| Avian leukosis is caused by the viral infections of avian leukosis virus(ALV) and avian sarcoma virus(ASV). According to the differences of virus gp85 protein antigenicity related to the host specificity and cross neutralization test, etc., avian leukosis mainly divided into A,B, C, D, E and J subgroup. Thereinto, the J subgroup has the strongest pathogenicity and infectious to commercial chicken. ALV-J spread mainly through horizontal contact and congenital infection to laying hens, broiler chickens and our local strains, and result in lower flock performance, harm the immune suppression, cancer death, etc. Especially the subclinical infection caused by ALV-J result in poor performance of infected chickens and immune suppression. And thus, the chickens are more susceptible to other viruses and bacteria,causing significant economic losses poultry industry. To date, no effective vaccines are available because of the intricate genetic sequence and antigenic variability of ALV-J.Persistent surveillance and elimination of infected chickens remain the routine method to control ALV-J spread. Although some large breeding companies have successfully eradicated ALV-J from breeding flocks, a certain incidence of ALV-J infection remains in the poultry industry, such as in raw flocks, in which the eradication of ALV-J was omitted, and in local breeds, especially in some developing countries. Thus, an effective way to prevent and control ALV-J infection is crucial to reduce the losses caused by ALV-J infection. Taishan Pinus massoniana pollen polysaccharide(TPPPS), a natural plant macromolecule, demonstrates immune regulating activity and anti-viral potential. However, whether TPPPS can significantly inhibit ALV-J infection remains unclear.The focus of the present study was to explore the inhibitory effect and mechanism of TPPPS on ALV-J infection. We examined for the first time the anti-ALV-J activities of TPPPS in cells cultured in vitro and then investigated the interaction between TPPPS and ALV-J to reveal the probable mechanism that interferes with viral replication. Moreover, we artificially established a congenitally infected chicken model to simulate the natural infection of ALV-J and then assessed the anti-viral effects of TPPPS in vivo. The inhibitory effect ofTPPPS on tumor development induced by challenging the chickens with the highly oncogenic strain ALV-J was also evaluated. This study was divided into the following three parts:1. Antiviral activity mechanism analysis of TPPPS in vitro1.1 The antiviral activity of TPPPS in vitroTo examine the anti-viral characteristics of TPPPS against ALV-J, we determined the viral replication kinetics in DF-1 cells treated with different TPPPS concentrations, and then determine the reaction phase of TPPPS in the whole process of viral replication. These results imply that TPPPS can remarkably inhibit ALV-J replication in vitro and that the inhibitory effect of TPPPS was optimal at the viral adsorption phase.1.2 Transmission electron microscope(TEM) assayThe viral adsorption capability to the DF-1 cells treated with TPPPS was assessed by TEM tomography. The result shows that TPPPS treatment reduced the number of viral particles that are in the phase of membrane fusion on the cell membrane surface of the infected cells and the virions in the the endosomes in ALV-J-infected cells. This phenomenon indicates that TPPPS reduced viral adsorption and infection to the cells.Subsequently, we again employed TEM to investigate the interaction between TPPPS and ALV-J particles. The result shows that the viral particles treated with TPPPS were found to gather into clusters, and the virions exhibited irregular shapes, low or none electron-density cores, and scattered fragments around the cluster, which indicates a direct impact of TPPPS to ALV-J particles.2. The immunomodulation and antiviral effects of TPPPS on congenitally ALV-J-infected chickens2.1 Establishment of a congenitally ALV-J-infected chicken modelClinically, ALV-J spreads mainly through the congenitally infected embryos to their hatched chickens, which induce immunosuppression and persistent viremia. In this study,seven-day-old chicken embryos were challenged with different concentrations(101,102,103,104 TCID50) of ALV-J to compare the hatchability. After hatching, the maximum challenge dose that allowed nearly 100% survival of the chickens was 103 TCID50.2.2 Antiviral effects of TPPPS on congenitally ALV-J-infected chickensIn our previous studies, we found that TPPPS has anti-viral activity against subgroup B ALV. In this study, we analysised the anti-ALV-J activities of TPPPS in congenital infected chickens. Our findings revealed that successive TPPPS administration remarkably reduced viral shedding to the external environment. Moreover, the initial viral shedding was delayed for approximately 2 weeks, thereby greatly reducing the risk of early horizontal transmission to uninfected chickens. We also found that TPPPS administration significantly reduced the viral loads of the infected chickens in vivo as reflected mainly in the immune organs. These results indicate the significant anti-ALV-J activites of TPPPS in vivo.2.3 Immunomodulation of TPPPS on congenitally ALV-J-infected chickensIt is reported that TPPPS has immune recovery effect to immune suppression mice caused by cyclophosphamide. In this study, congenital ALV-J infection in the early embryonic developmental stage rapidly induced a distinct immune suppression in the chickens after hatching, especially in the first 2 weeks. However, sustained TPPPS delivery can substantially relieve the impaired immunity, as characterized by increase in immune organ index, enhance in lymphocyte number and cytokine production. These results indicate that TPPPS can be used as an immune enhancer and play role in reducing the immune suppression caused by ALV-J.3. Inhibitory activity of TPPPS against acute oncogenic ALV-J infection-induced tumorsThe acute oncogenic Fu-J virus is a replication-defective virus that can rapidly induces tumors owing to its specific v-fps oncogene. To investigate the inhibitory activity of TPPPS against ALV-J infection-induced tumors, we administered TPPPS before and/or after challenging the chickens with tumor leachate containing Fu-J virus, and monitored the mortality, clinical symptoms, and tumor development daily. The results demonstrated that TPPPS remarkably delayed tumor occurrence and reduced tumor development in theoncogenic ALV-J-infected chickens, although TPPPS cannot completely eliminate tumors.Pre-administration of TPPPS can exert a strengthening tumor-suppressive effect before viral infection... |
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