The emergence and evolution of avian influenza A viruses (AIVs) pose significant challenges to both public health and animal husbandry worldwide. Here, we characterized a novel reassortant highly pathogenic avian influenza virus (HPAIV), clade 2.3.4.4b H5N6, that was isolated from a mandarin duck in South Korea in December 2023. Phylogenetic and molecular analyses show that the hemagglutinin (HA) gene of the 23-JBN-F12-36/H5N6 virus clustered with HPAIV clade 2.3.4.4b H5N1 viruses, which were circulating in South Korea and Japan in 2022–2023. The M and polymerase acidic (PA) genes also revealed a close association with the HPAIV clade 2.3.4.4b H5N1 AIV that was identified previously in South Korea during November 2022. Notably, the neuraminidase (NA) gene of the 23-JBN-F12-36/H5N6 virus was estimated to have its origins in the HPAIV clade 2.3.4.4h H5N6 prevalent in poultry in China, and it is clustered with the AIVs that are associated with human infection cases. Taken together, these results show that the virus has been produced by reassortment with H5N1 HPAIV, which is prevalent in wild birds; H5N6 HPAIV, which is circulated in poultry in China; and the internal genes of low pathogenic avian influenza viruses (LPAIVs). In light of the reassortment of HPAIVs circulating in existing wild birds and HPAIVs circulating in poultry in China within the 2.3.4.4b H5Nx clade, it is imperative to strengthen active surveillance across wild bird populations, poultry farms, and live poultry markets, and to inform for the effective design of improved prevention and control strategies.