Inactivation of porcine epidemic diarrhea virus as a SARS-CoV-2 surrogate at sub-zero temperatures by isochoric freezing

Abstract

Given the resilience of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on frozen food, there is a risk that contaminated products could serve as vectors for viral transmission. Yet, methods capable of inactivating the virus at sub-zero temperatures without compromising the food's taste and quality are scarce. The high-pressure environment that arises spontaneously during isochoric freezing has demonstrated efficacy in suppressing or inactivating harmful microorganisms, such as bacteria; however, its effectiveness against coronavirus and the mechanisms involved remain unclear. In this study, we employed Porcine epidemic diarrhea virus (PEDV) as a proxy for SARS-CoV-2 to examine the effects of isochoric freezing on PEDV infectivity and to evaluate post-treatment alterations in the integrity of viral nucleic acids and envelopes, as well as changes in antigenic properties. Our experimental findings indicate that after a 6-h isochoric freezing treatment at −20 °C and 201 MPa, the titer decreased by 1.18 log₁₀(TCID₅₀/mL). While the viral nucleic acid remained intact post-treatment, the envelope's integrity was significantly impaired, accounting for the loss of infectivity. Moreover, the antigenic properties of the virus showed a slight increase following isochoric freezing treatment. This study pioneers the exploration of isochoric freezing technology's inactivation effects on coronaviruses, including preliminary mechanisms, offering novel perspectives for managing coronavirus contamination in frozen food and highlighting the potential of isochoric freezing in vaccine inactivation processes.