Immunogenic analysis of two DNA vaccines of avian reovirus mediated by attenuated Salmonella typhimurium in chickens
ABSTRACT Avian reovirus (ARV) is an important pathogen in poultry industry and causes great economic losses. As attenuated Salmonella typhimurium is already being used as an effective vehicle for the transfer of DNA vaccines, so in this study we evaluated two DNA vaccines mediated by S. typhimurium on their ability of eliciting antibody production. SPF chickens were respectively immunized with SL7207 (pVAX-σB), SL7207 (pVAX-σC) and SL7027 (pVAX-σB-σC) three times. The results showed that the antibody production was highly dependent on the immunizing times, detectable antibodies of serum antibody IgG and small intestinal mucosal antibody IgA were generated at week 4 and were further improved at week 6 and antibody titers in group SL7207 (pVAX-σC) were higher than that in group SL7207 (pVAX-σB), demonstrating that SL7207 (pVAX-σC) was more powerful than SL7207 (pVAX-σB) in antibody production. The higher antibody titer in SL7027 (pVAX-σB-σC) than that in SL7207 (pVAX-σC) group showed that co-expressing σB and σC could improve antibody production. IFN-γ detection showed that significant higher IFN-γ was generated both in groups SL7027 (pVAX-σB-σC) and SL7207 (pVAX-σC). Subsequent challenge showed that SL7207 (pVAX-σB), SL7207 (pVAX-σC) and SL7027 (pVAX-σB-σC) conferred 50%, 75% and 87.5% respectively.
[Show abstract] [Hide abstract]
ABSTRACT: SYBR Green I based real-time RT-PCR assay was developed for the detection and quantification of duck reovirus (DRV) using ABI PRISM 7500 sequence detection system. The assay was carried out using a set of primer designed to amplify highly conserved sequences of S2 gene of DRV. A 10-fold dilution series assay using a plasmid containing the cDNA of DRV S2 gene demonstrated the high sensitivity of the assay with a lowest detection limit of ≤1.48 copies/μL. Standard deviation and coefficient of variation were low for both intra-assay and inter-assay variability. The assay performance was evaluated on 80 samples obtained from artificially infected Cherry Valley ducklings and 10 field specimens compared with the conventional RT-PCR assay. It was shown that 10 artificially infected samples tested negative in gel-based assay were positive for the real-time RT-PCR. DRV could be detected in all eight different tissues collected from the ducklings infected artificially. In contrast, the higher detection rate was obtained in the bursa of fabricius (90%), lung (90%), spleen (80%), and thymus (70%) than that in the liver (30%) as well as in the pancreas (10%). This method was rapid, specific, and sensitive for the detection of DRV and will be useful in veterinary diagnostic applications.Avian biology research 08/2014; 7(3). DOI:10.3184/175815514X14064587194329 · 0.90 Impact Factor
[Show abstract] [Hide abstract]
ABSTRACT: Muscovy duck reovirus (MDRV) is a highly pathogenic virus in waterfowl and causes significant economic loss in the poultry industry worldwide. Because the host innate immunity plays a key role in defending against virus invasion, more and more attentions have been paid to the immune response triggered by viral infection. Here we found that the genomic RNA of MDRV was able to rapidly induce the production of interferons (IFNs) in host. Mechanistically, MDRV infection induced robust expression of IFNs in host mainly through RIG-I, MDA5 and TLR3-dependent signaling pathways. In addition, we observed that silencing VISA expression in 293T cells could significantly inhibit the secretion of IFNs. Remarkably, the production of IFNs was reduced by inhibiting the activation of NF-κB or knocking down the expression of IRF-7. Furthermore, our study showed that treatment of 293T cells and Muscovy duck embryo fibroblasts with IFNs markedly impaired MDRV replication, suggesting that these IFNs play an important role in antiviral response during the MDRV infection. Importantly, we also detected the induced expression of RIG-I, MDA5, TLR3 and type I IFN in Muscovy ducks infected with MDRV at different time points post infection. The results from in vivo studies were consistent with those in 293T cells infected with MDRV. Taken together, our findings reveal that the host can resist MDRV invasion by activating innate immune response involving RIG-I, MDA5 and TLR3-dependent signaling pathways that govern IFN production. Copyright © 2014 Elsevier B.V. All rights reserved.Veterinary Microbiology 12/2014; 175(2-4). DOI:10.1016/j.vetmic.2014.12.004 · 2.73 Impact Factor