title: Application of qualitative and quantitative real-time PCR, direct sequencing, and terminal restriction fragment length polymorphism analysis for detection and identification of polymicrobial 16S rRNA genes in ascites.
abstract: Qualitative and quantitative 16S rRNA gene-based real-time PCR and direct sequencing were applied for rapid detection and identification of bacterial DNA (bactDNA) in 356 ascites samples. bactDNA was detected in 35% of samples, with a mean of 3.24 log copies ml(-1). Direct sequencing of PCR products revealed 62% mixed chromatograms predominantly belonging to Gram-positive bacteria. Terminal restriction fragment length polymorphism (T-RFLP) results of a sample subset confirmed sequence data showing polymicrobial DNA contents in 67% of bactDNA-positive ascites samples.
authors.affiliation: University Hospital Leipzig, Division of Gastroenterology and Hepatology, Leipzig, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Krohn
authors.forename: Stephan
authors.initials: S
authors.lastname: Böhm
authors.forename: Cornelius
authors.initials: C
authors.lastname: Engelmann
authors.forename: Jan
authors.initials: J
authors.lastname: Hartmann
authors.forename: Annika
authors.initials: A
authors.lastname: Brodzinski
authors.forename: Antonis
authors.initials: A
authors.lastname: Chatzinotas
authors.forename: Katharina
authors.initials: K
authors.lastname: Zeller
authors.forename: Delia
authors.initials: D
authors.lastname: Prywerek
authors.forename: Ingo
authors.initials: I
authors.lastname: Fetzer
authors.forename: Thomas
authors.initials: T
authors.lastname: Berg
mesh_terms: D001201:Ascites / Q000175:diagnosis* / Q000382:microbiology*; D004269:DNA, Bacterial / Q000235:genetics; D004275:DNA, Ribosomal / Q000235:genetics; D020459:Genes, rRNA / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods; D012150:Polymorphism, Restriction Fragment Length / Q000235:genetics*; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S