Project description:Microorganisms form biofilms containing differentiated cell populations. To determine factors driving differentiation, we study protein distributions in bacterial biofilms using shotgun proteomics. Notably, zinc- and manganese-depleted portions of the biofilm repress the production of anti-staphylococcal molecules. Exposure to calprotectin (a host protein known to sequester metal ions at infectious foci) recapitulates responses occurring within metal-deplete portions of the biofilm and promotes interaction between P. aeruginosa and Staphylococcus aureus. Consistent with these results, the presence of calprotectin promotes co-colonization of the murine lung, and polymicrobial communities are found to co-exist in calprotectin-enriched airspaces of a cystic fibrosis lung explant. These findings, which demonstrate that metal fluctuations are a driving force of microbial community structure, have clinical implications because of the frequent occurrence of P. aeruginosa and S. aureus co-infections.
Project description:Pseudomonas aeruginosa and Staphylococcus aureus are often co-isolated in persistent infections. The goal of this study was to determine how secreted products from S. aureus affect gene expression in P. aeruginosa. Therefore, media control or S. aureus supernatant was added to P. aeruginosa cultures at 25% total volume and gene expression was measured at 20 min, 1 h, and 2 h using RNA-seq. Overall, after addition of S. aureus supernatant, there was an upregulation in genes involved in metal deprivation and intermediate metabolite uptake.
Project description:Pseudomonas aeruginosa and Staphylococcus aureus are often co-isolated in persistent infections. The goal of this study was to determine how secreted products that were identified in S. aureus supernatant affect gene expression in P. aeruginosa. Therefore, media control, the indicated products in media, or S. aureus supernatant was added to P. aeruginosa cultures at 25% total volume and gene expression was measured at 20 min and 2 h using RNA-seq. The individual products induced distinct pathways in P. aeruginosa. The products in combination recapitulated much of the differential gene expression seen in P. aeruginosa in response to S. aureus supernatant.
Project description:Staphylococcus aureus and Pseudomonas aeruginosa are bacterial pathogens that have been shown to co-exist in biofilms related to numerous infections. Although the interaction between these two species is competitive, both partially benefit from the coexistence. In this study, we exhaustively characterized the interaction between Staphylococcus aureus and Pseudomonas aeruginosa by utilizing a proteomics approach, individually targeting the surface-associated proteins (surfaceome), and proteins secreted or otherwise liberated to the extracellular space (exoproteome). To that end, the conditions to co-culture S. aureus and P. aeruginosa in vitro were optimized and a high-resolution proteomics approach was applied to compare surface-associated and extracellular protein profiles between mono- and co-cultured biofilms.
Project description:Bronchial epithelial cells represent the first line of defense against invading airborne pathogens. They are important contributors to innate mucosal immunity and provide a variety of anti-microbial effectors. To investigate the role of epithelial cells upon infection of airway pathogens, we stimulated BEAS-2B cells for 4 h with UV-inactivated bronchial pathogens including Staphylococcus aureus, Pseudomonas aeruginosa and Respiratory Syncitial Virus (RSV) that among other receptors can strongly activate TLR2, TLR4 and TLR3, respectively. Experiment Overall Design: All conditions were done in triplicates except for Staphylococcus aureus, were two replicates were done. As a control, unstimulated BEAS-2B were used. Altogether 11 arrays were hybridized.
Project description:Pseudomonas aeruginosa and Staphylococcus aureus are often co-isolated in persistent infections. The goal of this study was to determine how secreted products from S. aureus affect gene expression in surface-associated P. aerguinosa undergoing emergent motility. Therefore, media salts control or S. aureus supernatant was added to agar plates at 25% total volume. P. aeruginosa was inoculated on the agar and gene expression was measured from the leading edge after 17 h incubation using RNA-seq. P. aeruginosa moving on the agar containing S. aureus supernatant had an upregulation in genes involved in intermediate metabolite uptake and a downregulation in heme biosynthesis, response to heat, Type III Secretion System, and aerobic respiration pathways.
Project description:Purpose: To uncover immune mediated genes and pathways by Pseudomonas aeruginosa infection that are modulated by the homeodomain PITX1/UNC-30, which plays a vital role in the GABAergic signaling in C. elegans Methods: RNA was extracted from synchronized Pseudomonas aeruginosa infected L4 stage unc-30(ok613) and WT using Qiagen extraction kits and following standard methods. The animals were grown on OP50 at 20 C and infected at 25 C. Results: RNA seq analyses shows enriched and signficant Pseudomonas aeruginosa mediated upregulated immune, neuropeptide and metabolism genes and pathways that are dependent of GABAergic signaling Conclusions: Our study uncovered GABAgergic signaling to be modulator of the innate immunity in C elegans during Pseudomonas aeruginosa infection
Project description:Staphylococcus aureus USA300 and Pseudomonas aeruginosa PAO1 were cultured in microaerobiosis (sealed bottles using a 1:4 medium-to-flask volume ratio without agitation) in Triptic Soy Broth (TSB; Oxoid) supplemented with 0.5% KNO3. S. aureus and P. aeruginosa monocultures were inoculated at a DO600 of 0.05 and incubated for 2h at 37°C. After that, cultures were split in two and furtherly incubated for 2h at 37°C (control) or 39°C (heat shock). A similar protocol was employed for co-culture experiments, where S. aureus and P. aeruginosa were co-inoculated each at a DO600 of 0.05 and followed the same protocol.
Project description:The goal of this study was to determine the impact of metal deprivation (such as the metal deprivation induced by calprotectin treatment) on the physiology of Pseudomonas aeruginosa under multiple growth conditions. The RNA-seq analysis was designed to reveal the impact of calprotectin treatment on P. aeruginosa physiology during planktonic growth.
Project description:Staphylococcus aureus and Pseudomonas aeruginosa are the most prevalent pathogens that colonize structurally abnormal airways such as those in Cystic Fibrosis (CF) and other chronic obstructive lung diseases. Although these bacteria seem to succeed one another, CF patients acquire coinciding P. aeruginosa and S. aureus pulmonary infections, being co-infection usually associated with decreased lung function and increased frequency of pulmonary exacerbations. In addition, P. aeruginosa and S. aureus pathogens adopt a biofilm mode of growth, which contributes to high tolerance to antibiotic treatment and the recalcitrant nature of these chronic coinfections, leading to significant patient morbidity and mortality. Interactions between P. aeruginosa and S. aureus have been widely studied and it is commonly admitted that P. aeruginosa outcompetes S. aureus, perhaps outcompeting S. aureus for limited nutrients or producing anti-staphylococcal compounds, having S. aureus a minimal contribution to the overall course of the infection. However, the molecular mechanisms behind these interactions are largely unknown. Herein, we decided to characterize the full transcriptome of these dual-species biofilms, to unveil important molecular interactions that can occur between these two bacterial species that are relevant for the pathogenesis of the entire consortia. Our data provide novel insights into the role of interspecies interactions in the pathogenesis of P. aeruginosa and S. aureus co-infections and will contribute to future studies by the research community.