Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

Search BRD Papers

Clinical evaluation of the cobas SARS-CoV-2 test and a diagnostic platform switch during 48 hours in the midst of the COVID-19 pandemic.

Author(s): Poljak M, Korva M, Knap Gašper N, Fujs Komloš K, Sagadin M, Uršič T, Avšič Županc T, Petrovec M

Publication: J Clin Microbiol, 2020, Vol. , Page

PubMed ID: 32277022 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study compared detection of SARS-CoV-2 using LightMix assays which include a standard RNA extraction followed by real-time PCR-based assays and the fully-automated, sample-to-result Cobas 6800 method. Specimens were collected from 217 patients referred for SARS-CoV-2 testing using nasopharyngeal (211 specimens) or combined nasopharyngeal/oropharyngeal (6 specimens). Swabs were collected in the Universal Transport Medium System (UTM-RT). Immediately after arrival in the lab, RNA was extracted using the MagNA Pure Compact Nucleic Acid Isolation Kit I on a MagNa Pure Compact instrument and SARS-CoV-2 was detected using the LightMix Modular SARS and Wuhan CoV E-gene Kit (76 bp fragment of CoV E-gene) and the LightMix Modular Wuhan CoV RNA-dependent RNA RNA Polymerase (RdRP) Kit (100 bp fragment of RDRP). Results were considered positive if RdRP was positive and presumptive positive if E-gene was found without RdRP. Presumptive positives were asked to submit a second specimen. Specimens that were negative for SARS-CoV-2 were tested for the presence of influenza virus A (H1, H3, H5, and H7), influenza virus B (Yamagata and Victoria  lineages), respiratory syncytial virus (types A and B), rhinovirus (types A, B, and C), enterovirus (types A, B, C, and D), human bocavirus 1, human parainfluenza virus (types 1–4), human parechovirus (types 1–8), human adenovirus (groups B, C, and E, and some from groups A and D), human coronavirus (229E, HKU-1, NL63, and OC43), and human metapneumovirus (types A and B) using the RT-PCR based Respiratory Viruses 16-well assay. Specimens positive for coronavirus in this assay were further analyzed using in-house coronavirus type-specific one-step RT-PCR assays for coronavirus 229E, HKU-1, 132 NL63, and OC43. The remaining specimen in UTM-RT was frozen at -30°C for 1-17 days (median 3 days) and thawed 1 h before testing using the RT-PCR based Cobas 6800 system. Using the Cobas 6800 system, a positive was defined based on positive amplification of ORF1 and a presumptive positive was detection of the E gene only. Additional specimens from 502 patients [nasopharyngeal (489 patients) or combined 150 nasopharyngeal/oropharyngeal swab (13 patients)] in UTM-RT were transported to the laboratory (median time 1 h 32 min), immediately vortexed, and aliquoted for testing using the RT-PCR based Cobas 6800 method and the LightMix RT-PCR assays.

   Summary of Findings:

   Of the 217 patients in the initial study, 64 were found to have SARS-CoV-2, 17 had other coronaviruses (nine NL63, five 229E, two HKU-1, and one OC43), 46 others had other respiratory viruses (14 rhinovirus, nine respiratory syncytial virus, eight human metapneumovirus, eight influenza B virus, six influenza virus A, and one human parechovirus) and four had multiple viruses (RSV and hCoV, RSV and hRV,  hRV and hCoV-229E, and Flu A and hMPV). The remining 86 were negative for all viruses investigated. Diagnostic agreement between the LightMix RT-PCR and follow-up Cobas 6800 method (after frozen storage) was found in 98.1% of specimens (211 of 215) with three of four discordant diagnoses being positive in the LightMix assay but negative in the Cobas assay, all of which had low viral loads. The remaining specimen was positive by Cobas and negative by LightMix assay. In the 502 specimens analyzed by the two methods at the same time (no storage), there was 99.6% agreement (499 of 501) between the methods. The two discordant cases included one low positive using Cobas but negative using LightMix and a Cobas low presumptive positive/LightMix negative. CT values for SARS-CoV-2 were very strongly correlated (r²=0.96) between the assay types in the 63 patients with SARS-CoV-2.

   Biospecimens

   • Cell - Nasal cavity
   • Cell - Nasopharynx

   Preservative Types

   • None (Fresh)
   • Frozen

   Diagnoses:

   • Pneumonia/Respiratory Infection

   Platform:

   Analyte	Technology Platform
   RNA	Real-time qRT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Real-time qRT-PCR Specific	Technology platform	Standard RNA extraction real-time PCR (LightMix) Fully automated extraction and analysis (Cobas 6800)
   Storage	Storage duration	Not stored Frozen for 1-17 days

   View more