Subscribe to RSS
![](/products/assets/desktop/img/oa-logo.png)
DOI: 10.1055/s-0043-1760752
Performance Evaluation of Different RT-PCR Kits for the Direct Detection of SARS-CoV-2 in Preheated Specimens
Funding None.![](https://www.thieme-connect.de/media/10.1055-s-00044881/202303/lookinside/thumbnails/10-1055-s-0043-1760752_2251244-1.jpg)
Abstract
Background 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.
Methods 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 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.
Conclusion 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.
Publication History
Article published online:
30 January 2023
© 2023. The Indian Association of Laboratory Physicians. This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/)
Thieme Medical and Scientific Publishers Pvt. Ltd.
A-12, 2nd Floor, Sector 2, Noida-201301 UP, India
-
References
- 1 Gordon DE, Jang GM, Bouhaddou M. et al. A SARS-CoV-2 protein interaction map reveals targets for drug repurposing. Nature 2020; 583 (7816): 459-468
- 2 “World Health Organization (WHO) Coronavirus disease (COVID-19) Dashboard,” December 2020. Accessed October 28, 2022 at: https://covid19.who.int/
- 3 Davies NG, Abbott S, Barnard RC. et al; CMMID COVID-19 Working Group, COVID-19 Genomics UK (COG-UK) Consortium. Estimated transmissibility and impact of SARS-CoV-2 lineage B.1.1.7 in England. Science 2021; 372 (6538): x
- 4 Davies NG, Jarvis CI, Edmunds WJ, Jewell NP, Diaz-Ordaz K, Keogh RH. CMMID COVID-19 Working Group. Increased mortality in community-tested cases of SARS-CoV-2 lineage B.1.1.7. Nature 2021; 593 (7858): 270-274
- 5 Tegally H, Wilkinson E, Giovanetti M. et al. Detection of a SARS-CoV-2 variant of concern in South Africa. Nature 2021; 592 (7854): 438-443
- 6 Corman VM, Landt O, Kaiser M. et al. Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR. Euro Surveill 2020; 25 (03) 2000045
- 7 Visseaux B, Collin G, Houhou-Fidouh N. et al. Evaluation of three extraction-free SARS-CoV-2 RT-PCR assays: a feasible alternative approach with low technical requirements. J Virol Methods 2021; 291: 114086
- 8 Smyrlaki I, Ekman M, Lentini A. et al. Massive and rapid COVID-19 testing is feasible by extraction-free SARS-CoV-2 RT-PCR. Nat Commun 2020; 11 (01) 4812
- 9 Ganguli A, Mostafa A, Berger J. et al. Rapid isothermal amplification and portable detection system for SARS-CoV-2. Proc Natl Acad Sci U S A 2020; 117 (37) 22727-22735
- 10 Jalandra R, Yadav AK, Verma D. et al. Strategies and perspectives to develop SARS-CoV-2 detection methods and diagnostics. Biomed Pharmacother 2020; 129: 110446
- 11 Mahmoud SA, Ganesan S, Ibrahim E. et al. Evaluation of RNA extraction-free method for detection of SARS-CoV-2 in salivary samples for mass screening for COVID-19. BioMed Res Int 2021; 2021: 5568350
- 12 Nishimura N, Nakayama H, Yoshizumi S. et al. Detection of noroviruses in fecal specimens by direct RT-PCR without RNA purification. J Virol Methods 2010; 163 (02) 282-286
- 13 Herraez-Hernandez E, Alvarez-Perez M, Navarro-Bustos G. et al. HPV Direct Flow CHIP: a new human papillomavirus genotyping method based on direct PCR from crude-cell extracts. J Virol Methods 2013; 193 (01) 9-17
- 14 Li L, He JA, Wang W. et al. Development of a direct reverse-transcription quantitative PCR (dirRT-qPCR) assay for clinical Zika diagnosis. Int J Infect Dis 2019; 85: 167-174
- 15 Azmi I, Faizan MI, Kumar R. et al. A saliva-based RNA extraction-free workflow integrated with Cas13a for SARS-CoV-2 detection. Front Cell Infect Microbiol 2021; 11: 632646
- 16 Bruce EA, Huang ML, Perchetti GA. et al. Direct RT-qPCR detection of SARS-CoV-2 RNA from patient nasopharyngeal swabs without an RNA extraction step. PLoS Biol 2020; 18 (10) e3000896
- 17 Merindol N, Pépin G, Marchand C. et al. SARS-CoV-2 detection by direct rRT-PCR without RNA extraction. J Clin Virol 2020; 128: 104423
- 18 Hasan MR, Mirza F, Al-Hail H. et al. Detection of SARS-CoV-2 RNA by direct RT-qPCR on nasopharyngeal specimens without extraction of viral RNA. PLoS One 2020; 15 (07) e0236564
- 19 Lübke N, Senff T, Scherger S. et al. Extraction-free SARS-CoV-2 detection by rapid RT-qPCR universal for all primary respiratory materials. J Clin Virol 2020; 130: 104579
- 20 Kriegova E, Fillerova R, Kvapil P. Direct-RT-qPCR detection of SARS-CoV-2 without RNA extraction as part of a COVID-19 testing strategy: from sample to result in one hour. Diagnostics (Basel) 2020; 10 (08) 605
- 21 Wee SK, Sivalingam SP, Yap EPH. Rapid direct nucleic acid amplification test without RNA extraction for SARS-CoV-2 using a portable PCR thermocycler. Genes (Basel) 2020; 11 (06) 664
- 22 Chu AW, Chan WM, Ip JD. et al. Evaluation of simple nucleic acid extraction methods for the detection of SARS-CoV-2 in nasopharyngeal and saliva specimens during global shortage of extraction kits. J Clin Virol 2020; 129: 104519
- 23 Barza R, Patel P, Sabatini L, Singh K. Use of a simplified sample processing step without RNA extraction for direct SARS-CoV-2 RT-PCR detection. J Clin Virol 2020; 132: 104587
- 24 Brotons P, Perez-Argüello A, Launes C. et al. Validation and implementation of a direct RT-qPCR method for rapid screening of SARS-CoV-2 infection by using non-invasive saliva samples. Int J Infect Dis 2021; 110: 363-370
- 25 Pastorino B, Touret F, Gilles M, de Lamballerie X, Charrel RN. Heat inactivation of different types of SARS-CoV-2 samples: what protocols for biosafety, molecular detection and serological diagnostics?. Viruses 2020; 12 (07) 735
- 26 Alcoba-Florez J, González-Montelongo R, Íñigo-Campos A. et al; The Microbiology Technical Support Team. Fast SARS-CoV-2 detection by RT-qPCR in preheated nasopharyngeal swab samples. Int J Infect Dis 2020; 97: 66-68
- 27 Miranda JP, Osorio J, Videla M, Angel G, Camponovo R, Henríquez-Henríquez M. Analytical and clinical validation for RT-qPCR detection of SARS-CoV-2 without RNA extraction. Front Med (Lausanne) 2020; 7: 567572