Recombinase Polymerase Amplification (Rpa) As an Isothermal Molecular-Point of Care Test (Im-Poct) for Rapid Identification of P. Aeruginosa Among Patients With Respiratory Infection Publisher



R Azizian REZA ; E Jafari ERFANEH ; B Pourakabri BABAK ; S Mamishi SETAREH ; S Hoseinkhani SAMAN ; R Hosseinpoursadeghi REIHANEH
Source: Molecular Biotechnology Published:2025

Abstract

Acute Respiratory Infections (ARIs) are a leading cause of pediatric mortality, with Pseudomonas aeruginosa as a key pathogen. This study evaluated Recombinase Polymerase Amplification (RPA)-ELISA for rapid detection of P. aeruginosa in respiratory samples (n = 101). Compared to Real-Time PCR (98.8% sensitivity, 100% specificity), RPA-ELISA showed 97.6% sensitivity and 90% specificity, detecting pathogens at 41 ng/µL within 30 min. The method’s simplicity and speed support its use as a screening tool in resource-limited settings, though confirmatory testing remains advised. Respiratory samples (sputum, BAL, throat cultures) were collected from pediatric patients (August–December 2022). P. aeruginosa was identified via culture, biochemical tests, and Real-Time PCR (targeting gyrB). RPA-ELISA was optimized for P. aeruginosa detection using labeled primers/probes. Among 101 samples, 83 (82.2%) were P. aeruginosa. RPA-ELISA demonstrated 97.6% sensitivity (95% CI: 91.7–99.7%) and 90% specificity (95% CI: 55.5–99.7%) compared to Real-Time PCR (98.8% sensitivity, 95% CI: 93.5–100%; 100% specificity, 95% CI: 69.2–100%). The method detected P. aeruginosa at a concentration of 41 ng/µL with a 30-min turnaround time. Positive results were indicated by a color change in the ELISA wells, contrasting with negative controls. RPA-ELISA is a promising IM-POCT for P. aeruginosa, offering high sensitivity and rapid results. It is suitable for resource-limited settings but requires confirmatory testing due to moderate specificity. © 2025 Elsevier B.V., All rights reserved.