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H5Nx clade 2.3.4.4 highly pathogenic avian influenza viruses (HPAIVs) have been disseminated to wide geographic regions since 2014. In 2016, five distinct genotypes (C-1 to C-5) of clade 2.3.4.4c H5N6 HPAIVs were detected in South Korea. In this study, we evaluated the pathogenicity, susceptibility to infection, and transmissibility of the two strains representing the C-1 and C-4 genotypes of the H5N6 viruses, which have different PA and NS gene, in domestic ducks. Although the susceptibility to infection of domestic ducks to the two strains was similar, the C-4 genotype virus induced higher mortality in ducks than C-1 genotype virus. A higher titer of viral shedding were detected in ducks challenged with the C-4 genotype virus compared with the C-1 genotype virus. These results indicated that the reassortment of HPAIVs with prevailing low pathogenic avian influenza viruses could effect on the pathogenicity in ducks. • The two genotypes of H5N6 HPAI viruses showed different pathogenicity in ducks. • The C-4 genotype virus induced higher mortality in ducks than C-1 genotype virus. • The reassortment of HPAIVs could effect on the pathogenicity in ducks. [ABSTRACT FROM AUTHOR]