Jain Rajeev Kumar, Perumal Nagaraj, Chaurasia Deepti, Shrivastava Rakesh, Ahirwar Kamlesh Kumar, Sharma Archa, Kapoor Garima, Lalwani Jaya
State Virology Laboratory, Gandhi Medical College, Bhopal, Madhya Pradesh, India.
Department of Microbiology, Gandhi Medical College, Bhopal, Madhya Pradesh, India.
J Lab Physicians. 2023 Jan 30;15(3):383-391. doi: 10.1055/s-0043-1760752. eCollection 2023 Sep.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has created high demand for molecular kits and consumables for mass screening of suspected individuals. Direct real-time polymerase chain reaction (RT-PCR) assay without nucleic acid extraction has several advantages in saving testing time and cost and helps in the rapid reporting of SARS-CoV-2. The present study evaluated the analytical performance of four SARS-CoV-2 RT-PCR for direct RT-PCR testing using preheated specimens. A total of 100 clinical specimens were selected and divided into three different groups: (1) group I: 20 SARS-CoV-2 positive specimens with high viral load, viz., low Ct values (< 30 Ct), (2) group II: 50 SARS-CoV-2 positive specimens with low viral load, viz., high Ct values (> 30 Ct), and (3) group III: 30 SARS-CoV-2 negative specimens. Specimens were heat-inactivated at 70°C for 10 minutes and cooled down at 4°C and were evaluated for standard and direct RT-PCR method by using ViralDtect-II Multiplex Real-Time PCR kit, TaqPath COVID-19 Combo kit, COVIDsure Pro Multiplex RT-PCR kit, and Hi-PCR Coronavirus (COVID-19) Multiplex Probe PCR kit. Results showed that except ViralDtect-II kit, the other three TaqPath COVID-19 Combo kit, COVIDsure Pro kit, and Hi-PCR Coronavirus (COVID-19) RT-PCR kit were able to amplify all the SARS-CoV-2 genes in the direct RT-PCR method using preheated specimens. In group I specimens, 100% sensitivity was observed in all three RT-PCR kits. In group II specimens, COVIDsure Pro kit was found to be superior among other kits. Direct RT-PCR method during pandemic situation is valuable and cost effective for the detection of SARS-CoV-2. All three TaqPath COVID-19 Combo kit, COVIDsure Pro kit, and Hi-PCR Coronavirus (COVID-19) RT-PCR kit can be used for direct RT-PCR method and COVIDsure Pro kit performance was found to be superior among all.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）大流行使得对用于大规模筛查疑似个体的分子检测试剂盒和耗材的需求大增。无需核酸提取的直接实时聚合酶链反应（RT-PCR）检测在节省检测时间和成本方面具有诸多优势，有助于快速报告SARS-CoV-2检测结果。本研究评估了四种SARS-CoV-2 RT-PCR检测方法对预热样本进行直接RT-PCR检测的分析性能。
共选取100份临床样本并分为三组：（1）第一组：20份病毒载量高的SARS-CoV-2阳性样本，即低Ct值（<30 Ct）；（2）第二组：50份病毒载量低的SARS-CoV-2阳性样本，即高Ct值（>30 Ct）；（3）第三组：30份SARS-CoV-2阴性样本。样本在70°C加热灭活10分钟，然后在4°C冷却，使用ViralDtect-II多重实时PCR试剂盒、TaqPath COVID-19组合试剂盒、COVIDsure Pro多重RT-PCR试剂盒和Hi-PCR冠状病毒（COVID-19）多重探针PCR试剂盒对标准RT-PCR方法和直接RT-PCR方法进行评估。
结果显示，除ViralDtect-II试剂盒外，其他三种TaqPath COVID-19组合试剂盒、COVIDsure Pro试剂盒和Hi-PCR冠状病毒（COVID-19）RT-PCR试剂盒在使用预热样本的直接RT-PCR方法中均能扩增所有SARS-CoV-2基因。在第一组样本中，所有三种RT-PCR试剂盒的灵敏度均为100%。在第二组样本中，COVIDsure Pro试剂盒在其他试剂盒中表现更优。
在大流行期间，直接RT-PCR方法对于检测SARS-CoV-2具有重要价值且成本效益高。所有三种TaqPath COVID-19组合试剂盒、COVIDsure Pro试剂盒和Hi-PCR冠状病毒（COVID-19）RT-PCR试剂盒均可用于直接RT-PCR方法，且发现COVIDsure Pro试剂盒的性能在所有试剂盒中最为出色。
