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Direct detection of Pseudomonas aeruginosa from patients with healthcare associated pneumonia by real time PCR |
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| Authors: | Mohammad Mehdi Feizabadi Araz Majnooni Bizhan Nomanpour Bahram Fatolahzadeh Nafiseh Raji Somayeh Delfani Minoo Habibi Soroor Asadi Mahmood Parvin |
| Institution: | 1. Office of Research and Development, National Exposure Research Laboratory, United States Environmental Protection Agency, 26 West Martin Luther King Dr., Cincinnati, OH 45268, United States;2. Department of Internal Medicine, University of Cincinnati, College of Medicine, 231 Albert Sabin Way, Cincinnati, OH 45229, United States;3. U.S. Geological Survey, Denver Federal Center, P.O. Box 25585, Denver, CO 80225, United States;4. U.S. Geological Survey, 400 S. Clinton Street, Iowa City, IA 52240, United States |
| 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 109 CFU/ml to 102 CFU/ml. Standard curve of duplicated every dilution had slope 3.25 ± 0.1 and R2 > 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 103 CFU/ml in cases with clinical symptoms. Our method can be used for quantitative detection of P. aeruginosa from BAL and sputum specimens in 1 h and 10 min. |
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