The interspecies transmission of avian influenza viruses remains a significant public health concern. H6 viruses have gained attention following the first human infection by a chicken-origin H6N1 virus (A/Taiwan/02/2013, Hu/13), highlighting their zoonotic potential. To understand the evolutionary trajectory and mammalian adaptation of this Taiwan lineage, we compared two avian isolates (A/Chicken/Taiwan/CF19/2009, Ck/09; A/Chicken/Taiwan/2267/2012, Ck/12) and Hu/13 in vitro and in vivo. Hu/13 exhibited enhanced replication in MDCK cells, producing larger plaques and higher viral titers than Ck/09 and Ck/12. In BALB/c mice, Hu/13 demonstrated the highest pathogenicity and mortality, followed by Ck/12, while Ck/09 induced minimal morbidity. Hu/13 and Ck/12 replicated efficiently in respiratory tissues, eliciting robust cytokine responses and severe pulmonary lesions. In ferrets, Hu/13 showed relatively efficient transmission, infecting all direct physical-contact and two out of three airborne-contact ferrets, whereas Ck/09 failed to transmit. Histopathology confirmed escalating lung pathology from Ck/09 to Ck/12 and Hu/13. Whole-genome sequencing identified adaptive mutations in Hu/13 during ferret replication, though no canonical mammalian-adaptive changes (e.g., PB2-E627K or HA-Q226L) were detected. These findings demonstrate progressive mammalian adaptation, replication efficiency, and transmissibility within the Taiwan H6N1 lineage. Enhanced surveillance is crucial to monitor mammalian-adaptive mutations, informing pandemic preparedness and public health strategies.