Duck Reovirus Research Articles

Isolation and characterization of a duck reovirus strain from mature ducks in China

In 2018, a disease characterized by splenic hemorrhage and necrosis killed ducks in a duck farm in Guangxi province, China. A duck reovirus strain was isolated from the tissues of the dead ducks by inoculating duck embryos and BHK-21 cells. Electron microscopy of the cultured the isolate showed that the viral particles were nearly round in shape and approximately 70 nm in diameter, and they were designated DRV-GL18. Sequence analysis showed that the GL18 strain viral genome was 23,419 nt in length and had 10 dsRNA segments. Phylogenetic analysis of cDNA amplicons of segments encoding the protein σC which are outer capsid proteins showed that the isolate belongs to the branch of the epidemic strains of duck reovirus. The Recombination Detection Program (RDP) and SimPlot program analyses suggested potential genetic recombination events in the M2 segments. Pathogenicity experiments revealed that GL18 produced severe hemorrhaging in livers and necrosis in the spleen of infected SPF ducklings. A death rate of 50% in the experimental ducklings was calculated during the first 7 d, and the rest of the ducklings were observed to undergo spleen necrosis. These data suggested that GL18 is a duck reovirus isolate with severer pathogenicity, and it could be a candidate for development of vaccine. This is the first reported isolation of duck reovirus from mature ducks.

MiR-155-1 as a positive factor for novel duck reovirus replication by regulating SOCS5-mediated interferons

Diseases caused by novel duck reovirus (NDRV) have brought considerable economic losses to the poultry industry. MicroRNAs (miRNAs) have an impact on virus replication and antiviral immunity. However, the miRNA profile upon NDRV infection in duck embryo fibroblasts (DEFs) remains to be discovered. In this study, small RNA (sRNA) sequencing was performed to decipher the cellular miRNA response to NDRV infection. Based on 26 differentially expressed miRNAs (19 upregulated and 7 downregulated miRNAs) obtained from sequencing data and their target genes predicted by software, GO and KEGG analyses were performed to elucidate the functions of miRNAs in NDRV invasion, replication, and virus spread. “FoxO signaling pathway”, “autophagy”, and “Toll-like receptor signaling pathway” might participate in NDRV replication as revealed by KEGG enrichment analysis. The miR-155-1 sequence was found to be identical to rno-miR-155-5p and was sharply increased with the progression of NDRV infection. Moreover, NDRV-induced miR-155-1 could act as a positive factor for virus replication in DEFs, which inhibited type I interferon (IFN-I) production. Luciferase assay confirmed that miR-155-1 disturbed the abundance of suppressor of cytokine signaling (SOCS) 5 by targeting 3’-UTR. SOCS5, which is linked to increased IRF7 expression, restricts IFN expression and promotes NDRV replication in DEFs. Therefore, this study proposed that miR-155-1 was used by NDRV to restrict SOCS5 expression, attenuating the production of IFN-I and creating a favorable environment for virus replication.

Development and application of a multiplex qPCR assay for the detection of duck circovirus, duck Tembusu virus, Muscovy duck reovirus, and new duck reovirus.

A multiplex qPCR assay was developed to simultaneously detect duck circovirus (DuCV), duck Tembusu virus (DTMUV), Muscovy duck reovirus (MDRV), and novel duck reovirus (NDRV), but it did not amplify other viruses, including duck virus enteritis (DVE), infectious bursal disease virus (IBDV), avian reovirus (ARV), H5 avian influenza virus (H5 AIV), H7 avian influenza virus (H7 AIV), H9 avian influenza virus (H9 AIV), Newcastle disease virus (NDV), and Muscovy duck parvovirus (MDPV), and the detection limit for DuCV, DTMUV, MDRV, and NDRV was 1.51 × 101 copies/μL. The intra- and interassay coefficients of variation were less than 1.54% in the repeatability test with standard plasmid concentrations of 1.51 × 107, 1.51 × 105, and 1.51 × 103 copies/μL. The developed multiple qPCR assay was used to examine 404 clinical samples to verify its practicability. The positivity rates for DuCV, DTMUV, MDRV, and NDRV were 26.0%, 9.9%, 4.0%, and 4.7%, respectively, and the mixed infection rates for DuCV + DTMUV, DuCV + MDRV, DuCV + NDRV, MDRV + NDRV, DTMUV + MDRV, and DTMUV + NDRV were 2.7%, 1.2%, 1.2%, 1.0%, 0.5%, and 0.7%, respectively.

Epidemiological investigation of infectious diseases in geese on mainland China during 2018-2021.

Geese play an important role in agricultural economics, with China producing the vast majority of goose meat consumed worldwide annually. The variations in the avian viruses and co-infections result in substantial economic losses to the goose industry in China. To understand the evolutionary characteristics and co-infections of viruses, a broad epidemiological investigation of epizootic viruses of goose was carried out in nine provinces of China during 2018-2021. Here, the results indicated that, among the 1970 clinical samples, 50.81% (1001/1970) were positive for goose astrovirus (GAstV), 18.22% (359/1970) for avian orthoreovirus, 12.74% (251/1970) for goose parvovirus, 11.02% (217/1970) for H9N2 subtype avian influenza virus, 4.01% (79/1970) for Newcastle disease virus, and 2.08% (41/1970) for fowl adenovirus. Among the six viruses, co-infections comprised a large proportion (66.37%) in the field, of which the dual infection was more common. Additionally, phylogenetic analysis of GAstVs indicated that Chinese GAstVs had formed two distinct groups, that is, GAstV-1 and GAstV-2. GAstV-2 sub-genotype II-c had arisen as the dominant genotype in the whole country. Notably, all the H9N2-AIV isolated strains harboured the mammalian adaptation markers I155T, H183N, and Q226L (H3 numbering) in the HA gene, which promotes preferential binding to human-like α2-6-linked sialic acid receptors. And beyond that, we determined that the goose-origin Muscovy Duck Reovirus isolates, showing 51.7%-96% similarities to that of other waterfowl-origin orthoreovirus isolates in sequence homology analysis of the representative part of σC, are a new variant of waterfowl-origin orthoreovirus. These data provide valuable information about the prevalence of infectious diseases in geese on mainland China.

Development and application of a multiplex PCR method for simultaneous detection of waterfowl parvovirus, duck enteritis virus and goose astrovirus.

Waterfowl parvovirus, duck enteritis virus and goose astrovirus have become serious pathogens in waterfowl farming. Co-infections occasionally occur, and as a result, it ismuchharderto rapidlyandsimultaneously identify several pathogens using conventional PCR. According to the characteristics of the goose parvovirus (GPV) and muscovy duck parvovirus (MDPV) genome sequences, a universal PCR primer was designed using Rep1 as the target gene. The specific detection primers were designed based on the specific conserved regions of UL54 of the duck enteritis virus (DEV) gene and ORF1a of the goose astrovirus (GAstV) gene. The PCR reaction system and conditions were optimized, and the optimal annealing temperature was found to be 56.2℃. The volume ratio of the GPV-MDPV, GAstV and DEV primers (20μM) was 1:4:5. The established multiplex PCR detection method can simultaneously detect GPV, MDPV, DEV and GAstVwithin one reaction, and be negative for duck Tembusu virus, muscovy duck reovirus, duck hepatitis A virus type 3 and duck circovirus. The method with excellent sensitivity, specificity and repeatability was successfully applied to clinical samples, it is a useful platform for identifing co-infections of GPV, MDPV, DEV and GAstV in waterfowl.

Intergrated Transcriptomic and Proteomic Analysis Revealed the Differential Responses to Novel Duck Reovirus Infection in the Bursa of Fabricius of Cairna moschata.

The bursa of Fabricius is an immunologically organ against the invasion of duck reovirus (DRV), which is a fatal bird virus belonging to the Reoviridae family. However, responses of the bursa of Fabricius of Cairna moschata to novel DRV (NDRV) infection are largely unknown. Transcriptomes and proteomes of the samples from control and two NDRV strain (HN10 and JDm10) with different virulence were analyzed. Differentially expressed genes and differential accumulated proteins were enriched in the serine protease system and innate immune response clusters. Most of the immune-related genes were up-regulated under both JDm10/HN10 infections. However, the immune-related proteins were only accumulated under HN10 infection. For the serine protease system, coagulation factor IX, three chains of fibrinogen, and complements C8, C5, and C2s were significantly up-regulated by the HN10 infection, suggesting that the serine protease-mediated immune system might be involved in the resistance to NDRV infection. For the innate and adaptive immune system, RIG-I, MDA5, MAPK20, and IRF3 were significantly up-regulated, indicating their important roles against invaded virus. TLR-3 and IKBKB were only up-regulated in the liver cells, MAPK20 was only up-regulated in the bursa of Fabricius cells, and IRAK2 was only up-regulated in the spleen samples. Coagulation factor IX was increased in the bursa of Fabricius, not in the liver and spleen samples. The data provides a detailed resource for studying the proteins participating in the resistances of the bursa of Fabricius of duck to NDRV infections.

Genetic variation analysis of the sigma B protein gene of novel duck reovirus in southeastern China from 2011 to 2020

The novel duck reovirus (NDRV) disease first appeared in China in 2011. Infected ducks may be of various ages and breeds. Sigma B protein, which is the key component of NDRV's outer capsid, may trigger group-specific neutralizing antibodies linked to NDRV infection, pathogenicity, and immune defense. The sigma B protein gene of fourteen NDRV field strains was amplified by RT-PCR and cloned into the pMD-18 T vector for sequencing to examine the genetic variation in sigma B proteins of NDRVs in southeastern China between 2011 and 2020. The sigma B protein gene of the fourteen NDRV southeastern strains included in this analysis had 96.3 %–99.8 % nucleotide, and 96.2 %–99.7 % deduced amino acid sequence homology. Phylogenetic analysis revealed that the fourteen southeastern strains belonged to a well-supported lineage that included NDRV and Muscovy duck reovirus (MDRV) strains. However, Avian reovirus (ARV) formed a distinct genetic lineage in the gene tree. The sigma B protein gene sequences of NDRV strains found in southeastern China are substantially conserved, according to these findings. There is no significant geographical difference between NDRV southeastern strains and DRV strains in other regions of China. Our findings will add to the molecular epidemiological picture of NDRV strain spread in southeastern China between 2011 and 2020, laying the groundwork for potential in-depth research on vaccine collection and comprehensive prevention.

Interactions of Muscovy duck reovirus, gut microbiota, and host innate immunity: Transcriptome and gut microbiota analysis

It has been shown that Muscovy duck reovirus (MDRV) infection causes severe intestinal barrier damage and intestinal mucosal immune suppression. The health and balance of gut microbes is essential for the progression of intestinal infectious diseases. To investigate the interaction of MDRV, intestinal bacteria with host intestinal innate immunity, an MDRV contact-infection model was established in this study. High-throughput sequencing technology was used to sequence 16S rDNA and transcripts in ileal samples from experimental Muscovy ducklings. Our results suggest that intestinal opportunistic pathogens such as Streptococcus and Corynebacterium proliferated massively in MDRV-infected Muscovy ducklings. The body initiates antiviral and antibacterial immunity and actively fights the infection of gut microbes. The synthesis of peptidoglycan, lipopolysaccharide, and flagellin by intestinal bacteria activates the Toll-like receptor signaling pathway resulting in increased secretion of IFN-β, IL-1β, and IL-8. The RIG-I-like receptor signaling pathway is an important signaling pathway for the interaction between MDRV and the host. At the same time, we also observed that multiple genes in the JAK-STAT signaling pathway were significantly different. These genes are important targets for studying the immunosuppression caused by MDRV. In conclusion, we analyzed the interaction of MDRV, intestinal flora and host immune system during MDRV infection, which provides a basis for the further study on the mechanism of intestinal immunosuppression caused by MDRV.

Structure and expression identification of Cherry Valley duck IRF8

Interferon regulatory factor 8 (IRF8) is also known as interferon (IFN) consensus sequence binding protein (ICSBP), which plays an important role in IFN signal transduction. In this study, we cloned the full-length coding sequence of Cherry Valley duck IRF8 (duIRF8) and analyzed its structure. In addition, we tested the distribution of IRF8 in the tissues of healthy Cherry Valley ducks, and the changes in IRF8 expression levels in the tissues after virus infection. The results show that the open reading frame (ORF) of IRF8 is 1293 bp, encodes 430 amino acids, and have 3 conserved domains: the N-terminal DBD domain, the C-terminal IAD domain, and the NLS domain. Besides, from the analysis of the phylogenetic tree, it can be known that the duIRF8 has the highest homology with the anser cygnoides, and has less homology with the fish. Analyzing the distribution level of IRF8 in the tissues, it is found that the expression level of IRF8 in the liver of Cherry Valley duck is the highest. However, after infection with duck Tambusu virus, novel duck reovirus, and duck plague virus, the expression of IRF8 in the spleen and brain all showed up-regulation. These data indicate that IRF8 is involved in the host's innate immune response against virus in Cherry Valley duck.

Identification and antiviral effect of Cherry Valley duck IRF4

Interferon regulatory factor 4 (IRF4) is a multifunctional transcription factor that plays an important regulatory role in the interferon (IFN) signaling. IRF4 participates in the process of antivirus, Th cell differentiation and B cell maturation by regulating the expression of IFN and some lymphokines. In this study, Cherry Valley duck IRF4 (duIRF4) was cloned and its cDNA was analyzed. Expression of duIRF4 in a wide variety of tissues and changes in duIRF4 expression due to viral infection also was detected by quantitative real-time PCR. The results show that duIRF4 contains 1,341 bp of ORF encoding a protein with 446 amino acids and contains 3 domains: DNA-binding domain (DBD), IRF-association domain (IAD) and nuclear localization signal (NLS). Quantitative real-time PCR analysis showed that duIRF4 was evenly expressed in all tissues examined, with the highest expression in the spleen, followed by the bursa of Fabricius, and lower in the skin and brain. In addition, expression of duIRF4 in the brain and spleen was significantly upregulated after being infected by duck plague virus, duck Tembusu virus, and novel duck reovirus. These data suggest that duIRF4 may be involved in innate immune response.

Isolation and characterization of a naturally attenuated novel duck reovirus strain as a live vaccine candidate

Novel duck reovirus (NDRV) causes high morbidity in ducklings, and recovered ducklings are often remarkably stunted in growth. In this study, four NDRV strains were isolated from the NDRV outbreaks that occurred in different regions of Shandong province, China. The biological characteristics and pathogenicity of the four NDRV strains were elucidated, and the N20 was identified as a naturally attenuated strain. Three-day-old ducklings were immunized with live N20 strain (100 ELD50/duck), and challenged with 104.52 ELD50 of virulent N19 strain at 7 days post immunization. The vaccinated ducklings showed no evidence of clinical signs, gross and histopathological lesions, or loss of body weight, and 100 % protection against the virulent NDRV N19 infection. The NDRV-specific antibodies were generated in the immunized ducklings and could neutralize different NDRV strains. These results indicated that the N20 strain was a promising live attenuated vaccine candidate against highly pathogenic NDRV infection.

Isolation and characterization of a new goose orthoreovirus causing liver and spleen focal necrosis in geese, China.

Since July 2020, an infectious disease characterized by liver and spleen white focal necrosis has been spreading widely through geese farms in many regions of China. A novel goose orthoreovirus (GRV), designated GRV-GD2020, was isolated from the liver and spleen of dead geese. Phylogenetic analysis and sequence comparison revealed that all the genes of GRV-GD2020 clustered with other waterfowl-origin orthoreovirus. However, the gene constellation of GRV-GD2020 was not similar to that of any particular strain. Instead, the genomic segments of GRV-GD2020 showed 27.5-97.3% similarities to that of other waterfowl-origin orthoreovirus isolates. Waterfowl-origin orthoreovirus infections characterized by liver and spleen focal necrosis had not emerged in recent years. The re-emergence of the disease may be related to the recombination of the genome segments of Muscovy duck reovirus (MDRV), GRV, and new-type duck orthoreovirus. In summary, we determined that the GRV-GD2020 strain, causing goose liver and spleen focal necrosis, is a new variant of waterfowl-origin orthoreovirus.

Novel duck reovirus exhibits pathogenicity to specific pathogen-free chickens by the subcutaneous route

To study the pathogenicity of new duck reovirus (NDRV) to chickens, eighty 3-day-old SPF chickens were equally divided into two groups. The experimental group was inoculated with a NDRV challenge strain of 100 μL (10–5.00 ELD50/0.1 mL) by the subcutaneous (s.c.) route, and the control group was inoculated with 100 μL of sterile phosphate-buffered saline (PBS) by the same route. In the experimental group, chickens exhibited introflexion of claws, performing of splits, stunting syndrome, weight loss and death. Gross lesions such as enlargement and yellowish-white focal necroses were observed in the liver and spleen. Microscopic changes were typical including varying degrees of hepatocyte steatosis and necrosis, splenic lymphocyte necrosis, interstitial pneumonia. Viral loads were detected in lung, liver, heart, spleen, duodenum, burse and kidney. The liver and spleen viral loads remained a much higher level and maintained for a longer time, suggesting that these tissues might be the target organs. In summary, NDRV can cause systemic infections and death in chickens, which indicated that chickens may be infected by NDRV in poultry production.

Genomic sequences and pathogenic characteristics of two variant duck reoviruses associated with spleen necrosis

Emerging variant novel duck reovirus (NDRV) strains that cause spleen swelling and necrosis have seriously threatened the waterfowl industry since 2017. However, there is no report about the complete genomic sequence of emerging variant strains isolated from Cherry valley ducks. In this study, we acquired the complete genome sequences of two variant NDRV strains, SD19/6201 and SD19/6202, and analyzed their genetic and evolutionary relationship with other orthoreoviruses. The phylogenetic analysis of σC showed that all the Chinese NDRVs were clustered into two distinct branches. The SD19/6201 strain located in branch I with most of the Chinese NDRVs, while SD19/6202 was clustered in branch II with significantly different from the existing strains. Within the branch I, the NDRVs isolated in 2017 and thereafter clustered in a new subgroup. Comparison analysis of σC amino acid sequences indicated that ten amino acid differences were found between SD19/6201 and SD19/6202. Apart from the SD19/6201 and SD19/6202 strains, isolates in 2017 and thereafter had specific mutations at residues 132A, 138R, 158H, and 258A. These two NDRV strains showed different pathogenicity in SPF duck embryos and ducks. The viral loads in the spleen of infected ducks were significantly higher than those of other organs, which might be the reason why NDRV could cause obvious spleen necrosis in ducks. This study will help us to formulate effective prevention and control strategies against NDRV and enrich our understanding of the intra- and inter-species relationships of orthoreoviruses.

Rapid and visual detection of the emerging novel duck reovirus by using a specific and sensitive reverse transcription recombinase polymerase amplification method

Duck spleen necrosis disease (DSND) caused by Novel Duck Reovirus (NDRV), is an emerging infectious disease that causes severely threaten to duck industry. Currently, the popular conventional RT-PCR technique for detecting NDRV is time consuming. So, it is essential to develop a rapid and accurate molecular diagnosis techniques of the pathogen for the purpose to effective control of the disease. In our study, a simple, rapid and reliable detection method was developed by an isothermal reverse transcription-recombinase polymerase amplification (RT-RPA). The RT-RPA primers were designed targeting the S3 gene of NDRV, and its specificity was verified by testing a series of other waterfowl pathogens. A total of 20 field and experimental samples from infected ducklings were tested by the RT-RPA and compared with the results of the conventional RT-PCR and the quantitative RT-PCR simultaneously. The RT-RPA method could detect as little as 4.14 × 102 copies/μl of the target gene in the sensitivity analysis, which was 10×higher sensitive than the conventional RT-PCR. The major advantage of the RT-RPA method is that it could be performed as an isothermal reaction at 37 ℃ and completed within 20 min. In addition, no cross-reactivity was detected with other waterfowl-origin viruses. Also, the amplified products could be visualized faster, without the gel electrophoresis, by adding the SYBR Green I and observing them under an ultraviolet light. The newly developed RT-RPA method offers a simple, rapid and accurate for rapid detection of NDRV, which especially useful in on-site facilities and resource-limited areas.

DDX1 from Cherry valley duck mediates signaling pathways and anti-NDRV activity

Novel duck reovirus (NDRV) causes severe economic losses to the duck industry, which is characterized by hemorrhagic spots and necrotic foci of the livers and spleens. DEAD-box helicase 1 (DDX1) plays a critical role in the innate immune system against viral infection. However, the role of duck DDX1 (duDDX1) in anti-RNA virus infection, especially in the anti-NDRV infection, has yet to be elucidated. In the present study, the full-length cDNA of duDDX1 (2223 bp encode 740 amino acids) was firstly cloned from the spleen of healthy Cherry valley ducks, and the phylogenetic tree indicated that the duDDX1 has the closest relationship with Anas platyrhynchos in the bird branch. The duDDX1 mRNA was widely distributed in all tested tissues, especially in the duodenum, liver, and spleen. Overexpression of duDDX1 in primary duck embryo fibroblast (DEF) cells triggered the activation of transcription factors IRF-7 and NF-κB, as well as IFN-β expression, and the expression of the Toll-like receptors (TLR2, TLR3, and TLR4) was significantly increased. Importantly, after overexpressing or knocking down duDDX1 and infecting NDRV in DEF cells, duDDX1 inhibits the replication of NDRV virus and also regulates the expression of pattern recognition receptors and cytokines. This indicates that duDDX1 may play an important role in the innate immune response of ducks to NDRV. Collectively, we first cloned DDX1 from ducks and analyzed its biological functions. Secondly, we proved that duck DDX1 participates in anti-NDRV infection, and innovated new ideas for the prevention and control of duck virus infection.

Effects of Hericium erinaceus polysaccharide on immunity and apoptosis of the main immune organs in Muscovy duck reovirus-infected ducklings

To investigate the effects of Hericium erinaceus polysaccharide (HEP) on immunity in Muscovy duck reovirus (MDRV)-infected ducklings and explore its mechanism of action, an MDRV contact-infection model was established. Then, we investigated the influence of HEP on morphology of main immune organs in MDRV-infected ducklings by HE staining, while antioxidant capacity (T-AOC, MDA), serum protein levels (TP, ALB, GLO), complement levels (C3, C4) and antibody levels (IgA, IgM, IgG) were detected. Apoptotic indexes (apoptosisi rate and FAS-L) were also quantified by TUNEL method and immunohistochemical staining. Meanwhile, FADD and CytC (apoptosis-related genes), were tested by quantitative RT-PCR. Results showed that HEP could reduce the injuries of immune organs caused by MDRV. Additionally, HEP markedly diminished MDA (p < 0.01), while significantly increased T-AOC, TP, ALB, GLO, C3, C4, IgA, IgM and IgG (p < 0.01 or p < 0.05). Then, HEP shifted apoptosis time to an early MDRV-infected stage and reduced apoptosis at later MDRV-infected stage. This was associated with changes of FADD and CytC. Collectively, our data suggested that HEP could reduce the immunesuppression by many ways, such as decreasing organs' injuries, improving antioxidant capacity, serum proteins levels, antibody levels and complement levels, while diminish the apoptosis by lowering the FADD and CytC.

Muscovy duck reovirus promotes virus replication by inhibiting autophagy-lysosomal degradation pathway

Autophagy plays a momentous role in cellular responses against pathogens. However, the influence of the autophagy machinery on Muscovy duck reovirus (MDRV) infection is not yet confirmed. In this study, it was shown that MDRV infection significantly increased the number of autophagy-like vesicles in DF-1 cells under electron microscope and the LC3-I/LC3-II conversion, which was considered important indicators of autophagy. It was worth noting that the level of autophagy was positively correlated with MDRV replication. Further test results showed that MDRV-induced autophagy can promote virus replication in DF-1 cells, and both the envelope protein sigma A and non-structural protein sigma NS that play an important role in virus replication process can colocalize with the autophagosome marker molecule LC3-II by confocal immunofluorescence analysis. These results indicated that MDRV utilized the autophagosomes for replication. Through transfection of the dual fluorescent plasmid mcherry-EGFP-LC3 and fluorescence microscope observation, it was found that autophagosomes were more likely to fuse with lysosomes in MDRV-infected cells compared with the blank group. The phenomenon of pEGFP-LC3B fluorescent spot and LAMP1 co-localization appeared in MDRV infected cells, indicating that MDRV infection would promote the fusion of autophagosomes and the lysosomes. Conversely, accumulation of p62 was observed by immunoblotting, suggesting that autolysosomes does not exert effective degradation. MDRV infection triggered a incomplete autophagic response. Further studies found that the expression of LAMP1, a marker protein of late endosome/early lysosome, increased significantly in MDRV-infected cells, suggesting an increase in the number of immature lysosomes. In addition, the experiment detected the maturation of the lysosomal acid hydrolase Cathepsin D in the cells, and found that the expression of the 33 kDa mature form of Cathepsin D was significantly reduced after MDRV infection, indicating that MDRV inhibits the maturation of lysosomes. In general, MDRV infection induces autophagy of DF-1 cells, promotes the fusion of autophagosomes and lysosomes, inhibits autophagolysosome degradation, and promotes virus replication.

Quantitative Proteomic Analysis of Duck Embryo Fibroblasts Infected With Novel Duck Reovirus.

The novel duck reovirus (NDRV) can cause hemorrhage and necrosis on the spleen of Pekin ducks; this disease has resulted in great economic losses to the duck industry. However, the molecular pathogenesis of NDRV remains poorly understood. In the current study, the quantitative proteomic analysis of NDRV-infected duck embryo fibroblasts was performed to explore the cellular protein changes in response to viral infection through iTRAQ coupled with the liquid chromatography (LC)–tandem mass spectrometry (MS/MS) method. A total of 6,137 proteins were obtained in cell samples at 24 h post-infection. Of these, 179 differentially expressed proteins (DEPs) were identified (cutoff set to 1.5-fold change), including 89 upregulated and 90 downregulated proteins. Bioinformatics analysis showed that DEPs can be divided into the cellular component, molecular function, and biological process; they were mainly involved in signal transduction, infectious diseases, cell growth and death, and the immune system. The subcellular localization of most proteins was in the cytoplasm. Importantly, the expressions of signal transducer and activator of transcription 1 (STAT1) and various interferon-stimulated genes (ISGs) were upregulated after NDRV infection. The mRNA transcripts of some ISGs were consistent with proteomic data, showing an increased trend. Results of our study suggested that NDRV infection can elicit strong expression changes of cellular proteins and activate the expression of ISGs from the point of quantitative proteomic analysis. The study provides a new insight into the understanding of NDRV pathogenesis.

Duck HMGB2 Mediates Signaling Pathways in the Innate Immunity of Hosts Against Viral Infections.

High-mobility group box 2 (HMGB2) belongs to the HMG-box family that participates in a variety of biologic processes. Recent studies have suggested that HMGB2 plays an important role in the innate immunity of fish. Cherry Valley duck is the main duck bred for meat consumption in China, but there is limited research available on the impact of duck HMGB2 (duHMGB2) in antiviral innate immunity. Here, duHMGB2 genes were first cloned and analyzed from the spleen of Cherry Valley ducks. We show that duHMGB2 is widely distributed in most tissues of healthy ducks, and duHMGB2 was differentially expressed in three organs (the spleen, brain, and lung) of ducks during different viral infections. duHMGB2 is mainly expressed in the nucleus of duck embryo fibroblast (DEF) cells. However, duHMGB2 is released into the cytoplasm after viral infection. DuHMGB2 induced expression of several genes that regulate the immune response. Moreover, duHMGB2 activated and upregulatede transcription factor NF-κB promoter activity. We also used single gene manipulations (knockout or overexpression) to confirm that duHMGB2 can inhibit the replication of duck plague virus, duck Tembusu virus, and the novel duck reovirus in DEF cells. These data show that duHMGB2 can activate the antiviral innate immunity of the host. Thus, duHMGB2 may be considered an immune adjuvant against infectious diseases in duck.