Determining the Stability of Human Betacoronavirus OC43 on Different Surfaces

Abstract

Scientists continue to study SARS‐CoV‐2, the virus currently responsible for a worldwide pandemic, to determine the best methods to inactivate the virus on surfaces and reduce the risk of contact transmission. Its classification as a Biosafety Level (BSL) ‐3 pathogen complicates research since few facilities have the capacity for this level of containment. Human betacoronavirus OC43 (OC43) is a good surrogate for SARS‐CoV‐2 based on its genetic similarity and OC43’s BSL‐2 classification. Two industry specific surfaces were inoculated with OC43 and sampled at timepoints that reflect typical industry timelines for sanitation. The objective was to determine the stability of OC43 on typical meat facility surfaces.OC43 was propagated on human ileocecal colorectal adenocarcinoma cells (HRT‐18, ATCC CCL‐244). Infectious virus was quantified by indirect immunofluorescence TCID50 assays on HRT‐18 cells. Stainless steel grade 316 and polyethylene (poly top) coupons were inoculated with 100 ul of 2.51 x 106 TCID50/ml virus by pipette in a grid pattern at room temperature (20ºC) in an active biosafety cabinet in triplicate for each timepoint. Coupons were rinsed with 5 ml of sterile filtered beef extract buffer (BEB7), composed of Tris base, glycine, magnesium chloride, and beef extract pH adjusted to 7.0, at 0, 1, 4, 8, and 16 hours post inoculation (hpi). The collected rinsate was aliquoted and stored at ‐80ºC until the TCID50 assays were conducted in batch. The 16 hpi stainless steel samples were created at the time of the poly top experiment. The TCID50 assays were run in duplicate for each sample and TCID50/ml was calculated and averaged for each replicate. Percent recovery of infectious virus was determined for each timepoint.Virus was observed to have partially dried at 1 hpi and was completely dry at all following timepoints on both surfaces. Few infectious virions were lost between inoculation and 0 hpi collection from both surfaces. Mean percent recovery (MPR) of infectious OC43 from stainless steel and poly top at 0 hpi was 153% and 90%, respectively. At 1 hpi, the MPR from stainless steel was 65% while poly top was 123%. The variability of the assay explains these findings. Stainless steel and poly top had average recoveries of 60% and 25% at 4 hours, respectively. The lowest MPR for stainless steel was 17% at 8 hpi while the lowest MPR for poly top was 7% at 16 hpi. A higher MPR of 43% from stainless steel at 16 hpi reflects this timepoint being conducted on a different day.Infectious OC43 is still detectable on common meat facility surfaces several hours after inoculation, but our current knowledge of enveloped viruses asserts that they are inviable after drying. A maximum decrease of only 1 log TCID50 between inoculation and the 16 hpi samples means that a person could potentially be infected between cleanings if it is possible to transfer dried infectious virus from a surface to a mucous membrane. However, this is a conjecture that requires further study since the infectious dose of SARS‐CoV‐2 is unknown and no experiment to determine if this contact transfer is possible has been conducted. Currently, similar experiments with SARS are underway.