title: Truncated human cytidylate-phosphate-deoxyguanylate-binding protein for improved nucleic acid amplification technique-based detection of bacterial species in human samples.
abstract: A trunk of human cytidylate-phosphate-deoxyguanylate-binding protein/CXXC finger protein 1 (CFP1), immobilized onto an aminohexyl-Sepharose column, can be used as a preanalytical tool for the selective enrichment of bacterial DNA from mixed solutions with high amounts of human background DNA for nucleic acid amplification technique-based detection of pathogens. The transcriptional activator protein exhibits a high affinity for nonmethylated CpG dinucleotide motifs, which are differentially distributed in prokaryotic and higher eukaryotic genomes. The feasibility of the affinity chromatography (AC) step was tested with DNA from severely septic patients. AC using 16S rRNA gene primers substantially increased PCR sensitivity. Approximately 90% of eukaryotic DNA was removed, which significantly increased the signal-to-noise ratio. Threshold cycle values revealed that sensitivity was elevated at least 10-fold. The change in the ratio of bacterial DNA to human DNA increased from 26% to 74% the likelihood of culture-independent PCR-based identification of bacterial presence. Compared to the results seen with blood culture (which is the clinical gold standard for systemic infections, exhibiting 28% positives), the combination of AC and PCR achieves a significant increase in sensitivity and contributes to shortening the time to results for the initiation of guided antibiotic therapy.
authors.affiliation: Institute of Medical Microbiology, Friedrich-Schiller-University, Jena, Germany.
authors.forename: Svea
authors.initials: S
authors.lastname: Sachse
authors.forename: Eberhard
authors.initials: E
authors.lastname: Straube
authors.forename: Marc
authors.initials: M
authors.lastname: Lehmann
authors.forename: Michael
authors.initials: M
authors.lastname: Bauer
authors.forename: Stefan
authors.initials: S
authors.lastname: Russwurm
authors.forename: Karl-Hermann
authors.initials: KH
authors.lastname: Schmidt
mesh_terms: D001419:Bacteria / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D001424:Bacterial Infections / Q000175:diagnosis*; D001431:Bacteriological Techniques / Q000379:methods*; D001769:Blood / Q000382:microbiology; D002846:Chromatography, Affinity / Q000379:methods*; D004269:DNA, Bacterial / Q000302:isolation & purification; D004268:DNA-Binding Proteins; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012680:Sensitivity and Specificity; D015534:Trans-Activators
chemical_list: C409062:CXXC1 protein, human; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D015534:Trans-Activators