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PROVIDER: PRJNA833222 | ENA |
REPOSITORIES: ENA
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Project description:Emerging known and unknown pathogens create profound threats to public health. Platforms for rapid detection and characterization of microbial agents are critically needed to prevent and respond to disease outbreaks. Available detection technologies cannot provide broad functional information about known and novel organisms. As a step toward developing such a system, we have produced and tested a series of high-density functional gene arrays to detect elements of virulence and antibiotic resistance mechanisms. Our first generation array targets genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for gene family detection and discrimination. When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family. In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms. This is the first report of a high density NimbleGen microarray system targeting microbial antibiotic resistance and virulence mechanisms. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples. Keywords: detection, pathogen, virulence mechanism
2008-04-02 | GSE11010 | GEO
Project description:Emerging known and unknown pathogens create profound threats to public health. Platforms for rapid detection and characterization of microbial agents are critically needed to prevent and respond to disease outbreaks. Available detection technologies cannot provide broad functional information about known and novel organisms. As a step toward developing such a system, we have produced and tested a series of high-density functional gene arrays to detect elements of virulence and antibiotic resistance mechanisms. Our first generation array targets genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for gene family detection and discrimination. When tested with organisms at varying phylogenetic distances from the four target strains, the array detected orthologs for the majority of targeted gene families present in bacteria belonging to the same taxonomic family. In combination with whole-genome amplification, the array detects femtogram concentrations of purified DNA, either spiked in to an aerosol sample background, or in combinations from one or more of the four target organisms. This is the first report of a high density NimbleGen microarray system targeting microbial antibiotic resistance and virulence mechanisms. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples. Keywords: detection, pathogen, virulence mechanism In this report, we describe the process used to design our first generation functional array for highly sensitive detection of virulence and antibiotic resistance gene families. We discuss the probe design algorithms, including virulence gene sequence selection, and our protocols for sample preparation, amplification, labeling, hybridization, and data analysis. We present the results from experiments designed to assess whether the array can detect virulence gene orthologs from organisms without perfect match probes on the array, using both targeted mismatch probes and hybridizations to DNA from other organisms. Also, we report the results from limit of detection studies, using known amounts of bacterial DNA spiked into aerosol samples to measure the minimal concentration required for detection of virulence elements against a complex background.
2008-04-02 | E-GEOD-11010 | biostudies-arrayexpress
Project description:Staphylococcus aureus is a Gram-positive human pathogen causing a variety of human diseases in both hospital and community settings. This bacterium is so closely associated with prophages that it is rare to find S. aureus isolates without prophages. Two phages are known to be important for staphylococcal virulence: the beta-hemolysin (hlb) converting phage and the Panton-Valentine Leukocidin (PVL) converting phage. The hlb-converting phage is found in more than 90% of clinical isolates of S. aureus. This phage produces exotoxins and immune modulatory molecules, which inhibit human innate immune responses. The PVL-converting phage produces the two-component exotoxin PVL, which can kill human leucocytes. This phage is wide-spread among community-associated methicillin resistant S. aureus (CA-MRSA). It also shows strong association with soft tissue infections and necrotizing pneumonia. Several lines of evidence suggest that staphylococcal prophages increase bacterial virulence not only by providing virulence factors but also by altering bacterial gene expression: 1) Transposon insertion into prophage regulatory genes, but not into the genes of virulence factors, reduced S. aureus killing of Caenorhabditis elegans.; 2) Although the toxins and immune modulatory molecules encoded by the hlb- converting phages do not function in the murine system, deletion of ϕNM3, the hlb-converting phage in S. aureus Newman, reduced staphylococcal virulence in the murine abscess formation model. 3) In a preliminary microarray experiment, prophages in S. aureus Newman altered the expression of more than 300 genes. In this research proposal, using microarray and high-throughput quantitative RT-PCR (qRT-PCR) technologies, we will identify the effects of the two important staphylococcal phages on the gene expression of S. aureus in both in vitro and in vivo conditions. This project is intended to be completed within one year. All the data – microarray, qRT-PCR and all the primer sequences- will be made available to public 6 month after completion. Data from this project will help us to understand the role of prophages in the S. aureus pathogenesis and can lead to development of a strategy to interfere with the pathogenesis process. Following strains were grown in TSA broth: Staphylococcus aureus USA300 (reference) Staphylococcus aureus USA300 with deletion of ϕSa2usa (Query) Staphylococcus aureus USA300 with deletion of ϕSa3usa (Query) Staphylococcus aureus USA300 Prophage-free mutant (Query) Staphylococcus aureus USA300 Prophage-free mutant lysogenized with ϕSa2mw (Query) Staphylococcus aureus USA300 Prophage-free mutant lysogenized with ϕSa3usa (Query) strain: Staphylococcus aureus USA300 Prophage-free mutant lysogenized with both ϕSa2mw and ϕSa3usa (Query) RNA samples were harvested at early log, midlog and stationary phase.Samples were hybridized on aminosilane coated slides with 70-mer oligos.
2011-02-28 | E-GEOD-26623 | biostudies-arrayexpress
Project description:Staphylococcus aureus (S. aureus) has already to be one of the most commonly identified bacteria that cause food poisoning. S. aureus colonization in humans can cause serious infections, toxinoses and life threatening diseases. The bacteriocin nisin has been extensively used as potential natural preservative in the food industry, but the overall transcriptional response mechanisms of S. aureus to nisin are still poorly understood. To detect the possible molecular mechanism of nisin against S. aureus, Affymetrix GeneChips were used to determine the global comparative transcription of S. aureus cells triggered by treatment with sub-inhibitory concentrations of nisin. Staphylococcus aureus planktonic cells were exposed for 60 minutes to nisin at concentration of 4 M-NM-<g/ml (1/2M-CM-^W MIC). 2 samples including 2 control samples are analyzed.
2015-02-10 | E-GEOD-65750 | biostudies-arrayexpress
Project description:Staphylococcus aureus is a major human pathogen and resistant to numerous clinically used antibiotics. The first antibiotic developed for S. aureus infections was the nonribosomal petide secondary metabolite penicillin. We discovered cryptic nonribosomal peptide secondary metabolites, the aureusimines, made by S. aureus itself that are not antibiotics, but function as small molecule regulators of virulence factor expression. Using established rules and codes for nonribosomal peptide assembly we predicted these nonribosomal peptides, and used these predictions to identify them from S. aureus culture broths. Functional studies using global microarray and mouse bacteremia models established that the aureusimines control virulence factor expression and are necessary for productive infections. This is the first report of the aureusimines and has important implications for the treatment of drug resistant S. aureus. Targeting aureusimine synthesis may provide novel anti-infectives.
2010-07-01 | GSE21373 | GEO
Project description:Staphylococcus aureus (S. aureus) has already to be one of the most commonly identified bacteria that cause food poisoning. S. aureus colonization in humans can cause serious infections, toxinoses and life threatening diseases. The bacteriocin nisin has been extensively used as potential natural preservative in the food industry, but the overall transcriptional response mechanisms of S. aureus to nisin are still poorly understood. To detect the possible molecular mechanism of nisin against S. aureus, Affymetrix GeneChips were used to determine the global comparative transcription of S. aureus cells triggered by treatment with sub-inhibitory concentrations of nisin.
2015-02-10 | GSE65750 | GEO
Project description:Staphylococcus aureus is responsible for a substantial number of invasive infections globally each year. These infections are problematic because they are frequently recalcitrant to antibiotic treatment. Antibiotic tolerance, the ability of bacteria to persist despite normally lethal doses of antibiotics, contributes to antibiotic treatment failure in S. aureus infections. To understand how antibiotic tolerance is induced, S. aureus biofilms exposed to multiple anti-staphylococcal antibiotics were examined using both quantitative proteomics and transposon sequencing. These screens indicated that arginine metabolism is involved in antibiotic tolerance within a biofilm and led to the hypothesis that depletion of arginine within S. aureus communities can induce antibiotic tolerance. Consistent with this hypothesis, inactivation of argH, the final gene in the arginine synthesis pathway, induces antibiotic tolerance. Arginine restriction was found to induce antibiotic tolerance via inhibition of protein synthesis. In a mouse skin infection model, an argH mutant has enhanced ability to survive antibiotic treatment with vancomycin, highlighting the relationship between arginine metabolism and antibiotic tolerance during S. aureus infection. Uncovering this link between arginine metabolism and antibiotic tolerance has the potential to open new therapeutic avenues targeting previously recalcitrant S. aureus infections.
2024-07-10 | GSE267626 | GEO
Project description:Staphylococcus aureus is a major human pathogen and resistant to numerous clinically used antibiotics. The first antibiotic developed for S. aureus infections was the nonribosomal petide secondary metabolite penicillin. We discovered cryptic nonribosomal peptide secondary metabolites, the aureusimines, made by S. aureus itself that are not antibiotics, but function as small molecule regulators of virulence factor expression. Using established rules and codes for nonribosomal peptide assembly we predicted these nonribosomal peptides, and used these predictions to identify them from S. aureus culture broths. Functional studies using global microarray and mouse bacteremia models established that the aureusimines control virulence factor expression and are necessary for productive infections. This is the first report of the aureusimines and has important implications for the treatment of drug resistant S. aureus. Targeting aureusimine synthesis may provide novel anti-infectives. Commerically available S. aureus GeneChips (Affymetrix) were used to compare biological replicates of early and late exponential phase wild type (Newman) and aureusimine defective (ausA) organisms.
2010-07-30 | E-GEOD-21373 | biostudies-arrayexpress
Project description:Staphylococcus aureus can infect a wide range of animals and pose as a serious threat to public health by transferring via animals or animal-derived food stuff. Even more importantly, multiple drug resistance development in the bacteria has resulted in therapeutic failure of a number of antibiotics. Therefore by realizing the need of time, this study was designed to investigate the underlying mechanisms of virulence and resistance in S. aureus. After screening through in vivo and in vitro virulence assays and susceptibility test, a highly virulent and multidrug resistant MRSA strain was selected for differential analysis by RNA-seq technology and gene expression results were verified by RT-qPCR. Up-regulation of crucial regulators like sarA and KdpDE seemed to play role in decreased expression of many exotoxin genes while enhanced the adhesion and cell wall protein expression, leading to strong biofilm production in the presence of inactivated agr system. In addition to resistance genes like blaZ, ermC and femA, up-regulation of vraS and multidrug ABC transporter genes contributed to the multidrug resistance in MRSA. Fluoroquinolone resistance was attributed to mutational changes in gyrA and parC genes. Our findings suggested that many virulence and resistance determinants in S. aureus are controlled by complex network of various regulators, and sarA is the most important of those as it adds to pathogenicity of the bacteria and ensures its survival in diverse environment. Further investigations are required to unveil these mechanisms in S. aureus. Four samples were analysed including 2 MRSA1679a test strain and 2 reference strain ATCC1 samples with two replicates of each.
2016-03-02 | E-GEOD-78764 | biostudies-arrayexpress