Detection of pathogenic Leptospira with rapid extraction followed by recombinase polymerase amplification (RPA) and quantitative polymerase chain reaction (qPCR) assay-A comprehensive study from Sri Lanka.
Hansi UduwawalaAresha ManamperiGayana P S GunaratnaLilani KarunanayakeArianna CerutiAhmed Abd El WahedLakkumar FernandoRanjan PremaratnaMenaka HapugodaPublished in: PloS one (2024)
Leptospirosis is the most widespread zoonosis in the world. The disease is more prevalent in tropical regions where the majority of developing countries are located. Leptospirosis is considered a protean manifestation zoonosis with severity of the disease ranging from a mild febrile illness to a severe and life-threatening illness. Clinical symptoms of leptospirosis overlap with other tropical febrile illnesses. Early, rapid, and definitive diagnosis is important for effective patient management. Since Polymerase Chain Reaction (PCR)-based assays are not readily available in most clinical settings, there is a need for an affordable, simple, and rapid diagnostic test. Quantitative PCR (qPCR) and Recombinase Polymerase Amplification (RPA) were implemented at the Faculty of Medicine, University of Kelaniya, and a prospective study to evaluate RPA for diagnosis of acute phase of leptospirosis was conducted. Results indicate that RPA and qPCR were positive in 81% (98/121) of the total positive and acute clinical samples. Of the 81 positive MAT confirmed patients 60 (74%) and 53 (65%) were positive with qPCR and RPA respectively. Retrospective evaluation revealed a high diagnostic accuracy (sensitivity-70% and specificity-87%) of RPA compared to MAT as the reference gold standard. Results further suggest that there is no significant difference between the two assays, qPCR and RPA-SwiftX (P = 0.40). Laboratory procedures for the extraction and detection by qPCR in the laboratory have been optimized to obtain results within 6 hours. However, the RPA-SwiftX method under field conditions took 35 minutes. The RPA-SwiftX method could replace the qPCR which shows similar sensitivity and specificity. Therefore, RPA established under the current study presents a powerful tool for the early and rapid diagnosis of leptospirosis at point-of-care.
Keyphrases
- loop mediated isothermal amplification
- end stage renal disease
- climate change
- newly diagnosed
- real time pcr
- label free
- squamous cell carcinoma
- high resolution
- chronic kidney disease
- liver failure
- sensitive detection
- structural basis
- depressive symptoms
- urinary tract infection
- drug induced
- mass spectrometry
- quantum dots
- sleep quality
- acute respiratory distress syndrome
- prognostic factors
- medical students