Replication cycle of duck hepatitis A virus type 1 in duck embryonic hepatocytes.

Duck hepatitis A virus type 1 (DHAV-1) is an important agent of duck viral hepatitis. Until recently, the replication cycle of DHAV-1 is still unknown. Here duck embryonic hepatocytes infected with DHAV-1 were collected at different time points, and dynamic changes of the relative DHAV-1 gene expression during replication were detected by real-time PCR. And the morphology of hepatocytes infected with DHAV was evaluated by electron microscope. The result suggested that the adsorption of DHAV-1 saturated at 90 min post-infection, and the virus particles with size of about 50 nm including more than 20 nm of vacuum drying gold were observed on the infected cells surface. What's more, the replication lasted around 13 h after the early protein synthesis for about 5h, and the release of DHAV-1 was in steady state after 32 h. The replication cycle will enrich the data for DVH control and provide the foundation for future studies.

The duck hepatitis virus 5'-UTR possesses HCV-like IRES activity that is independent of eIF4F complex and modulated by downstream coding sequences.

Duck hepatitis virus (DHV-1) is a worldwide distributed picornavirus that causes acute and fatal disease in young ducklings. Recently, the complete genome of DHV-1 has been determined and comparative sequence analysis has shown that possesses the typical picornavirus organization but exhibits several unique features. For the first time, we provide evidence that the 626-nucleotide-long 5'-UTR of the DHV-1 genome contains an internal ribosome entry site (IRES) element that functions efficiently both in vitro and in mammalian cells. The prediction of the secondary structure of the DHV-1 IRES shows significant similarity to the hepatitis C virus (HCV) IRES. Moreover, similarly to HCV IRES, DHV-1 IRES can direct translation initiation in the absence of a functional eIF4F complex. We also demonstrate that the activity of the DHV-1 IRES is modulated by a viral coding sequence located downstream of the DHV-1 5'-UTR, which enhances DHV-1 IRES activity both in vitro and in vivo. Furthermore, mutational analysis of the predicted pseudo-knot structures at the 3'-end of the putative DHV-1 IRES supported the presence of conserved domains II and III and, as it has been previously described for other picornaviruses, these structures are essential for keeping the normal internal initiation of translation of DHV-1.

Classification of duck hepatitis virus into three genotypes based on molecular evolutionary analysis.

The nucleotide sequences of the complete VP1, VP0, VP3, and partial 3D regions of seven duck hepatitis virus (DHV) serotype 1 (DHV-1) strains isolated in China between 2001 and 2007 and one DHV-1 strain originally obtained from ATCC were determined and compared with previously available DHV sequences in GenBank. Phylogenetic analysis on the basis of VP1 sequences demonstrated three distinct genetic groups. There was an excellent concordance among the genetic groups assigned based on the complete VP0, VP3, and the partial 3D regions. In view of the growing importance of molecular techniques in diagnosis, we propose that the three genetic groups should be termed DHV types A, B, and C. All DHV-1 strains grouped in genotype A, whereas the new serotype strains isolated in Taiwan and the new serotype strains isolated in South Korea clustered into genotypes B and C, respectively, suggesting a potential genetic correlates of serotype. In pairwise comparisons of complete VP1, VP0, and VP3 nucleotide and amino acid sequences and the partial 3D nucleotide sequence, DHVs of the same genotype were clearly distinguished from those of heterologous genotypes. Analysis of the amino acid sequences of the three capsid proteins demonstrated the presence of conserved elements that form the eight-stranded beta-barrel structures, as well as intervening domains that vary in sequence between strains of different genotypes as seen in other picorna viruses.

Molecular analysis of duck hepatitis virus type 1 reveals a novel lineage close to the genus Parechovirus in the family Picornaviridae.

Duck hepatitis virus type 1 (DHV-1) was previously classified as an enterovirus, based primarily on observed morphology and physicochemical properties of the virion. The complete nucleotide sequences of two strains (DRL-62 and R85952) of DHV-1 have been determined. Excluding the poly(A) tail, the genomes are 7691 and 7690 nt, respectively, and contain a single, large open reading frame encoding a polyprotein of 2249 aa. The genome of DHV-1 is organized as are those of members of the family Picornaviridae: 5' untranslated region (UTR)-VP0-VP3-VP1-2A1-2A2-2B-2C-3A-3B-3C-3D-3' UTR. Analysis of the genomic and predicted polyprotein sequences revealed several unusual features, including the absence of a predicted maturation cleavage of VP0, the presence of two unrelated 2A protein motifs and a 3' UTR extended markedly compared with that of any other picornavirus. The 2A1 protein motif is related to the 2A protein type of the genus Aphthovirus and the adjacent 2A2 protein is related to the 2A protein type present in the genus Parechovirus. Phylogenetic analysis using the 3D protein sequence shows that the two DHV-1 strains are related more closely to members of the genus Parechovirus than to other picornaviruses. However, the two DHV-1 strains form a monophyletic group, clearly distinct from members of the genus Parechovirus.