Direct Detection of Pseudomonas aeruginosa from Respiratory Samples of Children with Cystic Fibrosis

Abstract

Introduction: Pseudomonas aeruginosa is the most important cause of lung infection and major cause of morbidity and mortality among Cystic Fibrosis patients. Early detection of P. aeruginosa from clinical samples is a key element in patient management. Appropriate antimicrobial treatment will postpone the transition to chronic lung infection.
 Methods: Sputum and deep pharyngeal swab samples (DPS) of CF patients were inoculated into conventional agar plates and identified by Matrix-assisted laser desorption/ionization-time of flight mass spectrometry. DNA was extracted with a column based QIAamp DNA mini kit from the samples. Molecular detection was done by using 23S rDNA specific primers with Taqman probe for quantitative PCR (qPCR) and exoA gene specific primers for in house PCR.
 Results: A total number of 67 sputum and 33 DPS samples were included to the study. Median age of the patients was 6 (2-14) for DPS and 15(7-33) for sputum samples. Detection limit was 103 cfu/ml for qPCR which displayed 94% sensitivity and 93% specificity for sputum; 44% sensitivity and 100% specificity for DPS. In house PCR displayed 94% sensitivity and 93% specificity for sputum; 50% sensitivity and 100% specificity for DPS. Time reqired for conventional method was 48 hours, 6 hours for in- house PCR and 4 hours for qPCR.
 Conclusion: Although culture is a reliable detection technique, a more rapid and sensitive way to detect P. aeruginosa from CF airway samples is essential. In our study which culture is accepted as a gold standard; PCR results is promising especially for sputum samples. Validation of molecular methods is necessary before implementation in routine laboratories. Clinical implications of discrepant results between culture and PCR detection should be carefully evaluated and combining both approaches considering antibiotic treatment could be reasonable.