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Identification of Gram-positive bacteria using BC-GP assay

ABSTRACT: Signal intensities of BC-GP array for 105 speimens The Verigene Gram-Positive Blood Culture (BC-GP) nucleic acid assay is an automated microarray-based test, which can detect 12 Gram-positive bacterial genes and 3 resistance determinants using blood culture broths. We investigated signal intensities of microarray spots, and reclassified undetermined results where the automated system failed and various errors were called in blood culture specimens and spiked samples.

ORGANISM(S): Staphylococcus Streptococcus pneumoniae Staphylococcus hominis Listeria Micrococcus Streptococcus constellatus Streptococcus parasanguinis Streptococcus salivarius Streptococcus gordonii Corynebacterium striatum Streptococcus Streptococcus agalactiae Leuconostoc lactis Streptococcus mitis Staphylococcus epidermidis Streptococcus anginosus group Staphylococcus haemolyticus Streptococcus australis Streptococcus anginosus Staphylococcus capitis Streptococcus gallolyticus Micrococcus luteus Staphylococcus saprophyticus Staphylococcus simulans Corynebacterium afermentans Enterococcus gallinarum Streptococcus thermophilus Escherichia coli Staphylococcus caprae Klebsiella pneumoniae Streptococcus dysgalactiae Enterococcus faecalis Staphylococcus lugdunensis Bacillus licheniformis Listeria monocytogenes Streptococcus pyogenes Staphylococcus aureus Enterococcus faecium

PROVIDER: GSE57245 | GEO | 2014/05/30

SECONDARY ACCESSION(S): PRJNA246048

REPOSITORIES: GEO

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Project description:Objective: It is unclear whether the host response of gram-positive sepsis differs from gram-negative sepsis at a transcriptome level. Using microarray technology, we compared the gene-expression profiles of gram-positive sepsis and gram-negative sepsis in critically ill patients. Design: A prospective cross-sectional study. Setting: A 20-bed general intensive care unit of a tertiary referral hospital. Patients: Seventy-two patients admitted to the intensive care unit. Interventions: Intravenous blood was collected for leukocyte separation and RNA extraction. Microarray experiements were then performed examing the expression level of 19,232 genes in each sample. Measurements and Main Results: There was no difference in the expression profile between gram-positive and gram-negative sepsis. The finding remained unchanged even when genes with lower expression level were included or after statistical stringency was lowered. There were, however, ninety-four genes differentially expressed between sepsis and control patients. These genes included those involved in immune regulation, inflammation and mitochondrial function. Hierarchical cluster analysis confirmed that the difference in gene expression profile existed between sepsis and control patients, but not between gram-positive and gram-negative patients. Conclusion: Gram-positive and gram-negative sepsis share a common host response at a transcriptome level. These findings support the hypothesis that the septic response is non-specific and is designed to provide a more general response that can be elicited by a wide range of different micro-organisms. The study included seventy-two critically ill patients admitted to the intensive care unit (ICU) of Nepean Hospital, Sydney, Australia. Of these, fifty-five patients were diagnosed to have sepsis, as confirmed by microbiological culture. The remaining seventeen patients did not have sepsis and were therefore used as controls. The study was approved by the hospital ethics committee and informed consent was obtained from all patients or their relatives. Patient Samples. Whole blood was taken from each patient on admission to ICU. Neutrophils were separated from whole blood using density-gradient separation with Ficoll-PaqueP P(Amersham). Subsequent neutrophil RNA extraction was performed using guanidinium thiocyanate (Ambion). Microarray Experiment. The neutrophil RNA was converted to cDNA, fluorescently labeled and hybridized to its complimentary sequences on the microarray (Invitrogen). The fluorescent signals on each micrroarray were captured using the GenePix 4000B laser scanner (Axon Instruments). Expression level of each gene was represented by the intensity of its fluorescent signal. Data Extraction. All signal intensity values were processed using background-subtraction method. Prior to analysis, all values were log-transformed and normalized by fitting a print-tip group Lowess curve. Normalization minimizes bias due to dye chemistry, signal intensity or location of a gene on the array. It ensures the detection of genes that are truly differentially expressed, instead of those caused by experimental artifacts or variation in the hybridization process. After normalization, genes that had more than 50% of data missing were removed. We then selected genes that had at least 80% of the data showing two-fold changes from the geneâs median values. After filtering, 1617 genes were available for further analysis.

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Identification of Gram-positive bacteria using BC-GP assay

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