Extended storage times are an ineffective mitigation strategy for partial inactivation of ASF virus in corn- and soybean-based ingredients
By Jerry Shurson, Christian Ramirez-Camba, Pedro Urriola, and Declan Schroeder, University of Minnesota
As the African swine fever virus (ASFV) pandemic continues to persist in many countries around the world, the potential for this virus to enter North America and the U.S. is an ongoing concern. Because of the ongoing challenges of developing effective ASFV vaccines and the lack of any effective treatments if pigs become infected, developing and implementing strict biosecurity programs remains the only remaining viable option to prevent ASFV introduction into the U.S. swine industry.
Although the probability of ASFV transmission through feed is low compared with direct exposure to infected pigs, carcasses, tissues, and body fluids, studies have shown that experimentally inoculated feed with ASFV can cause disease when fed to pigs (Niederwerder et al., 2019). There is also experimental evidence showing that ASFV inoculated feed ingredients, such as soybean meal, can survive and be infective when using simulated environmental conditions of Trans-Pacific and Trans-Atlantic shipping models (Dee et al., 2018). Therefore, it is plausible that importing feed ingredients from countries endemic with ASFV may be contaminated with virus and pose as a potential route of introduction into the U.S.
However, our quantitative risk assessment of the likelihood of ASFV contaminated soybean meal and corn being imported into the U.S. is once every 21 to 1,563 years for soybean meal and once every 66 to 5,000 years for corn (Schambow et al., 2022). The wide variation in these estimates is due to high uncertainty from the lack of data on the prevalence and concentration of ASFV contamination in imported feed ingredients. This is because we do not have a system in place to accurately measure, monitor, prevent, and control ASFV and other swine virus contamination in global feed supply chains. These results also indicate that there is a low probability of ASFV contamination in imported soybean meal and corn, but the likelihood of contamination is not zero. Therefore, because of lack of data, high uncertainty, some potential for contamination, and documented evidence that ASFV can survive for long periods of time in some feed ingredients, we need to continue to be diligent in implementing more effective biosecurity protocols in feed supply chains.
One of the most popular biosecurity strategies currently being implemented is the use of extended storage time of imported feed ingredients from countries that are endemic with ASFV to allow for virus inactivation if it is present. Extended storage time is a requirement for imported feed ingredients in Canada (https://inspection.canada.ca/en/animal-health/terrestrial-animals/diseases/reportable/african-swine-fever/plant-based-feed-ingredients), but not in the U.S. Although research studies have shown that this strategy is effective for partially inactivating some swine viruses, such as PEDV, its effectiveness for inactivating ASFV in soybean meal and corn is unclear based on the low specificity and sensitivity of methodologies that have been used in only a few studies that have evaluated this potential mitigation strategy. The limited number of studies to evaluate ASFV stability during long-term commercial storage conditions is a result of the very strict and extensive biosecurity requirements for ASFV, which are approved in only a few U.S. research laboratories including the University of Minnesota.
To overcome the many challenges of working with ASFV directly in research settings, our research team developed, validated, and patented a surrogate virus assay using a similar algal-based megavirus called Emiliania huxleyi virus (EhV) as a safe and accurate substitute for ASFV (Balestreri et al., 2024). We also use viability PCR instead of conventional PCR to measure the presence of virus particles remaining when subjected to different time and temperatures of exposure. This method of quantifying virus particles provides greater specificity and sensitivity than standard PCR used in previous studies. Using EhV and viability PCR, we discovered that EhV and ASFV not only respond similarly to time and temperature exposure, but they are both extremely resistant to inactivation at temperatures as high as 100°C (212°F). This temperature is much greater than the most extreme temperatures found in commercial feed ingredient storage facilities and suggests that extended storage time and temperature under practical conditions may not be effective for inactivating ASFV (Balestreri et al., 2024).
Therefore, with funding support from the United Soybean Board, we conducted a study to evaluate the effects of storage time (1, 5, 60, and 120 days) and temperature (4, 24, or 34°C or 39, 75, 93°F, respectively) on EhV inactivation in solvent extracted soybean meal, extruded soybean meal, soybean hulls, corn, dried distillers grains with solubles (DDGS), high protein dried distillers grains (HP-DDG), corn fermented protein (CFP), and complete feed. Results showed minimal EhV inactivation (about 0.2 log) across all feed ingredients, and there were no differences in EhV inactivation across all time and temperature conditions for all ingredients because the reductions observed were within the experimental error of the assay (Table 1). These results are in agreement with our previous study (Palowski et al. 2022) which showed that no degradation of EhV occurred in experimentally inoculated conventional and organic soybean meal following a 23-day commercial U.S. truck transport simulation.
We also analyzed each feed matrix for crude protein (7.8-49.3%), crude fat (0.8-7.27%), neutral detergent fiber (8.7-58.6%), and ash (1.28-5.69%) concentrations and water activity (0.32-0.70 aw) to determine if there were correlations between chemical composition of feed matrices with EhV inactivation. Ash content was linearly associated with average EhV concentration at all time points and temperatures combined, but this potential protective effect in feed ingredients did not exceed the experimental error (Figure 1). These results indicate that the observed effect may be attributed to experimental error rather than providing a true protective effect of EhV inactivation. No other correlations between chemical components and EhV inactivation were observed.
In summary, the key findings of our research indicate a need to re-evaluate biosecurity protocols and mitigation strategies to prevent the potential introduction and transmission of ASFV through feed.
Our results indicate that ASFV and EhV are more resistant to inactivation at high temperatures (100°C, 212°F) than previous studies have shown.
Storing corn- and soybean-based ingredients for up to 120 days at temperatures up to 93°F are not effective for inactivating EhV (a surrogate for ASFV).
Viable EhV (ASFV) in corn and corn co-products is as stable as it is in solvent extracted and extruded soybean meal and soybean hulls for up to 120 days at temperatures up to 93°F.
It is difficult to predict ASFV (EhV) stability or inactivation in corn and soybean-based ingredients based on crude protein, crude fat, neutral detergent fiber, and ash content or water activity.
Prevention of ASFV contamination of imported ingredients and using effective chemical mitigants such as formaldehyde, medium chain fatty acids, or glycerol monolaurate if feed ingredients are suspected to be contaminated are the best current strategies available (Niederwerder et al., 2020; Jackman et al., 2020).
References
Balestreri, C., Schroeder, D.C., Sampedro, F., Marqués, G., Palowski, A., Urriola, P.E., van de Ligt, J.L.G., Yancy, H.F., Shurson, G.C. 2024. Unexpected thermal stability of two enveloped megaviruses, Emiliania huxleyi virus and African swine fever virus, as measured by viability PCR. Virology Journal 21:1. doi: 10.1186/s12985-023-02272-z
Dee, S.A., Bauermann, F.V., Niederwerder, M.C., Singrey, A., Clement, T., de Lima, M., Long, C., Patterson, G., Sheahan, M.A., Stoian, A.M.M., Petrovan, V., Jones, C.K., De Jong, J., Ji, J., Spronk, G.D., Minion, L., Christopher-Hennings, J., Zimmerman, J.J., Rowland, R.R.R., Nelson, E., Sundberg, P., Diel, D.G. 2018. Survival of viral pathogens in animal feed ingredients under transboundary shipping models. PLoS One 13:e0194509. doi: 10.1371/journal.pone.0194509
Jackman, J.A., Hakobyan, A., Zakaryan, H., Elrod, C.C. 2020. Inhibition of African swine fever virus in liquid and feed by medium-chain fatty acids and glycerol monolaurate. Journal of Animal Science and Biotechnology 11, 114. doi: 10.1186/s40104-020-00517-3
Niederwerder M.C., Stoian, A.M.M., Rowland, R.R.R., Dritz, S.S., Petrovan, V., Constance, L.A., Gebhardt, J.T., Olcha, M., Jones, C.K., Woodworth, J.C., Fang, Y., Liang, J., Hefley, T.J. 2019. Infectious dose of African swine fever virus when consumed naturally in liquid or feed. Emerging Infectious Diseases 25:891–7. doi: 10.3201/eid2505.181495
Niederwerder, M.C., Dee, S., Diel, D.G., Stoian, A.M.M., Constance, L.A., Olcha, M., Petrovan, V., Patterson, G., Cino-Ozuna, A.G., Rowland, R.R.R. 2020. Mitigating the risk of African swine fever virus in feed with anti-viral chemical additives. Transboundary and Emerging Diseases 68, 477–486. doi: 10.1111/tbed.13699
Palowski, A., Balestreri, C,, Urriola, P.E., van de Ligt, J.L.G., Sampedro, F., Dee, S., Shah, A., Yancy, H.F., Shurson, G.C., Schroeder, D.C. 2022. Survival of a surrogate African swine fever virus-like algal virus in feed matrices using a 23-day commercial United States truck transport model. Frontiers in Microbiology 13:1059118. doi: 10.3389/fmicb.2022.1059118
Schambow, R.A., Sampedro F., Urriola P.E., van de Ligt J.L.G., Perez A., Shurson G.C. 2022. Rethinking the uncertainty of African swine fever virus contamination in feed ingredients and risk of introduction into the United States. Transboundary and Emerging Diseases 69:157–175. doi: 10.1111/tbed.14358
Shurson, G.C., Ramirez-Camba, C.D., Urriola, P.E., Schroeder, D.C. 2024. Stability of a surrogate African swine fever-like algal virus in corn- and soybean-based feed ingredients during extended storage and in vitro digestion processes. Frontiers in Veterinary Science 11:1498977. doi:10.3389/fvets.2024.1498977
Jerry Shurson, Christian Ramirez-Camba, Pedro Urriola, and Declan Schroeder are with the University of Minnesota
