Disease Virus Infection In Chickens Research Articles

Experimental Velogenic Viscerotropic Newcastle Disease Virus Infection in Chickens Immunologically Impaired by Treatment with Cyclophosphamide

This study investigated whether lymphocytic depletion following chemical bursectomy influenced the severity of infection and development of lesions in chickens challenged with velogenic viscerotropic Newcastle disease virus (vvNDV). Cockerel chickens treated with cyclophosphamide on days 2, 3 and 4 post-hatch showed loss of weight, atrophy and lymphocytic depletion in the bursa of Fabricius and spleen. At 6 weeks of age, the chickens were assigned to four groups- Bursectomized intramuscularly vvNDV inoculated (BI), bursectomized uninfected (BU), non-bursectomized infected (NBI) and non-bursectomized uninfected (NBU) chickens. The BI and NBI chickens showed significant (P < .05) loss of weight than their uninfected controls. Depression, anorexia, greenish diarrhea, listlessness, tremor, and oculo-nasal discharges were observed in both infected groups, but were more severe and frequent in the NBI than in the BI chickens. Total mortalities were 100% and 95.5% for the NBI and BI chickens, respectively (P > .05). Lesions in both infected groups included atrophy of the bursa, spleen and thymus. Hemorrhages in the proventricular mucosa, intestines and cecal tonsils, as well as congestion and enlargement of the kidneys were significantly (P < .05) more severe and frequent in NBI than BI chickens. Histopathology showed necrosis and depletion of lymphocytes in the three lymphoid organs in both infected groups with more severity in the NBI than BI chickens. These results show that depletion of lymphocytes by treatment with cyclophosphamide may influence the severity of infection and development of lesions in vvNDV infection in cockerel chickens.

Temporal Dynamics of Purinergic Receptor Expression in the Lungs of Marek's Disease (MD) Virus-Infected Chickens Resistant or Susceptible to MD.

Marek's disease virus (MDV) is an economic concern for the poultry industry due to its poorly understood pathophysiology. Purinergic receptors (PRs) are potential therapeutic targets for viral infections, including herpesviruses, prompting our investigation into their role in MDV pathogenesis. The current study is part of an experimental series analyzing the expression of PRs during MDV infection. To address the early or short-acting P2 PR responses during natural MDV infection, we performed an "exposure" experiment where age-matched chickens were exposed to experimentally infected shedders to initiate natural infection. In addition, select non-PR regulatory gene responses were measured. Two groups of naïve contact chickens (n = 5/breed/time point) from MD-resistant (White Leghorns: WL) and -susceptible (Pure Columbian) chicken lines were housed separately with experimentally infected PC (×PC) and WL (×WL) chickens for 6 or 24 h. Whole lung lavage cells (WLLC) were collected, RNA was extracted, and RT-qPCR assays were used to measure specific PR responses. In addition, other potentially important markers in pathophysiology were measured. Our study revealed that WL chickens exhibited higher P1 PR expression during natural infection. WL chickens also showed higher expression of P1A3 and P2X3 at 6 and 24 h when exposed to PC-infected chickens. P2X5 and P2Y1 showed higher expression at 6 h, while P2Y5 showed higher expression at 6 and 24 h; regardless of the chicken line, PC chickens exhibited higher expression of P2X2, P2Y8, P2Y10, P2Y13, and P2Y14 when exposed to either group of infected chickens. In addition, MDV infection altered the expression of DDX5 in both WL and PC groups exposed to PC-infected birds only. However, irrespective of the source of exposure, BCL2 and ANGPTL4 showed higher expression in both WL and PC. The expression of STAT1A and STAT5A was influenced by time and breed, with major changes observed in STAT5A. CAT and SOD1 expression significantly increased in both WL and PC birds, regardless of the source of infection. GPX1 and GPX2 expression also increased in both WL and PC, although overall lower expression was observed in PC chickens at 24 h compared to 6 h. Our data suggest systemic changes in the host during early infection, indicated by the altered expression of PRs, DDX5, BCL2, ANGPTL4, and other regulatory genes during early MDV infection. The relative expression of these responses in PC and WL chickens suggests they may play a key role in their response to natural MDV infection in the lungs and long-term pathogenesis and survival.

Effects of Hydromethanol Extract of Stem Bark of Anacardium occidentale on Gross and Histopathological changes Associated with Experimental Velogenic Newcastle Disease Virus Infection in Chickens

Effects of Hydromethanol Extract of Stem Bark of Anacardium occidentale on Gross and Histopathological changes Associated with Experimental Velogenic Newcastle Disease Virus Infection in Chickens

Comparative transcriptome analysis of spleen of Newcastle Disease Virus (NDV) infected chicken and Japanese quail: a potential role of NF-κβ pathway activation in NDV resistance.

The online version contains supplementary material available at 10.1007/s13337-023-00833-y.

Integrated analysis of methylation profiles and transcriptome of Marek's disease virus-infected chicken spleens reveal hypomethylation of CD4 and HMGB1 genes might promote Marek's disease tumorigenesis

Marek's disease (MD) is a lymphoproliferative neoplastic disease caused by Marek's disease virus (MDV). Previous studies have showed that DNA methylation was involved in MD development, but systematic studies are still lacking. Herein, we performed whole genome bisulfite sequencing (WGBS) and RNA-seq in MDV-infected tumorous spleens (IN), noninfected spleens (NoIN), and survivor (SUR) spleens of chickens to identify the genes playing important roles in MD tumor transformation. We generated the first genome-wide DNA methylation profile of MDV-infected, noninfected, and survivor chickens. Combined the WGBS and RNA-Seq, we found that the expression of 25% differential expression genes (DEGs) were significantly correlated with methylation of CpG sites in their gene bodies or promoters. Further, we focused on the DEGs with differentially methylated regions (DMRs) on genes' body and promoter, and it showed the expression of 60% DEGs were significantly correlated with methylation of CpG sites in DMRs. Finally, we identified 8 genes, including CD4, CTLA4, DTL, HMGB1, LGMN, NUP210, RAD52, and ZAP70, and their expression was negatively correlated with methylation of DMRs in their promoters in both IN vs. NoIN and IN vs. SUR. These 8 genes showed specifically high expression in IN groups and clustered in module turquoise analyzed by WGCNA. Out of 8 genes, CD4 and HMGB1 were drop in QTLs associated with MD resistance. Thus, we overexpressed the 2 genes to simulate their high expression in the IN group and found they significantly promoted MDCC-MSB-1 cell proliferation, which revealed they might play promoting roles in MD tumorigenesis in IN due to their high expression induced by hypomethylation.

The brain-specific upregulation of CARD11 in response to avian brain-neurotropic virus infection serves as a potential biomarker

Avian neurotropic viruses are critical problems in poultry industry causing severe central nervous system (CNS) damage with neuroinvasive and neurovirulence properties. Biomarker of neurotropic viral intracranial invasion is of great application value for the diagnosis, but that of avian neurotropic viruses remains elusive. Previously, we found that chicken caspase recruitment domain family, member 11 (CARD11) was only upregulated in virulent Newcastle disease virus-infected chickens and in chicken primary neuronal cells. In this study, CARD11 was systemically expressed in chickens and pigeons detected by absolute qPCR and immunohistochemical (IHC) assay. After virus challenging, only avian neurotropic viruses (avian encephalomyelitis virus [AEV] and pigeon paramyxovirus type 1 [PPMV-1]) except Marek's disease virus (MDV) can invade brain and cause pathological changes. The relative mRNA expression of CARD11 was brain-upregulated in AEV- or PPMV-1-infected animals, rather than MDV and non-neurotropic viruses (fowl adenovirus serotype 4 [FAdV-4] and infectious bronchitis virus [IBV]). Similarly, the protein expression of CARD11 was only upregulated in the cerebra and cerebella infected by avian brain-neurotropic virus using IHC assay. And there were no correlations between the change level of CARD11 and viral load. Our preliminary data suggested that avian CARD11 may be a potential brain biomarker for avian brain-neurotropic virus invasion.

Review of Diagnostic and Vaccination Approaches of Infectious Bursal Disease of Poultry

Infectious bursal disease, also known as Gumboro disease, is a highly contagious and acute viral disease of poultry characterised by the destruction of lymphoid cells. Diagnosis of infectious bursal disease involves consideration of the flocks’ history, clinical signs, and lesions. The objectives of this paper are to highlight various commonly used diagnostic methods for infectious bursal disease and to review advances made in diagnostic methods and vaccination strategies for infectious bursal disease, with special emphasis on the strengths and weaknesses of each of those techniques. Isolation of infectious bursal disease virus followed by its serological assay and histopathological examination of the bursa is regarded as the gold standard method of infectious bursal disease diagnosis. Serological tests such as agar gel, immune diffusion, enzyme-linked immuno sorbent assay, and viral neutralisation tests are commonly used laboratory assays in diagnosing infectious bursal disease viruses. Recently, the most accurate and relatively fast diagnostic method, the molecular technique, is widely used. The molecular diagnostic technique is the simplest and most sensitive of the diagnostic techniques reviewed. The virus causes immunosuppression, so the infected chicken recovers from the acute disease but becomes more susceptible to infections by other pathogens. Therefore, prevention is important and vaccination has become the principal control measure of infectious bursal disease virus infection in chickens. Conventional attenuated live and killed vaccines are the most commonly used vaccines. With the advancement of knowledge and technology, new generation or genetically-engineered vaccines like deoxyribonucleic acid and subunit vaccines have been used. Various vaccination strategies, such as in ovo, at hatch, and post hatch vaccination, are used. Hatchery vaccination is becoming a common practice. Based on this review paper, more affordable and effective infectious bursal disease vaccines that are affordable and readily available must be identified with further cost-benefit analysis.

The Regulatory Microenvironment in Feathers of Chickens Infected with Very Virulent Marek’s Disease Virus

Vaccines against Marek’s disease can protect chickens against clinical disease; however, infected chickens continue to propagate the Marek’s disease virus (MDV) in feather follicles and can shed the virus into the environment. Therefore, the present study investigated if MDV could induce an immunoregulatory microenvironment in feathers of chickens and whether vaccines can overcome the immune evasive mechanisms of MDV. The results showed an abundance of CD4+CD25+ and CD4+ transforming growth factor-beta (TGF-β)+ T regulatory cells in the feathers of MDV-infected chickens at 21 days post-infection. In contrast, vaccinated chickens had a lower number of regulatory T cells. Furthermore, the expression of TGF-β and programmed cell death receptor (PD)-1 increased considerably in the feathers of Marek’s disease virus-infected chickens. The results of the present study raise the possibility of an immunoregulatory environment in the feather pulp of MDV-infected chickens, which may in turn favor replication of infectious MDV in this tissue. Exploring the evasive strategies employed by MDV will facilitate the development of control measures to prevent viral replication and transmission.

Characterization of chicken IFI35 and its antiviral activity against Newcastle disease virus.

Interferon-induced protein-35 kDa (IFI35) was an antiviral protein induced by interferon (IFN)-γ, which plays an important role in the IFN-mediated antiviral signaling pathway. Here, we cloned and identified IFI35 in the chicken for the first time. Chicken IFI35 (chIFI35) contains an open reading frame (ORF) of 1,152 bp encoding a protein of 384 amino acids containing two conserved Nmi/IFI35 domain (NID) motifs. Tissue distribution analysis of chIFI35 in healthy and Newcastle disease (ND) virus-infected chickens indicated a positive correlation between chIFI35 mRNA transcription and ND viral loads in various tissues. The role of chIFI35 in regulation NDV replication were further assessed by up- or down-regulated chIFI35 expression in DF-1 cells transfected with plasmid harboring chIFI35, pCMV-3HA-chIFI35 or shRNA targeting chIFI35, pshRNA-chIFI35 plasmids. NDV replications in DF-1 cells were significantly reduced or slightly increased by over- or under-expression of the chIFI35 protein, respectively, indicating the role of chIFI35 in anti-NDV infection. Moreover, chIFI35 also involved in regulation of viral gene transcription and IFNs expression. The collected data were meaningful for research of chicken antiviral immunity and shed light on the pleiotropic antiviral effect of chIFI35 during NDV infection.

The effect of ghrelin on the fibrosis of chicken bursa of fabricius infected with infectious bursal disease virus

The present study aimed to investigate the effect of ghrelin on the degree of bursa of Fabricius (BF) fibrosis in infectious bursal disease virus-infected chickens. Specific pathogen free (SPF) chicks were divided into four groups. One group was used as the control (“C”). The other three groups were inoculated with IBDV on the 19th day, of which two were injected intraperitoneally with 0.5 nmol (“LG”) or 1.0 nmol (“HG”) ghrelin/100 g weight from the 18th day to the 22nd day, and one was injected intraperitoneally with PBS (“I”). Hematoxylin-eosin staining, Masson's staining, and quantitative real-time PCR were used to determine the effects of ghrelin on the degree of inflammatory cell infiltration, the bursal fibrosis degree, and the expression of TGF-β and MMP-9 mRNA in IBDV-infected SPF chicks. The results showed that ghrelin administration reduced the number of infiltrated inflammatory cells in BF from 5 dpi and significantly attenuated the degree of fibrosis induced by IBDV from 2 dpi to 7 dpi (P < 0.05). Moreover, the TGF-β expression in the LG and HG groups were significantly or highly significantly lower (P < 0.05 or P < 0.01) than those of I group from 2 dpi to 5 dpi. In addition, ghrelin administration downregulated MMP-9 expression evoked by IBDV from 2 dpi to 7 dpi (P < 0.05 or P < 0.01). These results suggested that ghrelin attenuated the bursal fibrosis degree of IBDV-infected SPF chicks by reducing the number of inflammatory cells and by decreasing the expression of TGF-β and MMP-9, which shortened the process of bursa recovery.

Gut microbiota is associated with protection against Marek's disease virus infection in chickens

Marek's Disease Virus (MDV) infects chickens via respiratory route and causes lymphomas in internal organs including gastrointestinal tract. MDV infection causes a shift in the gut microbiota composition. However, interactions between the gut microbiota and immune responses against MDV infection are not well understood. Therefore, the current study was performed to understand the effect of the gut microbiota on Marek's disease (MD) pathogenesis. The findings showed that depletion of gut microbiota increased the severity of MD in infected chickens. In addition, an increase in the transcription of interferon (IFN)-α, IFN-β and IFN-γ in the bursa of Fabricius at 4 days post-infection (dpi) was observed in the gut microbiota depleted chickens. The observations in this study shed more light on the association between the gut microbiota and MDV infection in chickens. More research is needed to explore the mechanisms of involvement of the gut microbiota in immunity against MD in chickens.

RNA-seq analysis of viral gene expression in the skin of Marek’s disease virus infected chickens

Marek’s disease virus (MDV), a highly cell-associated oncogenic avian α-herpesvirus, is the causative agent of malignant transformation of T cells in domestic chickens. The latently infected CD4+CD8− T cells carry the virus through the blood stream and establish lymphomas in the skin, visceral organs and peripheral nerves. The feather follicle epithelium (FFE) is the only anatomical site where fully infectious enveloped virions are produced and eventually disseminated into the environment to infect contact birds. Therefore, skin and FFE play a critical role as being the common source of re-infection of birds sharing the same habitat. The molecular mechanism involved in the replication and assembly of MDV in the FFE leading to the production and release of cell-free infectious virus particles is unknown and to date no viral or host gene has been implicated in the process. To examine alterations in the expression pattern of viral genes, we performed RNA-seq on the skin samples of Marek’s disease virus-infected susceptible chickens at 10, 20, and 30 days post infection. For comparative analysis of the expression patterns of viral genes between the skin and spleen of the MD-susceptible and resistant lines, Real-Time RT-PCR was employed. In total, RNA-seq based analysis identified 42 viral genes that were differentially expressed in the skin of infected birds. Majority of the identified genes are involved in DNA replication, capsid, tegument, and envelop formation. Comparative analysis between the skin and spleen of MD-susceptible and resistant chicken lines, revealed significantly higher expression of the genes in the skin of either lines than the spleen. Furthermore, much higher expression of the genes was observed in the skin of the susceptible line than the resistant line.

Differential expression of type I interferon mRNA and protein levels induced by virulent Marek’s disease virus infection in chickens

Marek's disease virus (MDV), an α-herpesvirus targeting avian species, causes fatal Marek's disease (MD) in chickens. The host interferon (IFN) responses play a key role in resisting viral infection. However, host IFN responses following MDV infection in the chicken central immune organs (thymus and bursa of Fabricius), which contain numerous MDV target cells, is poorly understood. In this study, we performed animal experiments in specific pathogen-free chickens infected with two virulent MDV strains (BS/15 and Md5) or without infection as negative controls. Specifically, the type I IFN (IFN-α and IFN-β) transcriptional and proteomic expression levels at 7, 10, 14, 17, and 21 days post infection (dpi) were detected and analyzed. Our results indicated that the mRNA and protein expression levels of IFN-α and IFN-β in the thymus and bursa of Fabricius were mainly downregulated in cytolytic infection (such as 10 dpi) and reactivation (such as 17 dpi) stages, but not the latent (such as 14 dpi) stage of MDV infection, which was determined by comprehensively analyzing the MDV viral load and immune organ damage caused by MDV infection. These data suggest that MDV could inhibit the expression of host type I IFNs, which may be involved in the MDV-induced host immunosuppression and contribute to the immune escape of MDV from host immunity. Furthermore, we found that the downregulated expression of the host type I IFNs induced by BS/15 and Md5 infection was significantly different, which we speculated may be related to the diverse virulence and pathogenicity of MDV strains. In conclusion, our study demonstrated that MDV mostly inhibited the expression of type I IFNs in infected hosts, which may be associated to its pathogenesis.

Common microRNA–mRNA Interactions in Different Newcastle Disease Virus-Infected Chicken Embryonic Visceral Tissues

To investigate the roles and explore the altered expression of microRNAs (miRNAs) and mRNAs in chicken embryos in response to Newcastle disease virus (NDV) infection, deep sequencing was performed. Then, a conjoint analysis of small RNA-seq and mRNA-seq was performed to screen interactional miRNA–mRNA pairs during NDV infection. In total, 15 and 17 up- and downregulated miRNAs were identified that potentially targeted 4279 and 6080 mRNAs in NDV-infected chicken embryonic tissues, respectively; in addition, 595 upregulated and 480 downregulated mRNAs were identified. The conjoint analysis of the obtained data identified 1069 miRNA–mRNA pairs. Among these pairs, 130 pairs were related to immune or inflammatory responses. The relationship between gga-miR-203a and its target transglutaminase 2 (TGM2) was confirmed using a dual-luciferase reporter system and a real time quantitative polymerase chain reaction (RT-qPCR) assay. Overall, the discovery of miRNAs, mRNAs, and their potential pairing relationships, which may be involved in the regulation of NDV infection, will facilitate our understanding of the complex regulatory relationship between the host and the virus.

Atrophy of the lymphoid organs and suppression of antibody response caused by velogenic Newcastle disease virus infection in chickens.

This project was undertaken to study the immunosuppressive capabilities of velogenic viscerotropic pathotype of Newcastle disease virus (VVNDV) infection in cockerels. Two hundred six-week-old cockerels were divided into four groups. Groups B/VUC and C/VC were vaccinated with LaSota in drinking water at 6weeks of age. Groups C/VC and D/UC were challenged with VVNDV at 8weeks of age. Three days post challenge (PC), the cockerels in group D/UC came down with clinical signs which included depression and greenish diarrhoea. Total mortality was 74.6%. The C/VC cockerels showed no clinical signs. But both challenged groups showed significant weight loss, significant loss of total serum proteins, globulin and albumen (P < 0.05). These losses were more severe in the D/UC than in the C/VC. There was severe atrophy of the bursa, spleen and thymus in both groups. Histopathology showed severe necrosis and depletion of the lymphocytes in the three lymphoid organs. However, the lesions were more severe in the D/UC than in C/VC cockerels. On day 28, PC groups B/VUC, C/VIC and D/UIC were revaccinated with LaSota. The haemagglutination inhibition antibody response on days 35, 42 and 49 PC was very low in groups C/VIC and D/UIC when compared with B/VUC cockerels. These observations show that VVNDV infection both clinical and subclinical can cause immunosuppression and vaccine failure due to severe destruction of the lymphocytes in the lymphoid organs. This will be a serious problem for poultry production in those countries where the disease is enzootic.

Immune Responses in Cecal Tonsils of Marek's Disease Virus-Infected Chickens.

Marek's disease (MD) is a lymphoproliferative disease of domestic chickens that is caused by a highly cell-associated oncogenic α-herpesvirus, Marek's disease virus (MDV). MDV replicates in chicken lymphocytes and establishes a latent infection within CD4+ T cells. Clinical signs of MD include depression, crippling, weight loss, bursal/thymic atrophy, neurologic disorders, and rapid onset of T cell lymphomas that infiltrate lymphoid tissues, visceral organs, and peripheral nerves. The cecal tonsils (CTs) are considered the largest lymphoid aggregates of avian gut-associated lymphoid tissue. Along with Peyer's patches, CTs elicit protective immune responses against bacterial and viral pathogens in the intestinal tract of avian species. In this study, we investigated the effect of MDV infection on toll-like receptor (TLR) gene expression in CTs of MD-susceptible (72) and resistant (63) chicken lines. Real-time PCR gene expression profiling revealed that of the 10 TLRs tested, TLR2A, TLR3, TLR5, and TLR15 displayed significant differential expression patterns at different time points postinoculation. The expression levels of the remaining six genes were minimally affected by MDV infection in either line. Immunohistochemical analysis showed a severe depletion of B cells and CD4+ T cells in the CTs of susceptible line at 5 days postinfection (dpi), which recovered by 21 dpi. The destruction of B and T cells in the CTs of the resistant line was minimal at 5 dpi, which also recovered by 21 dpi. A significant infiltration of macrophages was observed after the depletion of B and T cells in the infected birds of both lines that could account for the differential TLR gene expression in the infected birds. The data presented provide further insight into the mechanism of MDV pathogenesis and tissue-specific immunologic responses to viral infection.

Temporal expression and DNA hypomethylation profile of CD30 in Marek's disease virus-infected chicken spleens

Marek's disease (MD) is a viral neoplastic disease of chickens caused by Marek's disease virus (MDV), which is serious threat to worldwide poultry industry. Our previous studies showed that the CD30 gene was hypomethylated in MD lymphoma. In this study, we further analyzed differential expression patterns and methylation levels of the CD30 gene between MDV-infected and noninfected spleens at 4, 7, 14, 21, and 28 d postinfection (dpi). The results showed that the expression of CD30 in MDV-infected spleens was significantly lower than that in noninfected spleens at 4 dpi. The expression of CD30 did not present significant difference between MDV-infected and noninfected spleens at 7 and 14 dpi. However, an increased expression of CD30 was presented in MDV-infected spleens at both 21 and 28 dpi. Simultaneously, CD30 showed a lower DNA methylation level in MDV-infected spleens at 14, 21, and 28 dpi. The results indicated that CD30 gene was involved in the whole process of MD tumorigenesis and upregulated expression of CD30 in MDV-infected spleens might be attributed to the hypomethylation of promoter of CD30 gene.

Comparison of the Serum Proteins and Immune Responses of Velogenic Newcastle Disease Virus Infected Chickens and Ducks

The effect of velogenic Newcastle disease virus (vNDV) on the immune responses and serum proteins was investigated in six-week-old ducks and chickens. Results showed that weight loss was markedly significant (p

Immunization with a Thermostable Newcastle Disease Virus K148/08 Strain Originated from Wild Mallard Duck Confers Protection against Lethal Viscerotropic Velogenic Newcastle Disease Virus Infection in Chickens

Newcastle disease (ND) is one of the most devastating poultry infections because of its worldwide distribution and accompanying economical threat. In the present study, we characterized the ND virus (NDV) K148/08 strain from wild mallard duck, with regard to safety, thermostability, immunogenicity, and protective efficacy against velogenic ND viral infection. The NDV K148/08 strain offered enhanced immunogenicity and safety relative to commercially available vaccine strains. The NDV K148/08 strain was safe in 1-day-old SPF chicks after vaccination using a coarse or cabinet-type fine sprayer. We demonstrated that the NDV K148/08 strain elicited high levels of antibody responses and provided protective efficacy against lethal NDV challenge. In addition, the thermostability of the NDV K148/08 strain was as high as that of the thermostable V4 strain. Therefore, the NDV K148/08 strain may be useful to ensure NDV vaccine performance and effectiveness in developing countries, especially in remote areas without cold chains.

Overexpression of microRNA gga-miR-21 in chicken fibroblasts suppresses replication of infectious bursal disease virus through inhibiting VP1 translation

Previously we have identified a series of cellular miRNA molecules up- or down-regulated in infectious bursal disease virus (IBDV) infected chicken embryo fibroblasts and Bursa of Fabricius with gene microarray analysis. Here we studied in detail a relatively well studied miRNA, gga-miR-21, for better understanding miRNAs involvement in IBDV-host interactions. Chicken pri-gga-miRNA-21 and a control miRNA Caenorhabditis elegans pri-cel-lin-4 gene were cloned into a lentiviral vector, respectively. The resulting recombinant lentiviruses were used to infect chicken fibroblast cell line DF-1, and two stable cell lines, DF-miR-21 (overexpressing gga-miR-21) and DF-lin-4 (overexpressing cel-lin-4), were selected. Replication of IBDV in DF-miR-21, DF-lin-4 and DF-1 cells were compared and molecular mechanism of IBDV replication alteration was explored using bioinformatics, reporter gene system, qRT-PCR and Western blot analysis. IBDV replication was markedly lower in DF-miR-21 than in DF-lin-4 or DF-1 cells. A gga-miR-21 target sequence was identified within IBDV VP1 gene (1713–1734bp). Fusion of a 520nt long partial IBDV VP1 gene containing the target with a luciferase gene resulted in significantly lower transient luciferase activity in DF-miR-21 cells as compared to that in DF-lin-4 or DF-1 cell. Following IBDV infection of the cell lines, VP1 protein level in DF-miR-21 cells was dramatically lower than that in DF-lin-4 or DF-1 cells but VP1 mRNA level was not different. The finding indicated that gga-miR-21 could suppress IBDV replication through down regulating IBDV VP1 expression at translational level.