Comparison of the sensitivity of culture, PCR and quantitative real-time PCR for the detection of Pseudomonas aeruginosa in sputum of cystic fibrosis patients.

BMC Microbiology
Pieter DeschaghtMario Vaneechoutte

Abstract

Pseudomonas aeruginosa is the major pathogen involved in the decline of lung function in cystic fibrosis (CF) patients. Early aggressive antibiotic therapy has been shown to be effective in preventing chronic colonization. Therefore, early detection is important and sensitive detection methods are warranted. In this study, we used a dilution series of P. aeruginosa positive sputa, diluted in a pool of P. aeruginosa negative sputa, all from CF patients--to mimick as closely as possible the sputa sent to routine laboratories--to compare the sensitivity of three culture techniques versus that of two conventional PCR formats and four real-time PCR formats, each targeting the P. aeruginosa oprL gene. In addition, we compared five DNA-extraction protocols. In our hands, all three culture methods and the bioMérieux easyMAG Nuclisens protocol Generic 2.0.1, preceded by proteinase K pretreatment and followed by any of the 3 real-time PCR formats with probes were most sensitive and able to detect P. aeruginosa up to 50 cfu/ml, i.e. the theoretical minimum of one cell per PCR mixture, when taking into account the volumes used in this study of sample for DNA-extraction, of DNA-elution and of DNA-extract in the PCR mixture. In this study, n...Continue Reading

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Citations

May 18, 2012·American Journal of Respiratory and Critical Care Medicine·Huzaifa AdamaliSeamas C Donnelly
Mar 19, 2013·American Journal of Respiratory and Critical Care Medicine·Keven M Robinson, John F Alcorn
Jun 4, 2013·Journal of Cystic Fibrosis : Official Journal of the European Cystic Fibrosis Society·J L FothergillC Winstanley
Apr 13, 2015·Journal of Infection and Public Health·Hosseine AghamollaeiNastaran S Barzi
Jun 15, 2011·Journal of Cystic Fibrosis : Official Journal of the European Cystic Fibrosis Society·Franziska A StressmannKenneth D Bruce
Jun 21, 2011·Journal of Cystic Fibrosis : Official Journal of the European Cystic Fibrosis Society·Pieter DeschaghtMario Vaneechoutte
Aug 24, 2010·Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases·Mohammad Mehdi FeizabadiMahmood Parvin
Jan 4, 2012·FEMS Immunology and Medical Microbiology·Vibeke B RudkjøbingThomas Bjarnsholt
Feb 13, 2010·Clinical Microbiology and Infection : the Official Publication of the European Society of Clinical Microbiology and Infectious Diseases·G B RogersK D Bruce
Sep 30, 2010·Clinical Microbiology and Infection : the Official Publication of the European Society of Clinical Microbiology and Infectious Diseases·G Greub
Aug 5, 2017·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Aisling TynanSeamas C Donnelly
Jul 30, 2019·The Journal of Antimicrobial Chemotherapy·Leif TueffersHinrich Schulenburg
Dec 28, 2018·BMC Infectious Diseases·Gianmarco MangiaterraFrancesca Biavasco
Feb 12, 2020·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Yiqi CaoBaiyu Zhang
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Mar 12, 2021·Medical Mycology·Ruben MorillaCarmen de la Horra

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Methods Mentioned

BETA
PCR
electrophoresis
PCRs
genotyping

Software Mentioned

Generic

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