Duck hepatitis A virus (DHAV) causes acute hepatitis and mortality resulting in high economic losses in the duck farms industry. The current study describes the outbreak of DHAV in vaccinated duck farm in North Egypt during 2019 and molecular characterization of 3’utranslated region UTR and viral protein VP1 genes. Thirty samples was collected from commercial Pekin duck farms of 7-28 days old in 6 governorates showed a history of nervous signs and sudden deaths. DHAV was typing by RT-PCR for 3’UTR and VP1 genes revealed the 20 positive farms with first detection of DHAV genotype-3 in 18 samples and the classic genotype-1 in two samples only. The phylogenetic analysis of VP1, 3’UTR genes of the 9 selected strains representative to 6 governorates revealed that seven strains were clustered with DHAV-3 Chinese, Korean-Vietnamese within different subgroup with 92.4-93.7% amino acid (A.A.) identity, such strains were distinguishable from vaccine strain of DHAV-1 used in Egypt with 74.4% A.A. The other strains were closely related to the DHAV-1 Asian strain and the vaccine strain used in Egypt with 98.7-99.6% A.A. identity for VP1 gene with different clustering than recently isolated DHAV-1 Egyptian strains. The VP1 gene of DHAV-3 had one hypervariable region (HVR) with 10 amino acid mutations when compared with DHAV3/DN2/Vietnam/ 2011 but DHAV-1 had 3 HVR with one amino acid mutation in HVRII when compared with DHAV-1 vaccine strain. In conclusion, a new introduction of DHAV-3 with the classical DHAV-1 was recorded in Pekin duck farms in North Egypt that genetically distant from vaccinal strain.

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