A One Health Investigation into H5N1 Avian Influenza Virus Epizootics on Two Dairy Farms

Background: In early April 2024 we studied two Texas dairy farms which had suffered incursions of H5N1 highly pathogenic avian influenza virus (HPAIV) the previous month. Methods: We employed molecular assays, cell and egg culture, Sanger and next generation sequencing to isolate and characterize viruses from multiple farm specimens (cow nasal swab, milk specimens, fecal slurry, and a dead bird). Results: We detected H5N1 HPAIV in 64% (9/14) of milk specimens, 2.6% (1/39) of cattle nasal swab specimens, and none of 17 cattle worker nasopharyngeal swab specimens. We cultured and characterized virus from eight H5N1-positive specimens. Sanger and next-generation sequencing revealed the viruses were closely related into other recent Texas epizootic H5N1 strains of clade 2.3.4.4b. Our isolates had multiple mutations associated with increased spillover potential. Surprisingly, we detected SARS-CoV-2 in a nasal swab from a sick cow. Additionally, 14.3% (2/14) of the farm workers who donated sera were recently symptomatic and had elevated neutralizing antibodies against a related H5N1 strain. Conclusions: While our sampling was limited, these data offer additional insight into the large H5N1 HPAIV epizootic which thus far has impacted at least 96 cattle farms in twelve US states. Due to fears that research might damage dairy businesses, studies like this one have been few. We need to find ways to work with dairy farms in collecting more comprehensive epidemiological data that are necessary for the design of future interventions against H5N1 HPAIV on cattle farms. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This project was supported in part by the Agriculture and Food Research Initiative Competitive Grant from the American Rescue Plan Act, award number 2023-70432-39558, through USDA APHIS and Professor Gregory C. Grays startup funding from the University of Texas Medical Branch. The findings and conclusions in this presentation are those of the authors and should not be construed to represent any official USDA or US Government determination or policy. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: This research was approved by the University of Texas Medical Branch IRB, Protocol 23-0085 I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data needed to evaluate the conclusions in the paper are present in the paper or in the supplementary materials. Viral sequences will be deposited in NCBI when the manuscript is published in a peer-reviewed journal. Researchers with BSL3Ag-approved laboratories may request the live viruses isolated from milk and from a dead female great-tailed grackle (Quiscalus mexicanus) described in this paper by contacting Dr. Kenneth Plante, PhD, [email protected], of UTMBs World Reference Center for Emerging Viruses and Arboviruses. Additional data, study instruments, or specimens may be requested from the corresponding author. Farm location, farm identification and personal identifying data for human are protected by nondisclosure agreement and will not be shared. The sharing of specimens or data will require the signing of material transfer agreement.

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