Discrimination and isolation of the virus from free RNA fragments for the highly sensitive measurement of SARS-CoV-2 abundance on surfaces using a graphene oxide nano surface

Hyun Jin Yoo,Wen Ying Cui,Changyoon Baek,Yeon-Sook Kim,Wonseok Chung,Junhong Min,Yun Guang Li,Yong-Beom Shin

doi:10.1186/s40580-021-00281-8

Hyun Jin Yoo, Wen Ying Cui + Show 6 more

Open Access

https://doi.org/10.1186/s40580-021-00281-8

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Abstract

It is highly important to sensitively measure the abundance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on various surfaces. Here, we present a nucleic acid-based detection method consisting of a new sample preparation protocol that isolates only viruses, not the free RNA fragments already present on the surfaces of indoor human-inhabited environments, using a graphene oxide-coated microbead filter. Wet wipes (100 cm2), not cotton swabs, were used to collect viruses from environmental surfaces with large areas, and viruses were concentrated and separated with a graphene oxide-coated microbead filter. Viral RNA from virus was recovered 88.10 ± 8.03% from the surface and free RNA fragment was removed by 99.75 ± 0.19% from the final eluted solution. When we tested the developed method under laboratory conditions, a 10-fold higher viral detection sensitivity (Detection limit: 1 pfu/100 cm2) than the current commercial protocol was observed. Using our new sample preparation protocol, we also confirmed that the virus was effectively removed from surfaces after chemical disinfection; we were unable to measure the disinfection efficiency using the current commercial protocol because it cannot distinguish between viral RNA and free RNA fragments. Finally, we investigated the presence of SARS-CoV-2 and bacteria in 12 individual negative pressure wards in which patients with SARS-CoV-2 infection had been hospitalized. Bacteria (based on 16 S DNA) were found in all samples collected from patient rooms; however, SARS-CoV-2 was mainly detected in rooms shared by two patients.

Highlights

The outbreak of coronavirus disease of 2019 (COVID19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), began in China at the end of 2019 and has become a pandemic [1, 2]
Nucleic acid (NA)-based diagnosis, using ribonucleic acid (RNA) amplification, is currently one of the most popular methods to measure the viral abundance in both patients with suspected infections and the environment, because this method is more sensitive than antigen or antibody-based tools as it directly utilizes and amplifies the target viral RNA [12, 13]
3.1 Surface modification with graphene oxide Glass beads were coated with GO to separate viral RNA and free RNA fragment

Summary

Introduction

The outbreak of coronavirus disease of 2019 (COVID19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), began in China at the end of 2019 and has become a pandemic [1, 2]. Both the infectious pathogens and free RNA fragments were adsorbed onto nanostructured GO surface at low-pH condition. When 90% of the cells detached from the bottom, the virus was isolated by centrifugation at 3200×g for 10 min

New sample preparation protocol and current commercial protocol

Results and discussion

Investigation of surfaces artificially contaminated with HCoV‐229E

Disinfection methods

Conclusions