Dataset Information

Point-of-care detection of Neisseria gonorrhoeae based on RPA-CRISPR/Cas12a.

ABSTRACT: Gonorrhea, caused by Neisseria gonorrhoeae (N. gonorrhoeae), is a persistent global public health threat. The development of low-cost, point-of-care testing is crucial for gonorrhea control, especially in regions with limited medical facilities. In this study, we integrated CRISPR/Cas12a reaction with recombinase polymerase amplification (RPA) to provide a simple and adaptable molecular detection method for N. gonorrhoeae. The RPA-Cas12a-based detection system developed in this study enables rapid detection of N. gonorrhoeae within 1 h without the use of specialized equipment. This method is highly specific for identifying N. gonorrhoeae without cross-reactivity with other prevalent pathogens. Furthermore, in the evaluation of 24 clinical samples, the detection system demonstrates a 100% concordance rate with traditional culture, which is being used clinically as a reference method. Overall, the RPA-Cas12a-based N. gonorrhoeae detection has the advantages of rapidity, portability, low-cost, no special equipment required, and strong operability, and has a high potential for application as a self-testing and point-of-care diagnosis, which is critical for the clinical management of gonorrhea in developing countries lacking medical equipment.

SUBMITTER: Tu Q

PROVIDER: S-EPMC10224890 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Point-of-care detection of Neisseria gonorrhoeae based on RPA-CRISPR/Cas12a.

Tu Qianrong Q Cao Xiaoying X Ling Chao C Xiang Lili L Yang Ping P Huang Shifeng S

AMB Express 20230527 1

Gonorrhea, caused by Neisseria gonorrhoeae (N. gonorrhoeae), is a persistent global public health threat. The development of low-cost, point-of-care testing is crucial for gonorrhea control, especially in regions with limited medical facilities. In this study, we integrated CRISPR/Cas12a reaction with recombinase polymerase amplification (RPA) to provide a simple and adaptable molecular detection method for N. gonorrhoeae. The RPA-Cas12a-based detection system developed in this study enables rap ...[more]

PMID: 37243708

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Project description:BackgroundCOVID-19 has spread rapidly around the world, affecting a large percentage of the population. When lifting certain mandatory measures for an economic restart, robust surveillance must be established and implemented, with nucleic acid detection for SARS-CoV-2 as an essential component.MethodsWe tried to develop a one-tube detection platform based on RT-RPA (Reverse Transcription and Recombinase Polymerase Isothermal Amplification) and DNA Endonuclease-Targeted CRISPR Trans Reporter (DETECTR) technology, termed OR-DETECTR, to detect SARS-CoV-2. We designed RT-RPA primers of the RdRp and N genes following the SARS-CoV-2 gene sequence. We optimized reaction components so that the detection process could be carried out in one tube. Specificity was demonstrated by detecting nucleic acid samples from pseudoviruses from seven human coronaviruses and Influenza A (H1N1). Clinical samples were used to validate the platform and all results were compared to rRT-PCR. RNA standards and pseudoviruses diluted by different gradients were used to demonstrate the detection limit. Additionally, we have developed a lateral flow assay based on OR-DETECTR for detecting COVID-19.ResultsThe OR-DETECTR detection process can be completed in one tube, which takes approximately 50 min. This method can specifically detect SARS-CoV-2 from seven human coronaviruses and Influenza A (H1N1), with a low detection limit of 2.5 copies/µl input (RNA standard) and 1 copy/µl input (pseudovirus). Results of six samples from SARS-CoV-2 patients, eight samples from patients with fever but no SARS-CoV-2 infection, and one mixed sample from 40 negative controls showed that OR-DETECTR is 100% consistent with rRT-PCR. The lateral flow assay based on OR-DETECTR can be used for the detection of COVID-19, and the detection limit is 2.5 copies/µl input.ConclusionsThe OR-DETECTR platform for the detection of COVID-19 is rapid, accurate, tube closed, easy-to-operate, and free of large instruments.

Dataset Information

Point-of-care detection of Neisseria gonorrhoeae based on RPA-CRISPR/Cas12a.

Publications

Point-of-care detection of Neisseria gonorrhoeae based on RPA-CRISPR/Cas12a.

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