key: cord-303832-1kcqhgjw
authors: Dai, Manman; Li, Huanan; Yan, Nan; Huang, Jinyu; Zhao, Li; Xu, Siqi; Jiang, Shibo; Pan, Chungen; Liao, Ming
title: Long-term survival of salmon-attached SARS-CoV-2 at 4°C as a potential source of transmission in seafood markets
date: 2020-09-06
journal: bioRxiv
DOI: 10.1101/2020.09.06.284695
sha: 
doc_id: 303832
cord_uid: 1kcqhgjw

Several outbreaks of COVID-19 were associated with seafood markets, raising concerns that fish-attached SARS-CoV-2 may exhibit prolonged survival in low-temperature environments. Here we showed that salmon-attached SARS-CoV-2 at 4°C could remain infectious for more than one week, suggesting that fish-attached SARS-CoV-2 may be a source of transmission.

ABSTRACT 21 Several outbreaks of COVID-19 were associated with seafood markets, raising concerns that 22 fish-attached SARS-CoV-2 may exhibit prolonged survival in low-temperature environments. 23 Here we showed that salmon-attached SARS-CoV-2 at 4 o C could remain infectious for more 24 than one week, suggesting that fish-attached SARS-CoV-2 may be a source of transmission. countries have identified SARS-CoV-2 in meat or meatpacking workers (3-7), raising concerns 31 that fish-or meat-attached SARS-CoV-2 could be a potential source of COVID-19 transmission.

Therefore, it is essential to determine the survival time of SARS-CoV-2 in the low-temperature 33 environment of seafood markets.

In this study, we detected the titer (50% tissue culture infectious dose/mL, TCID 50 /mL) of 35 viable SARS-CoV-2 attached on salmon or untreated SARS-CoV-2 in culture medium stored at 36 4°C, the temperature in refrigerators or cold rooms for the temporary storage of fish, or 25°C, the 37 regular room temperature, respectively, using end-point titration assay on Vero E6 cells as 38 described previously (8). As shown in Figure A and B, salmon-attached SARS-CoV-2 remained 39 viable at 4°C and 25°C for 8 and 2 days, respectively, while the untreated SARS-CoV-2 in 40 culture medium remained infectious at 4°C and 25°C for more than 8 days. SARS-CoV-2 41 attached on salmon or suspended in culture medium stored at 4°C remained viable for at least 8 42 days, while these stored at 25°C resulted in attenuating infectivity very quickly. The result from 43 the experiment on samples stored at 25°C is consistent with that reported by van Doremalen et al. 44 They showed that SARS-CoV-2 remained viable in aerosols, or on the surface of copper, 45 cardboard, stainless steel, and plastic, at 21~23°C and 40% relative humidity for 3 ~ 24 hours (8), 46 confirming that the loss of SARS-CoV-2 viability is associated with increased temperature.

Imported and exported fish must be transported under a low-temperature (e.g., 0 ~ 4°C ) In conclusion, fish-attached SARS-CoV-2 can survive for more than one week at 4°C, the 56 temperature of refrigerators, cold rooms, or transport carriers for storage of fish before selling in 57 the fish or seafood market. This calls for strict inspection or detection of SARS-CoV-2 as a 58 critical new protocol in fish importation and exportation before allowing sales. dashed lines indicate the limit of detection, which were 10 2 TCID 50 /mL. 

Aerosol and Surface Stability of 147

SARS-CoV-2 as Compared with SARS-CoV-1

Individual salmon cubes (5x5x5 mm) were placed in a 50 mL tube containing 13 mL liquid of 121 SARS-CoV-2 at 3.16x10 6 TCID 50 /mL, and the tube was gently inverted 5 times. The salmon 122 cubes were transferred into the 10 cm dish and incubated for 15 seconds at room temperature and 123 then put on filter paper in another 10 cm dish to remove the excess virus liquid. Salmon cubes 124 were transferred to 1.5 mL freezing tubes and stored at 4°C and 25°C, respectively. On day 1, 2, 125 3, 4, 5, 6, 7, 8, 10, and 12, respectively, one freezing tube was taken out, to which 1 mL DMEM 126 culture medium was added, oscillated for 5 seconds, and then centrifuged at 6,000 rpm for 5 127 minutes at 4°C. About 0.5 mL of the liquid was transferred to a new freezing tube and kept at -128 80°C until viral titration ( Figure A) . The untreated virus in culture medium was stored at 4°C and 129 25°C, respectively. On day 1, 3, and 8, 1 mL of the viral liquid was taken out and put in a 130 freezing tube, which was kept at -80°C until viral titration ( Figure A) . The virus titer (50% tissue 131 culture infectious dose, TCID 50 per mL) was quantified by end-point titration assay on Vero E6 132 cells (1). The detection limit of the typical TCID 50 assay used in this study was 10 2 TCID 50 / mL 133 (1).