Alternatives to Viral Transport Medium for use in SARS-CoV-2 Sample Preparation

Abstract

Introduction: As the world struggles to manage and move forward from the clinical effects of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the ability to test for viral genomic RNA in patient samples is critical. Currently, the development and performance of SARS-CoV-2 clinical tests is impaired by a diminished supply chain of reagents needed for the tests, compelling labs to seek alternative, readily-available reagents as substitutes. Aim: To evaluate the suitability of Phosphate-Buffered Saline (PBS) and RNAlater™ as substitutes for sample transport media, to preserve the fidelity of viral RNA for use in a SARSCoV-2 RT-PCR assay. Materials and Methods: This molecular study was conducted in Dayton, Ohio (USA) using synthetic materials and deidentified remnant patient specimens. Simulated standard clinical laboratory storage conditions were used, including prolonged storage up to 72 hours at 2-8°C and a freeze/thaw cycle. PCR amplification performance was measured for PBS and RNAlater™ against transport medium as a reference using purified viral RNA. Performance differences were determined using repeated-measures two-way ANOVA with a 5% false discovery rate. Results: Results indicate that both solutions were suitable for testing viral RNA in the short term, but the viral RNA stored in PBS began to degrade after just 24 hours at 2-8°C. In contrast, RNAlater™ preserved the viral RNA out to 72 hours when stored at 2-8°C, with no statistically significant decrease in the detection limits compared to freshly-prepared viral RNA dilutions. A single freeze/thaw cycle raised the lower limit of detection for RNAlater™-preserved viral RNA slightly. Conclusion: The current (as of April 2020) CDC sample guidelines permit the use of PBS, but have not published data to support this claim. These results offer an alternative to the transport options outlined in many Emergency Use Authorisations (EUA) currently authorised for use in diagnostic testing and may be used for possible long-term storage solutions for studies investigating SARS-CoV-2.

Keywords

• molecular diagnostic techniques
• sars virus
• specimen handling