Department of Infectious Disease, College of Veterinary Medicine, University of Georgia, Athens, GA, USA.
Tifton Veterinary Diagnostic and Investigational Laboratory, University of Georgia, Athens, GA, USA.
Biotechniques. 2022 Jun;72(6):263-272. doi: 10.2144/btn-2022-0037. Epub 2022 May 12.
Coronavirus disease 2019 is a public health challenge requiring rapid testing for the detection of infections and transmission. Nucleic acid amplification tests targeting SARS coronavirus 2 (CoV2) are used to detect CoV2 in clinical samples. Real-time reverse transcription quantitative PCR is the standard nucleic acid amplification test for CoV2, although reverse transcription loop-mediated isothermal amplification is used in diagnostics. The authors demonstrate a sequence-specific reverse transcription loop-mediated isothermal amplification-based nucleic acid amplification assay that is finished within 30 min using minimally processed clinical nasal swab samples and describe a fluorescence-quenched reverse transcription loop-mediated isothermal amplification assay using labeled primers and a quencher oligonucleotide. This assay can achieve rapid (30 min) and sensitive (1000 plaque-forming units/ml) fluorescence detection of CoV2 (WA1/2020), B.1.1.7 (Alpha) and variants of concern Delta (B.1.617.2) and Omicron (B.1.1.529) in nasal samples.
2019 冠状病毒病是一项公共卫生挑战，需要快速检测以发现感染和传播。针对严重急性呼吸综合征冠状病毒 2 (SARS-CoV-2) 的核酸扩增试验用于检测临床样本中的 CoV2。实时逆转录定量 PCR 是 CoV2 的标准核酸扩增试验，尽管逆转录环介导等温扩增用于诊断。作者展示了一种基于序列特异性逆转录环介导等温扩增的核酸扩增检测方法，该方法使用经过最少处理的临床鼻拭子样本，在 30 分钟内完成，并描述了一种使用标记引物和淬灭寡核苷酸的荧光猝灭逆转录环介导等温扩增检测方法。该检测方法可在 30 分钟内快速（30 分钟）、灵敏（1000 噬菌斑形成单位/ml）地检测鼻样本中的 CoV2（WA1/2020）、B.1.1.7（Alpha）和关注的变异株 Delta（B.1.617.2）和奥密克戎（B.1.1.529）。
