The study investigates the pathogenicity and transmissibility of a highly pathogenic H5N1 influenza virus isolated from infected cow milk in mice and ferrets. The virus spread systemically in both species, including to the mammary glands, and bound to sialic acids expressed in human upper airways. In mice, the virus caused rapid disease induction and systemic dissemination, with high virus titers in respiratory and non-respiratory tissues. In ferrets, the virus also showed elevated body temperatures and weight loss, with high virus titers in respiratory and non-respiratory organs. The virus was inefficiently transmitted to exposed ferrets, with one of four animals seroconverting without detectable virus. The virus also transmitted vertically from infected lactating mice to their pups, but not to adult contacts. The virus bound to both α2,3 and α2,6 sialic acids, indicating dual receptor-binding specificity. These findings suggest that the bovine H5N1 virus may have features that facilitate infection and transmission in mammals, highlighting the public health risk of the current outbreak.The study investigates the pathogenicity and transmissibility of a highly pathogenic H5N1 influenza virus isolated from infected cow milk in mice and ferrets. The virus spread systemically in both species, including to the mammary glands, and bound to sialic acids expressed in human upper airways. In mice, the virus caused rapid disease induction and systemic dissemination, with high virus titers in respiratory and non-respiratory tissues. In ferrets, the virus also showed elevated body temperatures and weight loss, with high virus titers in respiratory and non-respiratory organs. The virus was inefficiently transmitted to exposed ferrets, with one of four animals seroconverting without detectable virus. The virus also transmitted vertically from infected lactating mice to their pups, but not to adult contacts. The virus bound to both α2,3 and α2,6 sialic acids, indicating dual receptor-binding specificity. These findings suggest that the bovine H5N1 virus may have features that facilitate infection and transmission in mammals, highlighting the public health risk of the current outbreak.