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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 08; 129:104519.JC

Abstract

BACKGROUND

The severe shortage of nucleic acid extraction kits during the current COVID-19 pandemic represents a key limiting factor in testing capacity.

OBJECTIVES

This study compared the results of SARS-CoV-2 RT-PCR using different simple nucleic acid extraction methods on nasopharyngeal and saliva specimens.

STUDY DESIGN

Fifty nasopharyngeal swab and saliva specimens previously tested positive for SARS-CoV-2 were retrieved. Three different methods of nucleic acid extraction were compared. The first method involves incubating the specimen with proteinase K, and then heat treatment at 98 °C for 5 min (PKH); the second method involves heat treatment at 98 °C for 5 min without proteinase K pre-incubation (heat only); the third method involves no pre-processing steps (direct). The products from all 3 methods were tested by SARS-CoV-2 RT-PCR.

RESULTS

PKH had significantly higher positive rate in SARS-CoV-2 RT-PCR (80 %) than those of heat only (58 %; P = 0.001) or direct (56 %; P = 0.002). The median Ct value was significantly earlier for PKH (median Ct: 37.0, IQR 31.7-40) than that of heat only (median Ct: 40, IQR 36.2-41; P < 0.0001) and direct (median Ct, 37.5; IQR 33.9-41.0; P = 0.0049). Subgroup analysis showed that PKH had higher detection rate, lower limit of detection and earlier Ct values than the other two groups for both NPS and saliva specimens.

CONCLUSIONS

PKH pre-processing resulted in the highest detection rate of SARS-CoV-2 by RT-PCR, and represents an alternative method for nucleic acid extraction when commercial extraction kits are not available.

Authors+Show Affiliations

State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China.State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China.State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China.Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China.State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China.State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China.State Key Laboratory for Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; Department of Microbiology, Queen Mary Hospital, Hong Kong Special Administrative Region, China. Electronic address: kelvinto@hku.hk.

Pub Type(s)

Comparative Study
Evaluation Study
Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

32629187

Citation

Chu, Allen Wing-Ho, 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." Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology, vol. 129, 2020, p. 104519.
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.
Chu, A. W., Chan, W. M., Ip, J. D., Yip, C. C., Chan, J. F., Yuen, K. Y., & To, K. K. (2020). 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. Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology, 129, 104519. https://doi.org/10.1016/j.jcv.2020.104519
Chu AW, 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. PubMed PMID: 32629187.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - 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. AU - Chu,Allen Wing-Ho, AU - Chan,Wan-Mui, AU - Ip,Jonathan Daniel, AU - Yip,Cyril Chik-Yan, AU - Chan,Jasper Fuk-Woo, AU - Yuen,Kwok-Yung, AU - To,Kelvin Kai-Wang, Y1 - 2020/06/23/ PY - 2020/05/30/received PY - 2020/06/16/revised PY - 2020/06/21/accepted PY - 2020/7/7/pubmed PY - 2020/8/12/medline PY - 2020/7/7/entrez KW - COVID-19 KW - Detection KW - Nucleic acid extraction KW - Proteinase K KW - RT-PCR KW - SARS-CoV-2 SP - 104519 EP - 104519 JF - Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology JO - J. Clin. Virol. VL - 129 N2 - BACKGROUND: The severe shortage of nucleic acid extraction kits during the current COVID-19 pandemic represents a key limiting factor in testing capacity. OBJECTIVES: This study compared the results of SARS-CoV-2 RT-PCR using different simple nucleic acid extraction methods on nasopharyngeal and saliva specimens. STUDY DESIGN: Fifty nasopharyngeal swab and saliva specimens previously tested positive for SARS-CoV-2 were retrieved. Three different methods of nucleic acid extraction were compared. The first method involves incubating the specimen with proteinase K, and then heat treatment at 98 °C for 5 min (PKH); the second method involves heat treatment at 98 °C for 5 min without proteinase K pre-incubation (heat only); the third method involves no pre-processing steps (direct). The products from all 3 methods were tested by SARS-CoV-2 RT-PCR. RESULTS: PKH had significantly higher positive rate in SARS-CoV-2 RT-PCR (80 %) than those of heat only (58 %; P = 0.001) or direct (56 %; P = 0.002). The median Ct value was significantly earlier for PKH (median Ct: 37.0, IQR 31.7-40) than that of heat only (median Ct: 40, IQR 36.2-41; P < 0.0001) and direct (median Ct, 37.5; IQR 33.9-41.0; P = 0.0049). Subgroup analysis showed that PKH had higher detection rate, lower limit of detection and earlier Ct values than the other two groups for both NPS and saliva specimens. CONCLUSIONS: PKH pre-processing resulted in the highest detection rate of SARS-CoV-2 by RT-PCR, and represents an alternative method for nucleic acid extraction when commercial extraction kits are not available. SN - 1873-5967 UR - https://www.unboundmedicine.com/medline/citation/32629187/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_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1386-6532(20)30261-4 DB - PRIME DP - Unbound Medicine ER -