Reticuloendotheliosis Virus Infection Research Articles

Effects of Reticuloendotheliosis virus on TLR-3/IFN-Β pathway in specific pathogen-free chickens

Birds infected by Reticuloendotheliosis virus (REV) are vulnerable to other microorganisms. This immunosuppression is related to the immune organs (thymus, bursa of Fabricius, and spleen) damaged by REV. The regulation of IFN-β greatly depends on pattern recognition receptor TLR-3 and nuclear factors IRF-7, NF-κB. To address if and how the TLR-3/IFN-β pathway is disturbed by REV, 60 one-day-old specific-pathogen-free chickens were intraperitoneally injected with RE virus dilution (n = 30) or stroke-physiological saline solution (n = 30). At 1, 3, 7, 21, and 28 days post-infection, after collecting thymuses, bursas, and spleens, we monitor the kinetics of TLR-3, IFN-β, NF-κB p65, and IRF-7 at transcriptional and translational levels using qPCR, Western blotting, and ELISA separately. As a result, compared with control chickens, the mRNA levels of TLR-3, IRF-7, and NF-κB p65 showed increasingly differences in the early period of REV infection. Synchronal changes occurred at translation levels. In the latter infection period, a decrease of NF-κB p65 was contemporaneous with a fall in IFN-β at both transcriptional and translational levels in the thymuses and bursas. These data suggest that the changes of IFN-β content are closely related to NF-κB p65 when REV invades chicken central immune organs. That reveals new insights into the immunosuppression mechanism of REV in avian.

PATHOGEN SURVEY AND PREDICTORS OF LYMPHOPROLIFERATIVE DISEASE VIRUS INFECTION IN WILD TURKEYS (MELEAGRIS GALLOPAVO).

Growing populations of Wild Turkeys (Meleagris gallopavo) may result in increased disease transmission among wildlife and spillover to poultry. Lymphoproliferative disease virus (LPDV) is an avian retrovirus that is widespread in Wild Turkeys of eastern North America, and infections may influence mortality and parasite co-infections. We aimed to identify individual and spatial risk factors of LPDV in Maine's Wild Turkeys. We also surveyed for co-infections between LPDV and reticuloendotheliosis virus (REV), Mycoplasma gallisepticum, and Salmonella pullorum to estimate trends in prevalence and examine covariance with LPDV. From 2017 to 2020, we sampled tissues from hunter-harvested (n=72) and live-captured (n=627) Wild Turkeys, in spring and winter, respectively, for molecular detection of LPDV and REV. In a subset of captured individuals (n=235), we estimated seroprevalence of the bacteria M. gallisepticum and S. pullorum using a plate agglutination test. Infection rates for LPDV and REV were 59% and 16% respectively, with a co-infection rate of 10%. Seroprevalence for M. gallisepticum and S. pullorum were 74% and 3.4%, with LPDV co-infection rates of 51% and 2.6%, respectively. Infection with LPDV and seroprevalence of M. gallisepticum and S. pullorum decreased, whereas REV infection increased, between 2018 and 2020. Females (64%), adults (72%), and individuals sampled in spring (76%) had higher risks of LPDV infection than males (47%), juveniles (39%), and individuals sampled in winter (57%). Furthermore, LPDV infection increased with percent forested cover (β=0.014±0.007) and decreased with percent agriculture cover for juveniles (β=-0.061±0.018) sampled in winter. These data enhance our understanding of individual and spatial predictors of LPDV infection in Wild Turkeys and aid in assessing the associated risk to Wild Turkey populations and poultry operations.

TMT-based proteomic analysis reveals integrins involved in the synergistic infection of reticuloendotheliosis virus and avian leukosis virus subgroup J

BackgroundCo-infection with the avian leukosis virus subgroup J (ALV-J) and the reticuloendotheliosis virus (REV) increases mutual viral replication, causing a more serious pathogenic effect by accelerating the progression of neoplasia and extending the tumor spectrum. However, the molecular mechanism underlying the synergistic replication of ALV-J and REV remains unclear.ResultsHere, we performed this study to compare the differentially expressed proteins among CEF cells infected with ALV-J, REV or both at the optimal synergistic infection time using TMT-based quantitative proteomics. We identified a total of 719 (292 upregulated and 427 downregulated) and 64 (35 upregulated and 29 downregulated) proteins by comparing co-infecting both viruses with monoinfecting ALV-J and REV, respectively. GO annotation and KEGG pathway analysis showed the differentially expressed proteins participated in virus-vector interaction, biological adhesion and immune response pathways in the synergistic actions of ALV-J and REV at the protein levels. Among the differentially expressed proteins, a large number of integrins were inhibited or increased in the co-infection group. Further, eight integrins, including ITGα1, ITGα3, ITGα5, ITGα6, ITGα8, ITGα9, ITGα11 and ITGβ3, were validated in CEF cells by qRT-PCR or western blot.ConclusionsThese findings proved that integrins may be key regulators in the mechanism of synergistic infection of REV and ALV-J, which will provide more insight into the pathogenesis of synergism of REV and ALV-J at protein level.

The expression profile of miR-222b-5p/MAPK10 in spleens of SPF chickens infected with REV-SNV at 28-42 dpi.

Reticuloendotheliosis virus (REV) is an avian oncogenic retrovirus that causes atrophy of immune organs, such as the spleen, thymus, and bursa of Fabricius, leading to severe immunosuppression. However, there is limited information describing the genes or microRNAs (miRNAs) that play a role in replicating REV-spleen necrosis virus (SNV). Our previous miRNA and RNA sequencing data showed that the expression of gga-miR-222b-5p was significantly upregulated in REV-SNV-infected chicken spleens of 7, 14, and 21 dpi compared to non-infected chicken spleens, but mitogen-activated protein kinase 10 (MAPK10), which is related to innate immunity, had the opposite expression pattern. To understand chicken cellular miRNA function in the virus-host interactions during REV infection, we used quantitative reverse transcription PCR (qRT-PCR) to determine whether the expression of gga-miR-222b-5p and MAPK10 in the spleen of specific-pathogen-free chickens at 28, 35, and 42 dpi was consistent with the first 3 time points, and dual-luciferase reporter assay was used to determine the targeting relationship between gga-miR-222b-5p and MAPK10. Results show that MAPK10 was downregulated at all 3 time points; however, significant difference (p⟨0.01) was noted only at 35 dpi. Moreover, the expression of gga-miR-222b-5p was upregulated; however, significant difference (p⟨0.01) was observed only at 28 and 35 dpi. A dual-luciferase reporter assay showed that MAPK10 is a direct target of gga-miR-222b-5p. This study suggests that gga-miR-222b-5p may target MAPK10 to promote the REV-SNV-induced tumorigenesis via the RLRs signaling pathway.

Tandem Mass Tag-Based Quantitative Proteomic Analysis of Chicken Bursa of Fabricius Infected With Reticuloendotheliosis Virus.

Reticuloendotheliosis virus (REV) is a type C avian retrovirus that causes immunosuppression, dwarf syndrome, and lymphoma in infected hosts. In this study, we used tandem mass tag (TMT) labeling and liquid chromatography–tandem mass spectrometry (LC-MS/MS) to characterize protein alterations in chicken bursa of Fabricius, before and after REV infection at 7, 14, 21, and 28 days. Our data showed that 1,127, 999, 910, and 1,138 differentially expressed proteins were significantly altered at 7, 14, 21, and 28 days after REV infection, respectively. Morphological analysis showed that REV infection reduced in cortical lymphocytes, bursal follicle atrophy, and nuclear damage. Bioinformatics analysis indicated these proteins were mainly involved with immune responses, energy metabolism, cellular processes, biological regulation, metabolic processes, response to stimuli, and multicellular organismal process. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway cluster analysis showed that post-infection, proteins were enriched in the cell cycle, Wnt signaling, antigen processing and presentation, cytokine receptor interaction, adenosine 3′,5′-cyclic monophosphate signaling pathway, and NF-κB signaling. In addition, we observed that peroxiredoxin 4 (PRDX4), peroxiredoxin 6 (PRDX6), glutathione peroxidase 3 (GPX3), catalase (CAT), and peroxidasin (PXDN) were involved in oxidative stress. Some heat shock protein (HSP) family members such as HSPH1, DNAJA4, HSPA8, and HSPA4L also changed significantly after REV infection. These findings help clarify interactions between REV and the host and provides mechanistic insights on REV-induced host immunosuppression.

Expression analysis of genes related to lipid metabolism in peripheral blood lymphocytes of chickens challenged with reticuloendotheliosis virus

The underlying molecular mechanism of lipid metabolism in peripheral blood lymphocytes from chicken infected with reticuloendotheliosis virus (REV) remains poorly understood. Therefore, this scientific question was explored in vitro and in vivo. The results indicated that triglyceride content was significantly reduced, but the free fatty acid content and carnitine palmitoyltransferase-1 activity were significantly increased in blood lymphocytes after REV infection. By RNA sequencing, 97 known differentially expressed genes (DEG) related to lipid metabolism or glycometabolism were screened via Gene Ontology term analysis. On the basis of these 97 DEG, enriched pathways, including the peroxisome proliferators-activated receptor (PPAR) signaling pathway, were identified. Among these 97 DEG, some representative genes were related to lipolysis and fatty acid utilization (PPARG, LPL, PLIN2, ACOX1, ACSL1, FABP3, and FABP4). However, other genes related to lipid biosynthesis (ACSL3, ACSL6, DGAT2, LPIN1, and LPIN2) were downregulated. The quantitative polymerase chain reaction results confirmed the accuracy of the RNA sequencing data, and the in vivo outcome supports theses in vitro results. Our findings revealed that REV regulates fatty acid and lipid metabolism in peripheral blood lymphocytes from chicken. After the lymphocytes were infected with REV, the exogenous fatty acids were preferentially used; genes involved in fatty acid utilization were upregulated via the PPAR pathway, whereas genes involved in lipid and fatty acid biosynthesis were downregulated.

An optimized secretory expression system and immunogenicity evaluation for glycosylated gp90 of avian reticuloendotheliosis virus

Reticuloendotheliosis is an important immunosuppressive disease, associated with avian reticuloendotheliosis virus (REV) infection, and causes notable economic losses worldwide. Glycoprotein gp90 is an important structural protein of REV, and considered to be the most important immunogenic antigen, which can induce neutralizing antibodies against REV. In this study, an optimized suspension culture system was developed and applied to secretory express the immunogenic surface antigen gp90. To achieve an optimal glycosylation, the gp90 was designed to secretory expressed into the supernatant of the cell culture, which also occurs in the natural protein maturation procedure of REV. Serum-free culture medium was introduced to simplify the purification process and reduce the production costs. Based on the purified glycosylated gp90, an oil-emulsion subunit REV vaccine candidate was developed and evaluated in chickens. The subunit gp90-based vaccine induced fast immune responses, high levels of antibodies (REV-specific antibody, gp90-specific antibody, and neutralizing antibody against REV), and preferential T helper 2 (Th2) (interleukin-4 secretion) not Th1 (interferon-γ secretion) response. Furthermore, the viremia induced by REV infection was significantly reduced in chickens immunized with the glycosylated gp90. Overall, an optimized secretory expression system for glycosylated gp90 was developed, and the glycosylated gp90 obtained in this study retained good immunogenicity and could be an attractive vaccine candidate to protect chickens against REV horizonal infection.

An outbreak in three-yellow chickens with clinical tumors of high mortality caused by the coinfection of reticuloendotheliosis virus and Marek’s disease virus: a speculated reticuloendotheliosis virus contamination plays an important role in the case

Both reticuloendotheliosis and Marek's disease are neoplastic diseases of chickens caused by reticuloendotheliosis virus (REV) and Marek's disease virus (MDV), respectively. The infection of REV or MDV may lead to clinical tumors and also result in immunosuppression and easily allow secondary infection by other pathogens. Here, we investigated a breeder flock of three-yellow chickens in southern China that had been vaccinated with CVI988/Rispens at hatching and had experienced depression, weakness, reduction in weight gain, and an increased death rate after 120 d of age. The morbidity and mortality were 20% and 10%, respectively, at 140 d of age when this infection was diagnosed. The necropsy of the birds revealed significant tumor-like lesions in the heart, liver, spleen, and ceca. Peripheral blood lymphocytes and tumor-like tissues were sampled for PCR detection and for histopathological observation, for virus isolation and the subsequent immunofluorescent assay on the cell cultures and for gene sequencing of the isolated viruses. A REV isolate GX18NNR1 and a MDV isolate GX18NNM5 were both recovered from the sampled bird. Further phylogenetic analysis based on the env gene of REV and the meq gene of MDV demonstrated that GX18NNR1 was closely related to the reference REV strain MD-2, which was isolated from a contaminated commercial turkey herpesvirus vaccine. In addition, the GX18NNM5 was found to belong to the Chinese very virulent MDV strains' cluster. The coinfection of REV and MDV may contribute to tumor outbreaks with high morbidity and mortality in three-yellow chicken flocks.

TRIM62 From Chicken as a Negative Regulator of Reticuloendotheliosis Virus Replication.

Emerging evidence suggests that the tripartite motif containing 62 (TRIM62), a member of the TRIM family, plays an important role in antiviral processes. The objective of the study was to explore the role of TRIM62 in reticuloendotheliosis virus (REV) infection and its potential molecular mechanism. We first demonstrated that the REV infection affected the TRIM62 expression first upregulated and then downregulated in CEF cells. Next, we evaluated the effect of TRIM62 on viral replication. Overexpression of TRIM62 decreased REV replication. On the contrary, silencing of endogenously expressed TRIM62 increased viral replication. Then, to explore the necessity of domains in TRIM62's negative regulation on viral replication, we transfected CEF cells with TRIM62 domain deletion mutants. Deletion domain partially abolished TRIM62's antiviral activity. The effect of SPRY domain deletion was the highest and that of coiled-coil was the lowest. Further, we identified 18 proteins that coimmunoprecipitated and interacted with TRIM62 by immunocoprecipitation and mass spectrometry analysis. Strikingly, among which, both Ras-related protein Rab-5b (RAB5B) and Arp2/3 complex 34-kDa subunit (ARPC2) were involved in actin cytoskeletal pathway. Altogether, these results strongly suggest that chicken TRIM62 provides host defense against viral infection, and all domains are required for its action. RAB5B and ARPC2 may play important roles in its negative regulation processes.

Regulatory mechanism of microRNA-155 in chicken embryo fibroblasts in response to reticuloendotheliosis virus infection

Reticuloendotheliosis virus (REV) infection of multiple avian species can lead to a number of diseases such as runting syndrome, immunosuppression and oncogenesis, causing major economic losses. MicroRNAs play important roles in post-transcriptional regulation, effectively inhibiting protein synthesis, and participating in many biological processes in cells, including proliferation, differentiation, apoptosis, lipometabolism, virus infection and replication, and tumorigenesis. Based on our previous high-throughput sequencing results, we explore the regulatory mechanisms of microRNA-155(miR-155) in chicken embryo fibroblasts (CEFs) in response to REV infection. Our results revealed expression of miR-155 in CEFs after REV infection upregulated in a time- and dose-dependent manner, indicating miR-155 plays a role in REV infection in CEFs indeed. After transfected with miR-155-mimic and miR-155-inhibitor, we found overexpression of miR-155 targeted caspase-6 and FOXO3a to inhibit apoptosis and accelerate cell cycle, thus improving viability of REV-infected CEFs. This result also verified the protective role of miR-155 in the viability of CEFs in the presence of REV. Knockdown of miR-155 also supported these above conclusions. Our findings uncover a new mechanism of REV pathogenesis in CEFs, and also provide a theoretical basis for uncovering new effective treatment and prevention methods for RE based on miR-155.

Two new IncRNAs regulate the key immune factor NOD1 and TRAF5 in chicken lymphocyte

Reticuloendotheliosis virus (REV) causes the atrophy of immune organs and immuno-suppression in chickens, but the underlying molecular mechanism of the immune response after infection by REV is not well understood. Presently, the RNA-seq was used to analyze the regulation of immune response to REV in chicken lymphocytes from peripheral blood. Overall, 134 differentially expressed long non-coding RNAs (lncRNAs) between cells with REV infection or without in vitro were screened. Based on the differentially expressed protein-coding genes, the nucleotide-binding oligomerization domain (NOD)-like receptor pathway related to immune regulation was enriched. Two lncRNAs (L11530 and L09863) were predicted to target the NOD1 and tumor necrosis factor receptor-associated factor 5 (TRAF5) gene, respectively, which are involved in the NOD-like receptor pathway with cis-regulation way. The in vitro results revealed the significantly up-regulated (P<0.01) levels of lncRNA-L11530 and its target gene, NOD1, and the significantly down-regulated (P<0.05) levels of lncRNA-L09863 and its target gene, TRAF5, in lymphocytes after REV infection. These changes also occurred in vivo in blood lymphocytes of chickens infected with REV. Further, L09863 and L11530 were respectively interfered, the expression levels of their target genes NOD1 or TRAF5 were significantly down-regulated, accompanied by the change of IL-8 and IL-18 secretions in lymphocytes. The NOD-like receptor pathway appears to be important in the immune response to REV, LncRNA-11530 and IncRNA-09863 might involve in the immune regulation on REV infection by targeting NOD1 or TRAF5 in blood lymphocytes of chickens. Our findings reveal a new regulation of lncRNAs (L11530 and L09863) on immunity in chicken peripheral blood lymphocytes for REV infection by changing the expression of the target genes via the NOD-like receptor pathway.

Proteome analysis of reticuloendotheliosis-virus-infected chicken embryo fibroblast cells through iTRAQ-based quantitative proteomics.

Reticuloendotheliosis virus (REV) is an important representative avian retrovirus. To improve our understanding of the host cellular responses to virus infection and the pathogenesis of REV infection, we applied isobaric tags for relative and absolute quantification (iTRAQ) labeling coupled with multidimensional liquid chromatography-tandem mass spectrometry to detect changes in protein levels in chicken embryo fibroblast cells (CEFs) that were infected with REV or mock infected. In total, 605 cellular proteins were differentially expressed, among which 196, 345, and 286 were differentially expressed in REV-infected CEFs at 1, 3, and 5days postinfection, respectively. Gene Ontology analysis indicated that the biological processes of the differentially expressed proteins were primarily related to cellular processes, metabolic processes, biological regulation, response to stimulus, and immune system processes and that the molecular functions in which the differentially expressed proteins were mainly involved were binding, catalytic activity, and enzyme regulator activity. Pathway analysis showed that a total of 143, 167, and 179 pathways, including protein digestion and absorption, focal adhesion, ECM-receptor interaction, cytokine-cytokine receptor interaction, Toll-like receptors, and JAK-STAT signaling, were enriched in REV-infected CEFs at 1, 3, and 5days postinfection, respectively. In conclusion, this study is the first to analyze the protein profile of REV-infected CEFs using an iTRAQ approach. The results of this study provide valuable information for better understanding the host response to REV infection.

Transcriptional analysis of host responses related to immunity in chicken spleen tissues infected with reticuloendotheliosis virus strain SNV

In avian species, the Reticuloendotheliosis virus (REV) causes severe immunosuppression and other symptoms, including avian dwarfing syndrome, and chronic tumors in lymphoid and other tissues. The pathogenesis of REV and its interaction with the host have yet to be fully elucidated with transcriptional studies on the changes in host gene expression after REV infection at the body level. In this study, the Spleen Necrosis Virus (SNV) was used to inoculate the one-day-old specific pathogen free (SPF) chicken to simulate congenital infection. We identified 1507 differentially expressed genes (DEGs) at 7, 14 and 21 dpi using Next Generation Sequencing (NGS) technology. Through the Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of these DEGs, it was found that DEGs were mainly involved in the categories of signal transduction, immune system and signaling molecules and interaction. Among them, Pattern recognition receptors (PRRs), chemokine, T cell receptor, JAK-STAT, TNF, and NF-kappa B signaling pathway, and the Hematopoietic cell lineage play an important role in the tumorigenic and immunosuppressive regulation of REV. In addition, a series of DEGs associated with inflammatory factors (CCL4, TNFRSF18, CDKN2), apoptosis (IRF1, PDCD1, WNT5A), innate immunity (TLR, MAD5, TRIM25), and adaptive immunity (LY6E, CD36, LAG3) were also discovered. We further verified 33 selected immune- relevant DEGs using quantitative RT-PCR (qRT-PCR). These findings provide new insights and research directions for revealing the pathogenesis of REV infection and the interaction between REV and the chicken immune system.

Genetic characterization of reticuloendotheliosis virus in chickens in Thailand

Reticuloendotheliosis virus (REV) causes an immunosuppressive, runting, and oncogenic disease in poultry, posing a significant threat to the poultry industry. In Thailand, an unidentified disease associated with runting-stunting syndrome and neoplasia, resembling REV infection, has been continuously observed in several chicken farms. However, REV infection in Thailand has never been reported. In this study, we investigated the occurrence and genetic characteristics of REVs in chickens in Thailand from 2013 to 2016. Of the 130 clinical samples obtained from 29 chicken farms from 9 provinces located in the major chicken-raising regions of Thailand, including the central, eastern, northern, and northeastern parts of Thailand, 51 samples (39.23%) and 21 farms (72.41%) were REV-positive. REV-positive samples were detected in all 9 provinces tested. Our results demonstrated that REV was extensively distributed in the major chicken-raising regions of Thailand. Phylogenetic analysis of the whole genome sequence showed that Thai REV was most closely related to Chinese, Taiwanese, and the US REV strains isolated from different avian species and clustered into REV subtype III. This finding indicates that REV subtype III was predominantly circulated in Thai chicken flocks. This study is the first report on REV infection in chickens in Thailand. Our findings raise the awareness of REV as another causative agent of runting and oncogenic disease in chickens in Thailand and highlight the wide distribution of REV infection among chickens worldwide.

Changes in apoptosis, proliferation and T lymphocyte subtype on thymic cells of SPF chickens infected with reticuloendotheliosis virus

Reticuloendotheliosis virus (REV), an avian retrovirus is able to infect a variety of birds and can cause immunosuppression. The aim of this study was to investigate the relationship of thymic lymphocytes apoptosis, proliferation and T cell subtype with immunosuppression. In this study, a hundred and twenty one-day old SPF chickens were randomly divided into control groups (group C) and a REV infection groups (group I). The chickens of group I received intraperitoneal injections of REV with 104.62/0.1 ml TCID50. On day 14, 21, 28 and 35 post-inoculation, the chickens of C group and I group were sacrificed by cardiac puncture blood collection, and the thymic lymphocytes was sterile collected. The proliferation ability of lymphocytes was tested by Cell Counting Kit-8. Flow cytometry was performed to detect apoptosis, cell cycle stage and the change in T cell subtype. The RNA genome copy numbers of REV virus were detected using real-time PCR. Real-time PCR and western blotting were performed to analyze the expression of CyclinD1 and Bcl-2. Our results showed that REV genome copy number steadily declined, the proliferation potential of thymic lymphocytes was inhibited, lymphocytes apoptosed, the ratio of CD4+/CD8+ decreased and the expression of CyclinD1 and Bcl-2 were firstly inhibited, then rapidly recovered. Thus, immunosuppression lead by REV is closely related to the change of T cell subtype, apoptosis, and proliferation of thymic lymphocytes.

Assessment on reticuloendotheliosis virus infection in specific-pathogen-free chickens based on detection of yolk antibody.

Reticuloendotheliosis virus (REV) is the most frequent exogenous virus that contaminates attenuated vaccines. Therefore, it is extremely important to select REV-free specific-pathogen-free (SPF) chicken embryos. Generally, REV infection is assessed by detecting REV antibodies in SPF chickens. This present study seeks to evaluate REV infection by replacing serum antibody detection with yolk antibody detection. A cohort of 40 nineteen-week-old SPF chickens were artificially inoculated with REV, with 32 SPF chickens raised in another isolation environment served as a blank control. Eggs and serum from 23-week-old chickens were sampled, and yolks were diluted separately to ratios of 1:150, 1:200, 1:300 and 1:400, which were detected together with serum. We found that the yolk antibody detection findings at a dilution of 1:300 had the highest coincidence rate compared with that based on serum antibody measurements. At a dilution ratio of 1:300 for yolk antibody, 72 chickens were continuously observed for 10 weeks from 25- to 34-weeks-old. Our findings were based on serum antibody or yolk antibody detection, and the evaluation results were completely consistent. Therefore, all serum antibody-positive chickens were yolk antibody-positive, and vice versa. Accordingly, vaccine producers can estimate REV cleanliness in a poultry farm by sampling yolk antibody titers.

Integrated Analysis of miRNA and mRNA Expression Profiles in Spleen of Specific Pathogen-Free Chicken Infected with Avian Reticuloendotheliosis Virus Strain SNV.

The Reticuloendotheliosis virus (REV) primarily causes avian severe immunosuppression, in addition to other symptoms, which include avian dwarfing syndrome and chronic tumors in lymphoid and other tissue. To date, REV’s molecular mechanisms leading to immunosuppression have not been fully elucidated. In the current study, we aimed to elucidate the role of microRNAs (miRNA) in regulating gene expression during REV infections. Therefore, we used a high-dose spleen necrosis virus (SNV) model of REV to inoculate one-day-old specific pathogen-free (SPF) chickens, thereby inducing congenital infections. We analyzed miRNA and mRNA expression profiles using Next Generation Sequencing (NGS) in a total of 19 spleen samples that were collected at 7, 14, and 21 days post infection (dpi). The results showed that 63 differentially expressed miRNAs (DEmiRNAs) (30 known miRNAs and 33 novel miRNAs) and 482 differentially expressed target genes (DETGs) were identified. Integration analysis identified 886 known miRNA–mRNA and 580 novel miRNA–mRNA interaction pairs, which involved miRNAs that were inversely correlated with the above DETGs. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that the DETGs were considerably enriched in the immune-relevant pathways category, such as immune system, cell growth and death, signaling molecules and interaction, signal transduction, etc. We further verified selected immune-relevant miRNA and their DETGs while using quantitative RT-PCR (qRT-PCR). Overall, our data revealed valuable immune-related miRNA–mRNA interaction information that occurred during REV infections, thereby broadening our understanding of the REV-induced immunosuppression.

Survey of Reticuloendotheliosis Virus in Wild Turkeys (Meleagris gallopavo) in Texas, USA.

Reticuloendotheliosis virus (REV) is an immunosuppressive and sometimes oncogenic avian retrovirus that establishes lifelong infection in a wide range of avian species. REV-infected wild birds roaming near at-risk captive flocks, such as is the case for the highly endangered Attwater's Prairie Chicken (APC; Tympanuchus cupido attwateri), could act as a reservoir for viral transmission. In wild birds, prevalence rates of REV are low and appearance of associated disease is uncommon. During 2016-17, nearly half of all captive adult APC mortality at Fossil Rim Wildlife Center captive breeding facility in Glen Rose, Texas, US was attributed to REV infection. The unusually high REV prevalence rate prompted us to survey for this virus in wild galliforms throughout the region. From 2016-17, 393 blood samples collected from two subspecies of Wild Turkeys (Meleagris gallopavo) were tested for REV proviral DNA through amplification of the viral 3' long terminal repeat and segments of the viral pol gene. In REV-affected counties, 5% (5/98) of native Rio Grande Wild Turkeys (Meleagris gallopavo intermedia) were identified as REV-positive. In addition, we detected REV in one of 62 Eastern Wild Turkeys (Meleagris gallopavo silvestris) that had been imported during conservation efforts. To better determine protective measures, continued surveillance, including collection and genetic analysis of REV-infected samples, is necessary to identify sources of REV outbreaks in captive APC flocks.

The Seroprevalence Study of Reticuloendotheliosis Virus Infection in Chicken in Bangladesh

We showed seroprevalence of Reticuloendotheliosis virus (REV) in poultry in selected areas of Bangladesh.Total 3,555 serum samples were collected between 2014 and 2016 from 10 districts, 6 types of chicken and 144 flocks, tested for the specific antibodies directed against REV by indirect Enzyme Linked Immunosorbent Assay (ELISA). The overall seroprevalence of REV in the chickens tested were 21.13% (751/3,555) and 73.61% (106/144) flocks were seropositive. According to district, Gazipur had the highest (49.50%) and Khulna had the lowest (9.09%) seropositive rates.Additionally, the prevalence rates were highest 38.21% during onset of laying period, age between 19 and 24 weeks. The positivity rates were almost same round the year like 22.96% in summer, 20.70% in rainy and 20.43% in winter seasons.Furthermore, the seropositive rates in broiler flocks, broiler breeder flocks, commercial layer flocks, layer breeder flocks, sonali flocks, and domestic chicken flocks were 6.86%, 42.15%, 25.84%, 33.59%, 9.40%, and 6.49%, respectively. Conclusion: These results indicate that REV infections are widespread ubiquitous in all types of chickens and may appear as a potential threat for the rapidly growing poultry industry in Bangladesh. This is the first seroprevalencestudy on REV infections in chicken in Bangladesh.More prevalence study, molecular characterization and vaccine production from local isolates are emergency requirements to combat REV. The quality verification of imported vaccine must be strongly adopted.

Analysis of alternative splicing in chicken embryo fibroblasts in response to reticuloendotheliosis virus infection

ABSTRACTAlternative splicing (AS) plays a significant role in regulation of genomic expression at the transcriptional level and is involved in many important biological functions of cells, thus a gene can be spliced into distinct transcript variants then translated to many different kinds of protein. Reticuloendotheliosis virus (REV) is a kind of retrovirus that can infect multiple avian species, leading to runting syndrome, immunosuppression and oncogenesis. In this present study, we analyzed AS in REV-infected chicken embryo fibroblasts (CEFs) which were inoculated with the second generation of REV (group VB) and compared with normal CEFs (group C) by high-throughput RNA sequencing technology. A total of 6,939 genes which were alternatively spliced were detected, among them, skipped exon (SE) was the most common pattern. Moreover, 5,607 AS genes were detected as differentially expressed; compared with group C, group VB has 2,825 genes upregulated significantly and 2,782 genes downregulated significantly. These 5,607 differentially expressed AS genes are involved in many important biological processes. Many of them are involved in apoptosis and tumourigenesis. We also proved, by agarose gel electrophoresis, that AS events predicted by our study are authentic and AS is closely related with apoptosis and tumourigenesis in REV-infected CEFs. Our study provides the best analysis to date of the potential link between AS and CEFs in response to REV infection.Research highlightsTranscriptomics analysis of REV-infected CEFs using high-throughput sequencing.Potential link between alternative splicing and CEFs in response to REV infection.Skipped exon is the most common spliced pattern in REV-infected CEFs.Differentially expressed genes mainly involved in apoptosis and tumourigenesis.