title: Direct detection of Pseudomonas aeruginosa from patients with healthcare associated pneumonia by real time PCR.
abstract: Using oprL sequences, a TaqMan real time PCR was developed and used for quantitative detection of Pseudomonas aeruginosa from 99 broncoalveolar lavage and 11 sputum specimens collected from patients with health care associated pneumonia. All specimens were cultured on appropriate media to isolate bacteria. Twenty five specimens were positive by both methods. Polymicrobial infections were found in 13 specimens. Amplification of oprL in serial dilutions ranged from 10(9)CFU/ml to 10(2)CFU/ml. Standard curve of duplicated every dilution had slope 3.25±0.1 and R(2)>0.99 with SD 0.1. Our real time PCR assay showed high sensitivity (100%) and specificity (98.85%). This technique could detect and enumerate 100 bacteria directly from clinical specimens and showed that the threshold is 10(3)CFU/ml in cases with clinical symptoms. Our method can be used for quantitative detection of P. aeruginosa from BAL and sputum specimens in 1h and 10min.
authors: Department of Microbiology, School of Medicine, Tehran University of Medical Sciences, Iran. mfeizabadi@sina.tums.ac.ir
authors: Mohammad Mehdi
authors: MM
authors: Feizabadi
authors: Araz
authors: A
authors: Majnooni
authors: Bizhan
authors: B
authors: Nomanpour
authors: Bahram
authors: B
authors: Fatolahzadeh
authors: Nafiseh
authors: N
authors: Raji
authors: Somayeh
authors: S
authors: Delfani
authors: Minoo
authors: M
authors: Habibi
authors: Soroor
authors: S
authors: Asadi
authors: Mahmood
authors: M
authors: Parvin
mesh_terms: D001992:Bronchoalveolar Lavage Fluid / Q000382:microbiology; D003428:Cross Infection / Q000175:diagnosis* / Q000382:microbiology; D004269:DNA, Bacterial / Q000032:analysis; D006801:Humans; D057230:Limit of Detection; D018410:Pneumonia, Bacterial / Q000175:diagnosis* / Q000382:microbiology; D016133:Polymerase Chain Reaction; D011552:Pseudomonas Infections / Q000175:diagnosis* / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000302:isolation & purification*; D011957:Receptors, Opioid / Q000235:genetics; D012680:Sensitivity and Specificity; D013183:Sputum / Q000382:microbiology; D000094942:Nociceptin Receptor
chemical_list: D004269:DNA, Bacterial; D011957:Receptors, Opioid; D000094942:Nociceptin Receptor