Project description:Bile acids are not only crucial for the uptake of lipids, but also have widespread systematic ef-fects and shape the gut-microbiome composition. Bile acids can directly shape the gut-microbiome and can be modified by bacteria such as Eggerthella lenta which in turn plays a crucial role in host metabolism and immune response. We cultivated eight strains that represent a simplified human intestinal microbiome and inves-tigated the molecular response to bile acids, co-culturing with Eggerthella lenta and the combina-tion. We observed growth inhibition of particularly gram-positive strains during bile acid stress, which could be alleviated through co-culturing with Eggerthella lenta. The inhibition of growth was related to a decrease in membrane integrity and genotoxic effects of bile acids, which we investigated using zeta potential measurements in combination with proteomic and metabolomic analyses. Co-culturing with Eggerthella lenta alleviated stress through formation of oxidized and epimer-ized bile acids and the molecular response to co-culturing was seen to be strain specific. We also note that we could detect the recently described Microbial Bile Salt Conjugates in our cultures. This study highlights the significance of a potent bile acid modifier and how in-depth molecular analyses are required to decipher cross-communication between gut and host.
Project description:Industrial production of penicillins with the filamentous fungus Penicillium chrysogenum is based on an unprecedented effort in microbial strain improvement. Sequencing of the 32.19 Mb genome of P. chrysogenum Wisconsin54-1255 revealed many genes responsible for key steps in penicillin production. DNA microarrays were used to compare the transcriptomes of the sequenced strain and a penicillinG high-producing strain, grown in the presence and absence of the side-chain precursor phenylacetic acid. Transcription of genes involved in biosynthesis of valine, cysteine and alpha-aminoadipic acid, the amino-acid precursors for penicillin biosynthesis, as well as genes encoding microbody proteins, increased in the high-producing strain. Many key (intra)cellular transport processes involving penicillins and intermediates remain to be characterized at the molecular level. Genes predicted to encode transporters were strongly overrepresented among the genes transcriptionally upregulated under conditions that stimulate penicillinG production, illustrating potential for future genomics-driven functional analysis. Keywords: genetic modification
Project description:Listeria monocytogenes is a gram-positive, food-borne pathogen responsible for invasive infections with high overall mortality. Early host defenses encountered by L. monocytogenes following ingestion include low pH of the stomach and bile present in the small intestinal lumen. We hypothesized that “epidemic” strains are better able to withstand exposure to low pH and bile encountered in the gastrointestinal tract as compared to most “environmental” strains. Furthermore, we hypothesized that epidemic and environmental strains would have distinct transcriptional programs upon exposure to these conditions. Our treatments included 1 hr exposure to acid (pH 5.5 and 3.5) and bile (0.3%) stress. Strains were pre-exposed to pH 5.5 (1 hr) before being treated with pH 3.5. We used a collection of 12 previously characterized epidemic and environmental strains and each strainXtreatment combination included 3 biological replicates for each microarray experiment. All microarray experiments were two color competitive hybridizations that paired experimental conditions with the same strain at neutral pH for acid stress and pH 5.5 for bile stress. Transcriptomes of environmental strains exposed to acid and bile stress showed remarkably greater number of genes with differences of ≥2-fold expression levels as compared to epidemic strains (5 and 7, respectively). Environmental strains were characterized by up-regulation of several stress related genes and down-regulation of several cell envelope biosynthesis and virulence related genes, suggesting that complex regulatory networks orchestrate the cellular changes in the environmental strains to overcome stressful environments. The transcriptome of epidemic strains, in contrast, showed muted responses to these stress conditions implying their pre-adaptability to acid and bile stress encountered during natural infection that may enable epidemic strains to survive and become “primed” for subsequent colonization and infection in the lower gastrointestinal tract. Keywords: stress response, comparative transcriptomics, acid-adaptation, differential virulence, acid-stress response, bile-stress response
Project description:Little is known about the bacteria that reside in human gallbladder and the mechanisms that allow them to survive within this harsh environmental niche. Furthermore, certain bacterial species are considered to exhibit antagonistic activities whilst others may form mutualistic interactions through, for example, cross-feeding. We isolated two new strains from healthy human bile samples, one belonging to Ruminococcus gauvreauii, of Lachnospiraceae family, and other constituting a new specie in Ruminococcaceae family, named Ruminocoides biliarensis. The two strains differed markedly in their carbohydrate metabolism as R. gauvreauii mainly metabolised sugar alcohols, including inositol, to form acetate as unique fermentation product, and Rc. biliarensis mainly metabolised resistant starches to mainly form formate and acetate as fermentation end products. Both strains exhibited resistance to different bile salts, and the ability to sporulate. Amino acid and vitamin biosynthesis profiles also markedly differed between the two bile isolates. Finally,RNAseq was used to analyse the co-cultures of both isolates, to analyze the activities involved in the possible cross-feeding relationship.
Project description:Lactobacillus salivarius is a member of the indigenous microbiota of the human gastrointestinal tract (GIT). Tolerance to bile stress is crucial for intestinal lactobacilli to survive in the GIT and to exert their beneficial actions. In this work, the Next-Generation Sequencing platform Illumina HiSeq 2000 was used to investigate the global response to bile in L. salivarius Ren, a potential probiotic strain isolated from a healthy centenarian. In the presence of 0.75 g liter-1 oxgall, the transcription of nearly 200 genes was detected to be associated with bile stress, including genes involved in carbohydrate and amino acid metabolism, cell envelope and fatty acid biogenesis, transcription and translation. This study improves our understanding on bile stress response in L. salivarius Ren.