An International, Inter-Laboratory Ring Trial Confirms the Feasibility of an Open Source, Extraction-Less “Direct” RT-qPCR Method for Reliable Detection of SARS-CoV-2 RNA in Clinical Samples

  • Publication Date :
  • Publication Type : Journal Article
  • Author(s) : Mills MG, Bruce EA, Huang ML, Crothers JW, Hyrien O, Oura CAL, Blake L, Brown A, Hester S, Wehmas L, Mari B, Barby P, Lacoux C, Fassy J, Vial P, Vial C, Martinez JRW, Oladipo OO, Inuwa B, Shittu I, Meseko CA, Chammas R, Santos CF, Dionisio TJ, Garbrieri TF, Parisi VA, Mendes-Correa MC, dePaula AV, Romano CM, Goes LGB, Minoprio P, Campos AC, Cunha MP, Vilela APP, Nyirenda T, Mkakosya RS, Muula AS, Dumm RE, Harris RM, Mitchell CA, Pettit S, Botten J, & Jerome KR
  • Journal Name : medRxiv

RT-qPCR is used world-wide to test and trace the spread of SARS-CoV-2. Extraction-less or direct RT-PCR is an open-access qualitative method for SARS-CoV-2 detection from nasopharyngeal (NP) or oral pharyngeal (OP) samples with the potential to generate actionable data more quickly, at a lower cost, and with fewer experimental resources than full RT-qPCR. This study engaged ten global testing sites, including laboratories currently experiencing testing limitations due to reagent or equipment shortages, in an international inter-laboratory ring trial. Participating labs were provided a common protocol, common reagents, aliquots of identical pooled clinical samples and purified nucleic acids, and used their existing in-house equipment. We observed 100% concordance across labs in the correct identification of all positive and negative samples, with highly similar Ct values observed. The test also performed well when applied to locally collected patient NP samples, provided the viral transport media did not contain charcoal or guanidine, both of which appeared to potently inhibit the RT-PCR reaction. Our results suggest that open access, direct RT-PCR assays are a feasible option for more efficient COVID-19 testing as demanded by the continuing pandemic.

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