Duck Hepatitis A Research Articles

MOLECULAR CHARACTERIZATION OF A DUCK VIRUS HEPATITIS ISOLATE ISOLATED FROM SHARKIA GOVERNORATE.

In the present study a Blencher duckling flock with 70% recurrent mortality on each reared batch was examined for the cause of mortality. live duckling were showing nervous signs and opisthotonos, sacrificed samples revealed depressed areas on the liver marking the large blood vessels while Hemorrhagic lesion suggestive for DVH were only noticed on liver of dead duckling carcasses. Liver samples were collected for virus isolation trials and for immunofluorescence. Initial virus isolation in embryonated chicken eggs ECE after ultra-filtration through 400 nm membrane filter revealed small size hemorrhagic edematous embryos this lesion was consistent upon a series of ultra-membrane filtration through 200 and 100 nm membrane filters which indicate a Picorna virus. Immunofluorescence examination revealed positive results for duck virus hepatitis (DVH). Clinical samples examined by generic RT-PCR assays followed by partial sequence analysis of the 3D gene revealed that the isolate was characterized as Duck hepatitis A virus resembling the strain (DHV/Duck / Egypt / Al-Gharbia /2014) in the 3D protein gene whom its gene bank accession number was (KP202874). The current study stresses on the value of ultra-membrane filtration through a 100 nm membrane filters as a rapid diagnostic tool For DVH without the need for extra laboratory diagnostic work. Interpretation of sequence results revealed that. The isolated (DVH/EG. Bayoumieh-Sharkia-2015) is a Duck hepatitis type A virus. With 100% resemblance to the strain (DHV/Duck/ Al-Gharbia /2014) Egypt/ in 3D protein gene with gene bank accession number (KP202874) unpublished data. Sequence and phylogenetic analysis indicated that the (DVH/EG. Bayoumieh-Sharkia-2015) isolate is clustered in the DHAV serotype 1 but was distinguishable from the other isolated Egyptian strains with resemblance ranging between 63.1 - 63.8%.

Identification of a conserved neutralizing linear B-cell epitope in the VP1 proteins of duck hepatitis A virus type 1 and 3

Duck virus hepatitis (DVH), mainly caused by duck hepatitis A virus (DHAV), is a severe disease threaten to duck industry and has worldwide distribution. As the major structural protein, the VP1 protein of DHAV is able to induce neutralizing antibody in ducks. In this study, a monoclonal antibody (mAb) 4F8 against the intact DHAV-1 particles was used to identify the possible epitope in the three serotypes of DHAV. The mAb 4F8 had weak neutralizing activities to both DHAV-1 and DHAV-3, and reacted with the conserved linear B-cell epitopes of 75GEIILT80 in DHAV-1 VP1 and 75GEVILT80 in DHAV-3 VP1 protein, respectively, while not with DHAV-2 VP1. This was the first report about identification of the common conserved neutralizing linear B-cell epitope of DHAV-1 and DHAV-3, which will facilitate understanding of the antigenic structure of VP1 and the serologic diagnosis of DHAV infection.

Mouse Models of Hepatitis B Virus Pathogenesis.

The host range of hepatitis B virus (HBV) is limited to humans and chimpanzees. As discussed in the literature, numerous studies in humans and chimpanzees have generated a great deal of information on the mechanisms that cause viral clearance, viral persistence, and disease pathogenesis during acute or chronic HBV infection. Relevant pathogenetic studies have also been performed in those few species representing natural hosts of hepadnaviruses that are related to HBV, such as the woodchuck hepatitis virus and the duck hepatitis virus. Further insight has been gained from multidisciplinary studies in transgenic or humanized chimeric mouse models expressing and/or replicating HBV to varying degrees. We provide here a concise summary of the available HBV mouse models as well as of the contributions of these models to our understanding of HBV pathogenesis.

Determine the structure of phosphorylated modification of icariin and its antiviral activity against duck hepatitis virus A.

BackgroundOur previous research showed that icariin (1) and its phosphorylated structural modification (2) improved the survival and attenuated oxidative stress and liver dysfunction induced by duck virus hepatitis. In this paper, we were one step closer to determine the structure of phosphorylation icariin (2) by the FT-IR, HRESIMS and 13C NMR. Anti-DHAV activities of 1 and 2 were compared in duck embryonic hepatocytes (DEHs) cultured in vitro and by artificial infection method in vivo. Additionally, the antiviral mechanisms of replication/release in vitro and the DHAV gene expression in vivo of 1 and 2 were analyzed.ResultsCompound 2's molecular formula was C33H42O18P. The results indicated that 1 and 2 effectively resisted DHAV invading DEHs, that they decreased the mortality of ducklings challenged with DHAV, and that 2 performed more effectively. 1 and 2 performed evenly on DHAV release; however, 2 restrained virus replication far more effectively. Since the anti-DHAV mechanisms of 1 and 2in vitro probably involve suppression of replication and release, 2’s better performance in anti-DHAV may result from its far more effectively inhibiting virus replication.ConclusionsThe compound 2's chemical structure was defined as 8-prenylkaempferol-4'-methylether-3-rhamnosyl-7-(6'''-phosphate)-glycoside. 1 and 2 exhibited anti-virus activity on DHAV. Our results suggest that 1 and 2 might become an anti-virus plant material candidate.

Effects of Bush Sophora Root polysaccharide and its sulfate on immuno-enhancing of the therapeutic DVH

Bush Sophora Root polysaccharide (BSRPS) and its sulfate, sulfated Bush Sophora Root polysaccharide (sBSRPS), possess the antiviral activities against duck hepatitis A virus. However their antiviral mechanisms are still not clear. This paper reported their immuno-enhancing roles in the therapeutic effects for duck virus hepatitis (DVH). The effects of BSRPS and sBSRPS on stimulating lymphocyte proliferation were investigated by MTT methods. After that, ducklings were challenged with DHAV and treated with BSRPS and sBSRPS. Meanwhile, the total antibody (Ab), cytokines including interferon gamma (IFN-γ), hepatocyte growth factor (HGF), interleukin (IL)-2, IL-6 and IL-8 were determined by enzyme-linked immuno sorbent assay methods. The results showed that BSRPS owned a fine hepatoprotective effect with stable HGF producing ability. Sulfated modification was able to increase the proliferation rates of B and T lymphocytes and the secretions of total Ab, IFN-γ and IL-2, as comparison with those of BSRPS group. In summary, both of them exhibited immuno-enhancing effects on the therapeutic effects for DVH, and the capacity of sBSRPS was stronger than that of BSRPS.

Genotyping of duck hepatitis virus type 3 isolated in Vietnam during 2009-2013

Genotyping of duck hepatitis virus type 3 isolated in Vietnam during 2009-2013

Efficient strategy for constructing duck enteritis virus-based live attenuated vaccine against homologous and heterologous H5N1 avian influenza virus and duck enteritis virus infection.

Duck is susceptible to many pathogens, such as duck hepatitis virus, duck enteritis virus (DEV), duck tembusu virus, H5N1 highly pathogenic avian influenza virus (HPAIV) in particular. With the significant role of duck in the evolution of H5N1 HPAIV, control and eradication of H5N1 HPAIV in duck through vaccine immunization is considered an effective method in minimizing the threat of a pandemic outbreak. Consequently, a practical strategy to construct a vaccine against these pathogens should be determined. In this study, the DEV was examined as a candidate vaccine vector to deliver the hemagglutinin (HA) gene of H5N1, and its potential as a polyvalent vaccine was evaluated. A modified mini-F vector was inserted into the gB and UL26 gene junction of the attenuated DEV vaccine strain C-KCE genome to generate an infectious bacterial artificial chromosome (BAC) of C-KCE (vBAC-C-KCE). The HA gene of A/duck/Hubei/xn/2007 (H5N1) was inserted into the C-KCE genome via the mating-assisted genetically integrated cloning (MAGIC) to generate the recombinant vector pBAC-C-KCE-HA. A bivalent vaccine C-KCE-HA was developed by eliminating the BAC backbone. Ducks immunized with C-KCE-HA induced both the cross-reactive antibodies and T cell response against H5. Moreover, C-KCE-HA-immunized ducks provided rapid and long-lasting protection against homologous and heterologous HPAIV H5N1 and DEV clinical signs, death, and primary viral replication. In conclusion, our BAC-C-KCE is a promising platform for developing a polyvalent live attenuated vaccine.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-015-0174-3) contains supplementary material, which is available to authorized users.

Epidemiology and molecular characterisation of duck hepatitis A virus from different duck breeds in Egypt

Duck hepatitis virus (DHV) is an acute highly contagious disease of ducklings caused by three distinct serotypes of duck hepatitis A virus (DHAV), a member of the RNA family Picornaviridae, where serotype 1 is the most widespread serotype worldwide. To date, little if any is known about the prevalence and genetic characterisation of DHAV outside Asia. The current study describes surveillance on DHV in 46 commercial duck farms in Egypt with a history of high mortality in young ducklings from 3 to 15 day-old from 2012 to 2014. Clinical samples were examined by generic RT-PCR assays followed by partial sequence analysis of the 5′UTR, VP1 and 3D genes of the vaccine strain and 15 field viruses. The overall positive rate was 37% (n=17/46). All duck breeds (Pekin, Muscovy, Mallard and Green Winged) were susceptible to the disease with mortality ranged from 15% to 96.7%. Sequence and phylogenetic analyses indicated that the Egyptian strains cluster in the DHAV serotype 1 with Asian viruses and distinguishable from the vaccine strains. So far, this is the first report on the genetic characterisation of DHAV in Egypt. This study may be useful to better understand the epidemiology and evolution of DHAV.

Identification and expression analysis of the interferon-induced protein with tetratricopeptide repeats 5 (IFIT5) gene in duck (Anas platyrhynchos domesticus).

The interferon-induced proteins with tetratricopeptide repeats (IFITs) protein family mediates antiviral effects by inhibiting translation initiation, cell proliferation, and migration in the interferon (IFN) dependent innate immune system. Several members of this family, including IFIT1, IFIT2, IFIT3 and IFIT5, have been heavily studied in mammals. Avian species contain only one family member, IFIT5, and little is known about the role of this protein in birds. In this study, duck IFIT5 (duIFIT5) full-length mRNA was cloned by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of the cDNA ends (RACE). Based on the sequence obtained, we performed a series of bioinformatics analyses, and found that duIFIT5 was most similar to homologs in other avian species. Also, duIFIT5 contained eight conserved TPR motifs and two conserved multi-domains (TPR_11 and TPR_12). Finally, we used duck hepatitis virus type 1 (DHV-1) and polyriboinosinicpolyribocytidylic acid (poly (I:C)) as a pathogen or a pathogen-associated molecular pattern induction to infect three-day-old domestic ducklings. The liver and spleen were collected to detect the change in duIFIT5 transcript level upon infection by quantitative real-time PCR (qRT-PCR). DuIFIT5 expression rapidly increased after DHV-1 infection and maintained a high level, while the transcripts of duIFIT5 peaked at 8h after poly (I:C) infection and then returned to normal. Taken together, these results provide a greater understanding of avian IFIT5.

Genetic characterization of a novel astrovirus in Pekin ducks

Three divergent groups of duck astroviruses (DAstVs), namely DAstV-1, DAstV-2 (formerly duck hepatitis virus type 3) and DAstV-3 (isolate CPH), and other avastroviruses are known to infect domestic ducks. To provide more data regarding the molecular epidemiology of astroviruses in domestic ducks, we examined the prevalence of astroviruses in 136 domestic duck samples collected from four different provinces of China. Nineteen goose samples were also included. Using an astrovirus-specific reverse transcription-PCR assay, two groups of astroviruses were detected from our samples. A group of astroviruses detected from Pekin ducks, Shaoxing ducks and Landes geese were highly similar to the newly discovered DAstV-3. More interestingly, a novel group of avastroviruses, which we named DAstV-4, was detected in Pekin ducks. Following full-length sequencing and sequence analysis, the variation between DAstV-4 and other avastroviruses in terms of lengths of genome and internal component was highlighted. Sequence identity and phylogenetic analyses based on the amino acid sequences of the three open reading frames (ORFs) clearly demonstrated that DAstV-4 was highly divergent from all other avastroviruses. Further analyses showed that DAstV-4 shared low levels of genome identities (50–58%) and high levels of mean amino acid genetic distances in the ORF2 sequences (0.520–0.801) with other avastroviruses, suggesting DAstV-4 may represent an additional avastrovirus species although the taxonomic relationship of DAstV-4 to DAstV-3 remains to be resolved. The present works contribute to the understanding of epidemiology, ecology and taxonomy of astroviruses in ducks.

Outbreak of duck viral hepatitis in duckling flocks in Poland - case report

Abstract The aim of the study was to determine the aetiologic agent causing deaths in two flocks of Pekin ducklings at the age of 12 d in Poland. on the basis of clinical symptoms and pathological changes, viral hepatitis infection was suspected in the birds. During the necropsy, liver sections were collected, from which total cellular RNA was isolated. Then, real-time polymerase chain reaction (RT-PCR) was performed using primers complementary to the pre-S region of the duck hepatitis virus genome. In all liver samples, the presence of a 530 bp PCR product was detected. The RT-PCR demonstrated the presence of genetic material of duck hepatitis virus type 1 (DH type 1) in the examined ducklings.

The CD8α gene in duck (Anatidae): cloning, characterization, and expression during viral infection.

Cluster of differentiation 8 alpha (CD8α) is critical for cell-mediated immune defense and T-cell development. Although CD8α sequences have been reported for several species, very little is known about CD8α in ducks. To elucidate the mechanisms involved in the innate and adaptive immune responses of ducks, we cloned CD8α coding sequences from domestic, Muscovy, Mallard, and Spotbill ducks using reverse transcription polymerase chain reaction (RT-PCR). Each sequence consisted of 714 nucleotides and encoded a signal peptide, an IgV-like domain, a stalk region, a transmembrane region, and a cytoplasmic tail. We identified 58 nucleotide differences and 37 amino acid differences among the four types of duck; of these, 53 nucleotide and 33 amino acid differences were between Muscovy ducks and the other duck species. The CD8α cDNA sequence from domestic duck consisted of a 61-nucleotide 5' untranslated region (UTR), a 714-nucleotide open reading frame, and an 849-nucleotide 3' UTR. Multiple sequence alignments showed that the amino acid sequence of CD8α is conserved in vertebrates. RT-PCR revealed that expression of CD8α mRNA of domestic ducks was highest in the thymus and very low in the kidney, cerebrum, cerebellum, and muscle. Immunohistochemical analyses detected CD8α on the splenic corpuscle and periarterial lymphatic sheath of the spleen. CD8α mRNA in domestic ducklings was initially up-regulated, and then down-regulated, in the thymus, spleen, and liver after treatment with duck hepatitis virus type I (DHV-1) or the immunostimulant polyriboinosinic polyribocytidylic acid (poly I:C).

Roles of the antioxidant properties of icariin and its phosphorylated derivative in the protection against duck virus hepatitis.

BackgroundDuck viral hepatitis (DVH) is an acute disease of young ducklings with few convenient and effective veterinary drugs to treat. In pathology, present study mainly focused on the immune mechanism, but very few studies have concerned with the role of oxidative stress in the pathogenesis of DVH. To study the antioxidative and hepatoprotective effects of icariin and its phosphorylated derivative against DVH, we prepared phosphorylated icariin (p-icariin) using the sodium trimetaphosphate–sodium tripolyphosphate method. Ducklings were drunk with icariin and p-icariin after being challenged with duck hepatitis virus 1 (DHV-1). We recorded the number of dead ducklings, gross pathological changes in the liver, and changes in indices of oxidative stress and liver injury. The correlations between these indices were also analyzed.ResultsExposure to DHV-1 induced significant oxidative damage in ducklings. Administration of icariin or p-icariin attenuated liver pathological injury and significantly increased the survival rate, with better outcomes in ducklings treated with p-icariin than in those treated with icariin. Icariin and p-icariin also attenuated the changes in oxidative stress and liver injury. We found positive correlations among indices of oxidative stress (malondialdehyde and inducible nitric oxide synthase) and liver injury (alanine aminotransferase, alkaline phosphatase, and lactate dehydrogenase), suggesting that DHV-1 causes significant oxidative damage, which is related to the extent of hepatic injury.ConclusionsIcariin and p-icariin improved the survival and attenuated oxidative stress and liver dysfunction induced by DHV-1. These outcomes were better in ducklings treated with p-icariin than in those treated by icariin. The clinical effects of both components were related to their antioxidant activities.

Detection, differentiation, and VP1 sequencing of duck hepatitis A virus type 1 and type 3 by a 1-step duplex reverse-transcription PCR assay

Duck hepatitis A virus (DHAV) is an infectious pathogen causing fatal duck viral hepatitis in ducklings. Although both the inactivated vaccines and live attenuated vaccines have been used to protect ducklings, DHAV-1 and DHAV-3 still cause significant serious damage to the duck industry in China and South Korea. For rapid detection, differentiation, and epidemic investigation of DHAV in China, a genotype-specific 1-step duplex reverse-transcription (RT) PCR assay was established in this study. The sensitivity and specificity of the developed RT-PCR assay was evaluated with nucleic acids extracted from 2 DHAV reference strains, and 9 other infectious viruses and bacteria. The genotype-specific primers amplified different size DNA fragments encompassing the complete VP1 gene of the DHAV-1 or DHAV-3. The assay detected the liver samples collected from experimentally infected ducklings and dead ducklings collected from different regions of China. Sequence analysis of these DNA fragments indicated that VP1 sequences of DHAV-1 can be used to distinguish wild type and vaccine strains. The phylogenetic analysis of VP1 sequences indicated that the developed RT-PCR assay can be used for epidemic investigation of DHAV-1 and DHAV-3. The developed RT-PCR assay can be used as a specific molecular tool for simultaneous detection, differentiation, and sequencing the VP1 gene of DHAV-1 and DHAV-3, which can be used for understanding the epidemiology and evolution of DHAV.

Molecular cloning and expression analysis of ferritin, heavy polypeptide 1 gene from duck (Anas platyrhynchos).

H-ferritin is a core subunit of the iron storage protein ferritin, and is related to the pathogenesis of malignant diseases. A differential expressed sequence tag of the ferritin, heavy polypeptide 1 gene (FTH1) was obtained from our previously constructed suppression subtractive cDNA library from 3-day-old ducklings challenged with duck hepatitis virus type I (DHV-1). The expression and function of FTH1 in immune defense against infection remains largely unknown in ducks. In this study, the full-length duFTH1 cDNA was obtained using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends. It consisted of 153 basepairs (bp) 5'untranslated region (UTR), 183 bp 3'UTR, and 546 bp open reading frame that encodes a single protein of 181 amino acid residues. duFTH1 shares high similarity with FTH1 genes from other vertebrates. The amino acid sequence possesses the conserved domain of typical ferritin H subunits, including seven metal ligands in the ferroxidase center, one iron binding region signature, and a potential bio-mineralization residue (Thy(29)). Moreover, in agreement with a previously reported ferritin H subunit, we identified an iron response element in the 5'UTR. RT-PCR analyses revealed duFTH1 mRNA is widely expressed in various tissues. Real-time quantitative polymerase chain reaction analyses suggested that duFTH1 mRNA is significantly up-regulated in the liver after DHV-1 injection or polyriboinosinic polyribocytidylic acid (polyI:C) treatment, reaching a peak 4 h post-infection, and dropping progressively and returning to normal after 24 h. Our findings suggest that duFTH1 functions as an iron chelating protein subunit in duck and contributes to the innate immune responses against viral infections.

Correction: DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

Correction: DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

Apoptosis induction in duck tissues during duck hepatitis A virus type 1 infection

To investigate the role of apoptosis in duck viral hepatitis pathogenesis, 4- and 21-d-old ducks were inoculated with duck hepatitis A virus serotype 1 and killed at 2, 6, 12, 24, and 48 h postinfection. TdT-mediated dUTP nick-end labeling was used to detect apoptosis cells. Expression profiles of apoptosis-related genes including caspase-3, -8, -9, and Bcl-2 in spleen, bursa of Fabricius, liver, and the quantity of virus in blood were examined using real-time PCR. The TdT-mediated dUTP nick-end labeling analysis indicated there was a significant difference of apoptotic cells between treatments and controls. The same difference also appeared in virus amount variation in blood during infection. Gene expression analysis revealed that the apoptosis-related gene expression profile was different in the 2 groups, and also different between various organs. This study suggested that apoptosis may play an important role in duck hepatitis A virus serotype 1 infection, and apoptosis suppression might facilitate virus multiplication, resulting in the highest virus concentration in the host.

Correction: DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

Correction: DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

Bush Sophora Root polysaccharide and its sulfate can scavenge free radicals resulted from duck virus hepatitis

In order to study the antioxidant effect of Bush Sophora Root polysaccharide (BSRPS) and its sulfate on anti-duck virus hepatitis (DVH), sulfated Bush Sophora Root polysaccharide (sBSRPS) was prepared by chlorosulfonic acid–pyridine method. Ducklings were fed with BSRPS and sBSRPS after challenged DHAV. Death was monitored, evaluation indexes of peroxidative and hepatic injury at the initial (4th and 8th hour) and later (54th hour) stages were detected. The results showed a fine treatment effect of BSRPS and sBSRPS. Visual hepatic pathological injury severities were less serious after the treatment. At the initial stage, free radical levels in all groups were the same, and BSRPS and sBSRPS reduced the hepatic injury through inhibiting virus replication. At the later stage, mass free radicals were detected in VC group while free radical levels in BSRPS and sBSRPS groups were significantly lower than VC group. The antioxidant effect of BSRPS and sBSRPS might alleviate the hepatic injury.

DNA Methylation and Regulation of the CD8A after Duck Hepatitis Virus Type 1 Infection

BackgroundCluster of differentiation 8 (CD8) is expressed in cytotoxic T cells, where it functions as a co-receptor for the T-cell receptor by binding to major histocompatibility complex class I (MHCI) proteins, which present peptides on the cell surface. CD8A is critical for cell-mediated immune defense and T-cell development. CD8A transcription is controlled by several cis-acting elements and trans-acting elements and is also regulated by DNA methylation. However, the epigenetic regulation of CD8A in the duck and its relationship with virus infection are still unclear.ResultsWe investigated the epigenetic transcriptional regulatory mechanisms, such as DNA methylation, for the expression of the CD8A and further evaluated the contribution of such epigenetic regulatory mechanisms to DHV-I infection in the duck. Real-time quantitative polymerase chain reaction (RT-qPCR) revealed the highest level of CD8A expression to be in the thymus, followed by the lungs, spleen, and liver, and the levels of CD8A expression were very low in the kidney, cerebrum, cerebellum, and muscle in the duck. RT-qPCR also demonstrated that the CD8A mRNA was down-regulated significantly in morbid ducklings treated with DHV-1 and up-regulated significantly in non-morbid ducklings in all the tissues tested. In addition, hypermethylation of CD8A was detected in the morbid ducklings, whereas relatively low methylation of CD8A was evident in the non-morbid ducklings. The CD8A mRNA level was negatively associated with the CpG methylation level of CD8A and global methylation status.ConclusionsWe concluded that the mRNA level of the CD8A was negatively associated with the CpG methylation level of CD8A and global methylation status in the duck, suggesting that the hypermethylation of CD8A may be associated with DHV-1 infection. The first two CpG sites of the CD8A promoter region could be considered as epigenetic biomarkers for resistance breeding against duckling hepatitis disease in the duck.