Project description:The opportunistic pathogen, Staphylococcus aureus, encounters a wide variety of fluid shear levels within the human host, which may play a key role in dictating whether this organism adopts a commensal interaction with the host or transitions to cause disease. Using rotating-wall vessel bioreactors to create a physiologically-relevant, low fluid shear environment, S. aureus was evaluated for cellular responses that could impact its colonization and virulence. S. aureus cells grown in a low fluid shear environment initiated a novel attachment-independent biofilm phenotype and were completely encased in extracellular polymeric substances. Compared to controls, low-shear cultured cells displayed slower growth and repressed virulence characteristics, including decreased carotenoid production, increased susceptibility to oxidative stress, and reduced survival in whole blood. Transcriptional whole genome microarray profiling suggested alterations in metabolic pathways. Further genetic expression analysis revealed the down-regulation of the RNA chaperone Hfq, which parallels low fluid shear responses of certain Gram negative organisms. This is the first study to report an Hfq association to fluid shear in a Gram positive organism, suggesting an evolutionarily conserved response to fluid shear among structurally diverse prokaryotes. Collectively, our results suggest S. aureus responds to a low fluid shear environment by initiating a biofilm/colonization phenotype with diminished virulence characteristics, which could lead to insight into key factors influencing the divergence between infection and colonization during initial host pathogen interaction. Genetic expression profiles of Staphylococcus aureus cultured under low fluid shear conditions was compared to control cultures of S. aureus which was cultured in identical hardware in an orientation disrupting the low fluid shear effect. Samples from the same date of culture were compared (control 21:low 21 and control 30: low 30). S. aureus was cultured for 20 hours in either the low fluid shear or control orientated rotating wall vessel (RWV) bioreactor at which point the cells were removed and RNA extracted. At 20 hours, both cultures were in the same stage of growth (stationary phase) and at this point phenotypic differences between control and low fluid shear cultures were noted.
Project description:S. aureus has the propensity to survive a range of NaCl challenge conditions We used commercially available Affymetrix S. aureus GeneChips (part number 900514) to compare the gene expression properties of wild type cells during growth at no or high (2M) NaCl. S. aureus strain USA300-lac cells were grown to late exponential phase growth in the absence or presence of 2M NaCl, total bacterial RNA was isolated and subjected to GeneChip hybridization and analysis. We sought to determine the regulatory effects of high NaCl.
Project description:In the present study, we employed Affymetrix Staphylococcus aureus GeneChip arrays to investigate the dynamics of global gene expression profiles during the cellular response of Staphylococcus aureus to peracetic acid, which involved initial growth inhibition and subsequent partial recovery. Keywords: Time course
Project description:Staphylococcus aureus is an important human pathogen that causes life-threatening infections, and is resistant to the majority of our antibiotic arsenal. This resistance is complicated by the observation that most antibacterial agents target actively growing cells, thus, proving ineffective against slow growing populations, such as cells within a biofilm or in stationary phase. Recently, our group generated updated genome annotation files for S. aureus that not only include protein-coding genes but also regulatory and small RNAs. As such, these annotation files were used to perform a transcriptomic analysis in order to understand the metabolic and physiological changes that occur during transition from active growth to stationary phase; with a focus on sRNAs. We observed â¼24% of protein-coding and 34% of sRNA genes displaying changes in expression by â¥3-fold. Collectively, this study adds to our understanding of S. aureus adaptation to nutrient-limiting conditions, and sheds new light onto the contribution of sRNAs to this process. Bacterial cells were grown in TSB medium at 37°C with shaking for 3h (exponential growth phase) or 16h (stationary growth phase).
Project description:Staphylococcus aureus is one of the earliest pathogens that persists the airways of cystic fibrosis (CF) patients and contributes to increased inflammation and decreased lung function. In contrast to other staphylococci, S. aureus possesses two superoxide dismutases (SODs), SodA and SodM, with SodM being unique to S. aureus. Both SODs arm S. aureus for its fight against oxidative stress, a byproduct of inflammatory reactions. Despite complex investigations it is still unclear, if both enzymes are crucial for the special pathogenicity of S. aureus. To investigate the role of both SODs during staphylococcal persistence in CF airways, we analyzed survival and gene expression of S. aureus CF isolates and laboratory strains in different CF-related in vitro and ex vivo settings. Bacteria located in inflammatory and oxidized CF sputum transcribed high levels of sodA and sodM. Especially expression values of sodM were remarkably higher in CF sputum than in bacterial in vitro cultures. Interestingly, also S. aureus located in airway epithelial cells expressed elevated transcript numbers of both SODs, indicating that S. aureus is exposed to oxidative stress at various sites within CF airways. Both enzymes promoted survival of S. aureus during PMN killing and seem to act compensatory, thereby giving evidence that the interwoven interaction of SodA and SodM contributes to S. aureus virulence and facilitates S. aureus persistence within CF airways
Project description:More than 200 direct CodY target genes in Staphylococcus aureus were identified by genome-wide analysis of in vitro DNA binding. This analysis, which was confirmed for some genes by DNase I footprinting assays, revealed that CodY is a direct regulator of numerous transcription units associated with amino acid biosynthesis, transport of macromolecules and virulence. The virulence genes regulated by CodY fell into three groups. One group was dependent on the Agr system for its expression; these genes were indirectly regulated by CodY through its repression of the agr locus. A second group was regulated directly by CodY. The third group, which includes genes for alpha-toxin and capsule synthesis, was regulated by CodY in two ways, i.e., by direct repression and by repression of the agr locus. Since S. aureus CodY was activated in vitro by the branched chain amino acids and GTP, CodY appears to link changes in intracellular metabolite pools with the induction of numerous adaptive responses, including virulence. Affymetrix GeneChips were used to compare the transcript titers of S. aureus strains UAMS-1 (wild type) and corresponding agr- (strain CM18), codY- (strain MS1), and agr- codY- (strain CM19) isogenic mutant strains during exponential and stationary phase growth. At least two biological replicates were assessed for each strain and each growth phase.
Project description:Microarray expression profiling of S. aureus lavaged from murine lungs after residence times between 30 minutes and 6 hours compared to post-exponential phase or early log phase growth in Luria-Betani broth.