergistic effects in order to select probiotic bacteria for in vivo purposes. METHODS: Cytokine production in human peripheral blood mononuclear cells (PBMC) in response to stimulation with eleven different potentially probiotic bacterial strains from Streptococcus, Lactobacillus, Bifidobacterium, Lactococcus, Leuconostoc and Propionibacterium genera was analysed. Production and mRNA expression of TNF-alpha, IL-12, IFN-gamma and IL-10 were determined by ELISA and Northern blotting, respectively. RESULTS: All tested bacteria induced TNF-alpha production. The best inducers of Th1 type cytokines IL-12 and IFN-gamma were Streptococcus and Leuconostoc strains. All Bifidobacterium and Propionibacterium strains induced higher IL-10 production than other studied bacteria. Stimulation of PBMC with any bacterial combinations did not result in enhanced cytokine production suggesting that different bacteria whether gram-positive or gram-negative compete with each other during host cell interactions. CONCLUSION: The probiotic S. thermophilus and Leuconostoc strains are more potent inducers of Th1 type cytokines IL-12 and IFN-gamma than the probiotic Lactobacillus strains. Bacterial combinations did not result in enhanced cytokine production. (+info)
A double blind randomized controlled trial of a probiotic combination in 100 patients with irritable bowel syndrome.
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