Peptidoglycan- associated lipoprotein gene (oprL gene) as target for Taqman probe Detection of P. aeruginosa

Department of Microbiology, School of Medicine, Tehran University of Medical Sciences mfeizabadi@tums.ac.ir P. aeruginosa is an important nosocomial pathogen and in hospital associated pneumonia (HAP) is an emerging pneumonia category (Guidelines ATS/ISDA). Delay in HAP treatment is life threatening. We find out P. aeruginosa oprL gene (the peptidoglycan- associated lipoprotein: PAL, formerly protein H2) has conserved region that can be used for detection of P. aeruginosa by Taq man probe real time PCR. Despite the homology of C-terminal region of OprL from P. aeruginosa with P. putida (more than 60 %) and E. coli (40%), we designed primers and probe for species specific detection of our target organism with high sensitivity (100%) and specificity (98.85%). It could detect and enumerate 100 bacteria in specimens. The specificity of the primers were doubly checked against a panel of P. aeroginosa strains and other gram negative coccobacilli and gram positive cocci as well as the yeast that confirmed by culture methods as golden standard in the 101 bronco alveolar lavage (BAL) specimens. Negative predictive value (NPV) and Positive predictive value (PPV) were 100% and 99% respectively. Standard curve of duplicated every dilution had slope 3.25± 0.1 and R2 >0.99 with SD 0.1. Efficacy of PCR the test was 0.988. (E=10 (-1/slope)-1). We recommend to use real Time PCR for patient with sever hospital associated pneumonia as long as clinical findings.