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

Table 2 Comparison of the sensitivity of the different PCR formats for sputum dilutions extracted with easyMAG Generic 2.0.1 and proteinase K pretreatment

PCR format^a	Cycler^c	Primers	Probes	Annealing temperature (°C)^d	Last positive dilution
1. PCR + AGE^b	1	PAO1 S/PAO1 A	None	55	6
2. PCR + FCE	1	PAO1 S/PAO1 A	None	55	7
3. real-time PCR + SybrGreen	2	PAO1 S/PAO1 A	None	55	7
4. real-time PCR + HybProbes	2	oprL F/oprL R	oprL-LC-ROX/oprL-LC-FAM	57	8
5. real-time PCR + TaqMan probe^b	2	PAO1 S/PAO1 A	oprL TM	55	8
6. real-time PCR + TaqMan probe	3	Not specified	Not specified	60	8

^a AGE: Agarose gel electrophoresis + ethidium bromide staining; FCE: Fluorescent capillary electrophoresis on ABI310.
^b PCR formats that were used to compare the sensitivity of the different DNA-extraction protocols (Table 1).
^c 1: Veriti 96-Well Thermal Cycler, Applied BioSystems, Foster City, Ca.; 2: LightCycler 1.5, Roche, Basel, Switzerland; 3: ABI Prism 7000 Sequence Detection System, Applied BioSystems.
^d Annealing temperatures as specified by provider of primers and probes (PCR formats 1-5) or by provider of commercial kit (PCR format 6).

ISSN: 1471-2180