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RT-RPA- Pf Ago detection platform for one-tube simultaneous typing diagnosis of human respiratory syncytial virus.
- Source :
-
Frontiers in cellular and infection microbiology [Front Cell Infect Microbiol] 2024 Jul 25; Vol. 14, pp. 1419949. Date of Electronic Publication: 2024 Jul 25 (Print Publication: 2024). - Publication Year :
- 2024
-
Abstract
- Human respiratory syncytial virus (HRSV) is the most prevalent pathogen contributing to acute respiratory tract infections (ARTI) in infants and young children and can lead to significant financial and medical costs. Here, we developed a simultaneous, dual-gene and ultrasensitive detection system for typing HRSV within 60 minutes that needs only minimum laboratory support. Briefly, multiplex integrating reverse transcription-recombinase polymerase amplification (RT-RPA) was performed with viral RNA extracted from nasopharyngeal swabs as a template for the amplification of the specific regions of subtypes A (HRSV <subscript>A</subscript> ) and B (HRSV <subscript>B</subscript> ) of HRSV. Next, the Pyrococcus furiosus Argonaute ( PfAgo ) protein utilizes small 5'-phosphorylated DNA guides to cleave target sequences and produce fluorophore signals (FAM and ROX). Compared with the traditional gold standard (RT-qPCR) and direct immunofluorescence assay (DFA), this method has the additional advantages of easy operation, efficiency and sensitivity, with a limit of detection (LOD) of 1 copy/μL. In terms of clinical sample validation, the diagnostic accuracy of the method for determining the HRSV <subscript>A</subscript> and HRSV <subscript>B</subscript> infection was greater than 95%. This technique provides a reliable point-of-care (POC) testing for the diagnosis of HRSV-induced ARTI in children and for outbreak management, especially in resource-limited settings.<br />Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.<br /> (Copyright © 2024 Liao, Feng, Zhang, Yang, Zhan, Zeng, Huang, Lian, Ke, Cai, Zhang, Fang, Cai, Chen, Lin, Lin, Chen, Liu, Huang, Lin and Lin.)
- Subjects :
- Humans
Infant
Pyrococcus furiosus genetics
Pyrococcus furiosus isolation & purification
Argonaute Proteins genetics
Molecular Diagnostic Techniques methods
Nucleic Acid Amplification Techniques methods
Limit of Detection
Nasopharynx virology
Respiratory Tract Infections diagnosis
Respiratory Tract Infections virology
Child, Preschool
Respiratory Syncytial Virus, Human genetics
Respiratory Syncytial Virus, Human isolation & purification
Respiratory Syncytial Virus Infections diagnosis
Respiratory Syncytial Virus Infections virology
Sensitivity and Specificity
RNA, Viral genetics
Subjects
Details
- Language :
- English
- ISSN :
- 2235-2988
- Volume :
- 14
- Database :
- MEDLINE
- Journal :
- Frontiers in cellular and infection microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 39119294
- Full Text :
- https://doi.org/10.3389/fcimb.2024.1419949