Inactivation of two SARS-CoV-2 virus surrogates by electron beam irradiation on large yellow croaker slices and their packaging surfaces

Abstract

The detection of infectious SARS-CoV-2 in food and food packaging associated with the cold chain has raised concerns about the possible transmission pathway of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in foods transported through cold-chain logistics and the need for novel decontamination strategies. In this study, the effect of electron beam (E-beam) irradiation on the inactivation of two SARS-CoV-2surrogate, viruses porcine epidemic diarrhea virus (PEDV) and porcine transmissible gastroenteritis virus (TGEV), in culture medium and food substrate, and on food substrate were investigated. The causes of virus inactivation were also investigated by transmission electron microscopy (TEM) and Quantitative Real-time PCR (QRT-PCR). Samples packed inside and outside, including virus-inoculated large yellow croaker and virus suspensions, were irradiated with E-beam irradiation (2, 4, 6, 8, 10 kGy) under refrigerated (0 °C)and frozen (-18 °C) conditions. The titers of both viruses in suspension and fish decreased significantly (P < 0.05) with increasing doses of E-beam irradiation. The maximum D₁₀ value of both viruses in suspension and fish was 1.24 kGy. E-beam irradiation at doses below 10 kGy was found to destroy the spike proteins of both SARS-CoV-2 surrogate viruses by transmission electron microscopy (TEM) and negative staining of thin-sectioned specimens, rendering them uninfectious. E-beam irradiation at doses greater than 10 kGy was also found to degrade viral genomic RNA by qRT-PCR. There were no significant differences in color, pH, TVB-N, TBARS, and sensory properties of irradiated fish samples at doses below 10 kGy. These findings suggested that E-beam irradiation has the potential to be developed as an efficient non-thermal treatment to reduce SARS-CoV-2 contamination in foods transported through cold chain foods to reduce the risk of SARS-CoV-2 infection in humans through the cold chain.

Keywords: Electron beam irradiation; Inactivation; PEDV; SARS-CoV-2 virus surrogates; TGEV.

Fig. 1

Schematic diagram of E-beam irradiation. Fig. 1 (a) is the samples of large yellow croaker in the foam box before irradiation, Fig. 1 (b) is the foam box used for irradiation, Fig. 1 (c) is the virus samples in the foam box before irradiation, Fig. 1 (d) is the virus samples on the outside surface of the foam box before irradiation, and Fig. 1 (e) is the E-beam irradiation workshop. The potassium dichromate (silver) dosimeters were irradiated along with fish and virus samples to measure the actual dose. Fig. 1(f) is schematic of sample packaging and E-beam irradiation process.

Fig. 2

Virus titers of PEDV(A) and TGEV(B) in large yellow croaker and DMEM samples by E-beam irradiation. Different lowercase letters in different groups indicate a significant difference (P < 0.05, n = 3).

Fig. 3

Copy number of viral RNA detected by quantitative Real-time PCR in DMEM and fish. (A) Viral RNA copy numbers at different irradiation doses (2, 4, 6, 8, 10 kGy, 0 kGy for control group). The a indicates PEDV viral RNA copy numbers inside the package; b indicates PEDV viral RNA copy numbers outside the package; c indicates TGEV viral RNA copy numbers inside the package; d indicates TGEV viral RNA copy numbers outside the package. (B) RNA copy numbers of two viruses at different irradiation doses (5, 10, 15, 20, 25, 30 kGy, 0 kGy is the control group). All tests were repeated three times.

Fig. 4

Transmission electron microscope observation on PEDV in Vero cells and TGEV in PK-15 cells by ultrathin sectioning a、b、c were PEDV treated with 0, 4.0, and 10.0 kGy of E-beam irradiation, respectively; d, e and f were TGEV treated with 0, 4.0, and 10.0 kGy of E-beam irradiation, respectively. The arrow direction is the fibrin out of the envelope of the virus.

Fig. 5

Transmission electron microscopy observation on PEDV and TGEV by negative staining a、b、c were PEDV treated with 0, 4.0, and 10.0 kGy of E-beam irradiation, respectively; d, e and f were TGEV treated with 0, 4.0, and 10.0 kGy of E-beam irradiation, respectively. The arrow direction is the fibrin out of the envelope of the virus.

Fig. 6

Color values (L*a*b*ΔE) of refrigeration (A) and freezing (B) large yellow croaker by E-beam irradiation. Different lowercase letters in different groups indicate a significant difference (P < 0.05, n = 3).

Fig. 7

Sensory evaluation of irradiated refrigeration (A) and freezing (B) large yellow croaker. Different lowercase letters in different groups indicate a significant difference (P < 0.05, n = 3).

Fig. 8

2-dimensional principal component analysis of irradiated large yellow croaker.

Fig. 9

TBARS (A), TVB-N (B) and pH (C) of large yellow croaker by E-beam irradiated. Different lowercase letters in different groups indicate a significant difference (P < 0.05, n = 3).