Comparative analysis of environmental persistence of SARS-CoV-2 variants and seasonal coronaviruses.

Conducting persistence studies of infectious severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on environmental surfaces may require a biosafety level 3 (BSL-3) laboratory. We aimed to compare the environmental persistence of BSL-2 level human coronaviruses (229E, NL63, and OC43) and bovine coronavirus (BoCoV) with three SARS-CoV-2 variants (WA-1, Delta, and Omicron). OC43 (1.8 TCID₅₀/mL) and BoCoV (1.0 TCID₅₀/mL) had lower detection thresholds in cell culture assays compared to 229E (150 TCID₅₀/mL) and NL63 (2,670 TCID₅₀/mL) and were used for persistence tests at room temperature. Viable OC43 became undetectable (>5.2log₁₀) after 48 hours on stainless steel and plastic coupons but exhibited extended persistence up to 72 hours on touchscreen glass coupons. In contrast, BoCoV remained viable for up to 120 hours with <1.8 log₁₀ infectivity loss. Both OC43 and BoCoV showed a reduction of >5 log₁₀ on vinyl coupons after 48 hours. On stainless steel coupons, the viability of all three SARS-CoV-2 variants became undetectable (>2.3 log₁₀ reduction) after 48 hours, with minor differences in reduction levels at 24 hours, whereas on touchscreen glass coupons, the viable virus could be detected for up to 48 hours for WA-1 and Omicron and 72 hours for the Delta variant. Regardless of coupon or virus type, viral RNA titers increased <4.5 Ct values after 120 hours. Our data demonstrate distinct persistence characteristics between BoCoV and OC43, with neither fully mimicking SARS-CoV-2 variants. This variability along with the impact of surface types on viral persistence underscores the need for caution when using these viruses as surrogates for SARS-CoV-2.IMPORTANCEIn this study, we evaluated three human seasonal coronaviruses (OC43, NL63, and 229E) and one bovine coronavirus (BoCoV) as potential surrogate viruses for SARS-CoV-2. Our data suggest that among the four surrogate viruses tested, OC43 and BoCoV were the most promising candidates due to their assay sensitivity, ease of handling, and high genetic similarity to SARS-CoV-2. However, neither BoCoV nor OC43 fully mimicked the environmental persistence characteristics of SARS-CoV-2 variants highlighting the potential limitations of using surrogate viruses.