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Avian reovirus (ARV) infection is prevalent in farmed poultry and causes viral arthritis and severe immunosuppression. The spleen plays a very important part in protecting hosts against infectious pathogens. In this research, transcriptome and translatome sequencing technology were combined to investigate the mechanisms of transcriptional and translational regulation in the spleen after ARV infection. On a genome-wide scale, ARV infection can significantly reduce the translation efficiency (TE) of splenic genes. Differentially expressed translational efficiency genes (DTEGs) were identified, including 15 upregulated DTEGs and 396 downregulated DTEGs. These DTEGs were mainly enriched in immune regulation signaling pathways, which indicates that ARV infection reduces the innate immune response in the spleen. In addition, combined analyses revealed that the innate immune response involves the effects of transcriptional and translational regulation. Moreover, we discovered the key gene IL4I1, the most significantly upregulated gene at both the transcriptional and translational levels. Further studies in DF1 cells showed that overexpression of IL4I1 could inhibit the replication of ARV, while inhibiting the expression of endogenous IL4I1 with siRNA promoted the replication of ARV. Overexpression of IL4I1 significantly downregulated the mRNA expression of IFN-β, LGP2, TBK1 and NF-κB; however, the expression of these genes was significantly upregulated after inhibition of IL4I1, suggesting that IL4I1 may be a negative feedback effect of innate immune signaling pathways. In addition, there may be an interaction between IL4I1 and ARV σA protein, and we speculate that the IL4I1 protein plays a regulatory role by interacting with the σA protein. This study not only provides a new perspective on the regulatory mechanisms of the innate immune response after ARV infection but also enriches the knowledge of the host defense mechanisms against ARV invasion and the outcome of ARV evasion of the host's innate immune response.

The complete mitochondrial genome sequence of the Cenxi classical three-buff chicken was measured by PCR-based methods and analyzed in detail. Our research findings reveal that the entire mitochondrial genome of the Cenxi classical three-buff chicken is a circular molecule consisting of 16,786 bp (GenBank accession number: KM433666). The contents of A, T, C, and G in the mitochondrial genome were found to be 30.27%, 23.75%, 32.49% and 13.49%, respectively. The complete mitochondrial genome of the Cenxi classical three-buff chicken contains a typical structure, including 13 protein-coding genes, 2 rRNA genes, 22 tRNA genes and 1 control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships among Gallus gallus domesticus mitochondrial genomes.

Interferon-inducible transmembrane protein 3 (IFITM3) is an antiviral factor that plays an important role in the host innate immune response against viruses. Previous studies have shown that IFITM3 is upregulated in various tissues and organs after avian reovirus (ARV) infection, which suggests that IFITM3 may be involved in the antiviral response after ARV infection. In this study, the chicken IFITM3 gene was cloned and analyzed bioinformatically. Then, the role of chicken IFITM3 in ARV infection was further explored. The results showed that the molecular weight of the chicken IFITM3 protein was approximately 13 kDa. This protein was found to be localized mainly in the cytoplasm, and its protein structure contained the CD225 domain. The homology analysis and phylogenetic tree analysis showed that the IFITM3 genes of different species exhibited great variation during genetic evolution, and chicken IFITM3 shared the highest homology with that of Anas platyrhynchos and displayed relatively low homology with those of birds such as Anser cygnoides and Serinus canaria. An analysis of the distribution of chicken IFITM3 in tissues and organs revealed that the IFITM3 gene was expressed at its highest level in the intestine and in large quantities in immune organs, such as the bursa of Fabricius, thymus and spleen. Further studies showed that the overexpression of IFITM3 in chicken embryo fibroblasts (DF-1) could inhibit the replication of ARV, whereas the inhibition of IFITM3 expression in DF-1 cells promoted ARV replication. In addition, chicken IFITM3 may exert negative feedback regulatory effects on the expression of TBK1, IFN-γ and IRF1 during ARV infection, and it is speculated that IFITM3 may participate in the innate immune response after ARV infection by negatively regulating the expression of TBK1, IFN-γ and IRF1. The results of this study further enrich the understanding of the role and function of chicken IFITM3 in ARV infection and provide a theoretical basis for an in-depth understanding of the antiviral mechanism of host resistance to ARV infection. [ABSTRACT FROM AUTHOR]

Chicken parvovirus (ChPV) infection can cause runting-stunting syndrome (RSS) in chickens. There is currently no commercially available vaccine for controlling ChPV, and ChPV infection in chickens is widespread globally. The rapid detection of ChPV is crucial for promptly capturing epidemiological data on ChPV. Two monoclonal antibodies (mAbs), 1B12 and 2B2, against the ChPV NS1 protein were generated. A double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was developed for detecting ChPV based on the mAb 1B12 and an anti-chicken polyclonal antibody against the ChPV NS1 protein. The detection limit for the ChPV recombinant pET32a-NS1 protein was approximately 31.2 ng/mL. A total of 192 throat and cloaca swab samples were analyzed for ChPV by the established DAS-ELISA and nested PCR methods. The concordance rate between the DAS-ELISA and the nested PCR method was 89.1%. The DAS-ELISA can detect the ChPV antigen without any cross-reaction with FAdV-4, FAdV-1, NDV, AIV, MS, CIAV, aMPV, EDSV, IBV, or AGV2. The method also has high repeatability, with a coefficient of variation (CV) of less than 5%. These findings indicate that the DAS-ELISA exhibits high accuracy, good sensitivity, and specificity, making it suitable for viral detection, field surveillance, and epidemiological studies. [ABSTRACT FROM AUTHOR]

Avian parvoviruses cause several enteric poultry diseases that have been increasingly diagnosed in Guangxi, China, since 2014. In this study, the whole-genome sequences of 32 strains of chicken parvovirus (ChPV) and 3 strains of turkey parvovirus (TuPV) were obtained by traditional PCR techniques. Phylogenetic analyses of 3 genes and full genome sequences were carried out, and 35 of the Guangxi ChPV/TuPV field strains were genetically different from 17 classic ChPV/TuPV reference strains. The nucleotide sequence alignment between ChPVs/TuPVs from Guangxi and other countries revealed 85.2–99.9% similarity, and the amino acid sequences showed 87.8–100% identity. The phylogenetic tree of these sequences could be divided into 6 distinct ChPV/TuPV groups. More importantly, 3 novel ChPV/TuPV groups were identified for the first time. Recombination analysis with RDP 5.0 revealed 15 recombinants in 35 ChPV/TuPV isolates. These recombination events were further confirmed by Simplot 3.5.1 analysis. Phylogenetic analysis based on full genomes showed that Guangxi ChPV/TuPV strains did not cluster according to their geographic origin, and the identified Guangxi ChPV/TuPV strains differed from the reference strains. Overall, whole-genome characterizations of emerging Guangxi ChPV and TuPV field strains will provide more detailed insights into ChPV/TuPV mutations and recombination and their relationships with molecular epidemiological features. [ABSTRACT FROM AUTHOR]

Chicken parvovirus (ChPV), chicken infectious anaemia virus (CIAV) and fowl adenovirus serotype 4 (FAdV-4) are avian viruses that have emerged in recent years and have endangered the global poultry industry, causing great economic loss. In this study, a multiplex fluorescence-based loop-mediated isothermal amplification (mLAMP) assay for detecting ChPV, CIAV and FAdV-4 was developed to simultaneously diagnose single and mixed infections in chickens. Three primer sets and composite probes were designed according to the conserved regions of the NS gene of ChPV, VP1 gene of CIAV and hexon gene of FAdV-4. Each composite probe was labelled with a different fluorophore, which was detached to release the fluorescence signal after amplification. The target viruses were distinguished based on the colour of the mLAMP products. The mLAMP assay was shown to be sensitive, with detection limits of 307 copies of recombinant plasmids containing the ChPV target genes, 749 copies of CIAV and 648 copies of FAdV-4. The assay exhibited good specificity and no cross-reactivity with other symptomatically related avian viruses. When used on field materials, the results of the mLAMP assay were in 100% agreement with those of the previously published PCR assay. The mLAMP assay is rapid, economical, sensitive and specific, and the results of amplification are directly observable by eye. Therefore, the mLAMP assay is a useful tool for the clinical detection of ChPV, CIAV and FAdV-4 and can be applied in rural areas. [ABSTRACT FROM AUTHOR]

Avian reovirus (ARV) is a member of the genus Orthoreovirus in the family Reoviridae and causes a severe syndrome including viral arthritis that leads to considerable losses in the poultry industry. Innate immunity plays a significant role in host defense against ARV. Here, we explored the interaction between ARV and the host innate immune system by measuring mRNA expression levels of several genes associated with the MDA5 signaling pathway. The results showed that expression peaks for MDA5, MAVS, TRAF3, TRAF6, IRF7, IKKɛ, TBK1 and NF-κB occurred at 3 days postinfection (dpi). Moreover, type I IFN (IFN-α, IFN-β) and IL-12 expression levels peaked at 3 dpi, while type II IFN (IFN-γ), IL-6, IL-17 and IL-18 expression reached a maximum level at 1 dpi. IL-8 changed at 5 dpi, and IL-1β and TNF-α changed at 7 dpi. Interestingly, several key IFN-stimulated genes (ISGs), including IFITM1, IFITM2, IFITM5, Mx1 and OASL, were simultaneously upregulated and reached maximum values at 3 dpi. These data indicate that the MDA5 signaling pathway and innate immune cytokines were induced after ARV infection, which would contribute to the ARV-host interaction, especially at the early infection stage. [ABSTRACT FROM AUTHOR]

• Four host proteins interacting with ARV σA protein were screened by a yeast two-hybrid system. • NME2 was further validated as a σA-binding protein through co-immunoprecipitation. • The key interaction domain was identified at aa 121–416 in NME2 and at aa 71–139 in σA, respectively. • Overexpression of NME2 substantially inhibited ARV replication. Our present study aimed to identify host cell proteins that may interact with avian reovirus (ARV) σA protein and their potential effect on ARV replication. The ARV structural protein σA has been demonstrated to suppress interferon production and confirmed to activate the PI3K/Akt pathway. However, host cell factors interacting with σA to affect ARV replication remain unknown. In current study, a cDNA library of chicken embryo fibroblasts (CEFs) was constructed, and host cell proteins interacting with σA were screened by a yeast two-hybrid system. We identified four candidate cellular proteins that interact with ARV σA protein. Among them, Gallus NME/NM23 nucleoside diphosphate kinase 2 (NME2) was further validated as a σA-binding protein through co-immunoprecipitation. The key interaction domain was identified at amino acids (aa) 121–416 in NME2 and at aa 71–139 in σA, respectively. We demonstrated that overexpression of NME2 substantially inhibited ARV replication. In addition silencing NME2 by small interfering RNAs (siRNAs) resulted in marked enhancement of ARV replication. Our work has demonstrated that NME2 is a σA-binding protein that may affect ARV replication in CEF cells. [ABSTRACT FROM AUTHOR]