Project description:Commercial probiotic bacteria must be tested for acquired antibiotic resistance elements to avoid potential transfer to pathogens. The European Food Safety Authority recommends testing resistance using microdilution culture techniques previously used to establish inhibitory thresholds for the Bifidobacterium genus. Many Bifidobacterium animalis subsp. lactis strains exhibit increased resistance to tetracycline, historically attributed to the ribosomal protection gene tet(W). However, some strains that harbor genetically identical tet(W) genes show various inhibition levels suggesting that other genetic elements also contribute to observed differences. Here, we adapted several molecular assays to confirm the inhibition of B. animalis subsp. lactis strains Bl-04 and HN019, and employed RNA-seq to assess the transcriptional differences related to genomic polymorphisms. We detected specific stress responses to the antibiotic by correlating ATP concentration to viable genome copies from droplet digital PCR, and found that the bacteria were still metabolically active in high drug concentrations. Transcriptional analyses revealed that several polymorphic regions, particularly a novel multi-drug efflux transporter, were differentially expressed between the strains in each experimental condition, likely having phenotypic effects. We also found that the tet(W) gene was up-regulated only during sub-inhibitory tetracycline concentrations, while two novel tetracycline resistance genes were up-regulated at high concentrations. Furthermore, many genes involved in amino acid metabolism and transporter function were up-regulated while genes for complex carbohydrate utilization, protein metabolism, and CRISPR-Cas systems were down-regulated. These results provide high-throughput means for assessing antibiotic resistance and determine the genetic network that contributes to the global tetracycline response between two highly related probiotic strains.
Project description:Modulation of gut microbiota through probiotic supplementation is an interesting strategy to prevent obesity We use microarrays to study the global genome expression of C. elegans fed with the probiotic strain Bifidobacterium animalis sbsp. lactis CECT 8145
Project description:Bifidobacterium animalis subsp. lactis BB12 is a widely used probiotic bacterium. However the molecular mechanism by which this bacterium confers positive health effects to the host is largely unknown. The importance of lipoteichoic acids (LTA) in the cross-talk between the bacteria and human enterocyte-like Caco-2 cells was investigated in the present study. A microarray-based analysis of gene expression revealed induction of genes encoding enzymes involved in the synthesis of lipoteichoic acids of BB12. This observation was confirmed in phenotypic experiments, where a specific galactofuranosyl carbohydrate epitope present in the structure of lipoteichoic acids of BB12 was used as a detection marker. An exopolysaccharide EPS/LTA extract isolated from BB12 and commercially available pure LTAs from the human pathogen S. pyogenes were tested in competitive adhesion experiments. The adhesion of B. animalis subsp. lactis BB12 to Caco-2 cells was inhibited in presence of LTA. The strongest inhibition was observed in the concentration range 0.5-10 ug/ml for LTAs from S. pyogenes and 5-10 ug/ml for EPS/LTA extract from BB12. The inhibitory concentrations in both cases likely reflect the physico-chemical properties of LTAs with the highest efficacy below the critical micelle concentration, and therefore we ascribe the main inhibitory effect to LTAs for the BB12 isolated EPS/LTA extract as well. In conclusion we present increased biosynthesis of lipoteichoic acids of BB12 co-incubated with human Caco-2 cells and evidence that LTA serve as important cell wall factors involved in adhesion to these human enterocyte-like cells.