Abstract
Highly pathogenic avian influenza A(H5N1) virus exhibits a strong tropism for the bovine mammary gland, challenging our understanding of influenza A virus host range and tissue specificity. We performed experimental studies with an influenza A(H5N1) B3.13 genotype virus in female lactating dairy cattle to define the infectious dose, routes of exposure, and factors linked to morbidity and mortality. Here, we demonstrate that intramammary inoculation with as few as 10 TCID50 establishes a robust infection and shedding of high-titer virus in milk. Despite this low infectious dose, H5N1 does not readily transmit via contaminated milking equipment and close contact with infected animals. High-dose intramammary exposure results in severe disease and mortality, while respiratory and oral exposures are less likely to establish productive infection and associated morbidity. This study challenges current hypotheses of H5N1 transmission on dairy farms, raising important questions about potential agent, host, or environmental cofactors contributing to viral spread.
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Acknowledgements
We would like to thank PAAR facility staff (Alden Sewell, Kaitlynn Starr, Jonathan Zsoldos, and Michael Jeffers) for their support of this study. Thank you to Yehia Mohamed (Mo) Saif for his contributions to the calf study. We also thank John Franks, Jennifer DeBeauchamp, David Walker, Trushar Jeevan, and Tom Fabrizio for the initial isolation and characterization of the influenza A(H5N1) B3.13 genotype virus. We additionally thank the Ohio Department of Agriculture Animal Disease Diagnostic Laboratory (ODA ADDL) for performing whole genome sequencing on the B3.13 virus stock isolate. Figures supporting experimental design were created using https://BioRender.com and all data was graphed in Graphpad Prism (version 10.4.0).
Funding
Funding was provided by the Centers of Excellence for Influenza Research and Response (CEIRR), National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services under contract 75N93021C00016.
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Lee, C., Tarbuck, N.N., Cochran, H.J. et al. Dairy cows infected with influenza A(H5N1) reveals low infectious dose and transmission barriers. Nat Commun (2026). https://doi.org/10.1038/s41467-026-73490-6
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DOI: https://doi.org/10.1038/s41467-026-73490-6
