Apoptosis Differences Induced By Very Virulent And Attenuated Infectious Bursal Disease Virus

Infectious bursal disease(IBD) is an acute, highly contagious, and lethal infectious disease of young chickens caused by infectious bursal disease virus(IBDV). Infectious bursa disease virus infects chicken bursa, causing the immune suppression of chicken. There are two serotypes and only serotype I causes disease. Because of two major virus evolutions, the pathogenic serotype I virus includes classical strain, antigenic variant strain and very virulent IBDV(vv IBDV) strain. At present, the research on IBDV gene function and infection mechanism is mainly based on the cell adapted strains or classical strains. The difference in infection mechanism of the attenuated and very virulent strains needs to be further investigated.In order to study the differences in clinical characteristics, pathogenic tissue pathology, and apoptosis and necrosis in target cell between attenuated and very virulent strains, vv IBDV Gx and attenuated Gt were used to infect 3-week-old SPF chickens. Compared with attenuated Gt, vv IBDV Gx has a high mortality rate(80%) and causes rapid apoptosis and necrosis of target cells, and causes severe pathological changes in the bursa, which leads to serious damage to the immune system and acute death of chicken.In order to reduce the level of apoptosis of bursal B lymphocyte in vitro, and to investigate the infection mechanism of very virulent and attenuated strains of IBDV in bursal B cells. We try to express the functional domain of chicken CD154 in a secretory way. The open reading frame of chicken CD154 was cloned based on the sequence in Gen Bank. The tandem protein was comprised with the N-terminal of mouse CD8α and C-terminal of chicken CD154. Additionally, the signal peptide was fused to induce secretory expression of fusion protein. The results showed that the fusion protein expressed successfully and the amount of fusion protein increased at 48 hours post transfection. It is important that the fusion protein can secrete out of cells. The expression of functional domain of chicken CD154 supports the investigation of infection mechanism of different strains of IBDV in bursal B cells in vitro.Based on the two parts above, the attenuated and very virulent IBDV were used to infect the chicken primary B lymphocytes in vitro. TCID50 or ELD50 assay was employed to titrate the infectious virus titer of attenuated Gt or very virulent Gx, respectively. Additionally, real-time RT-PCR was used to quantify virus load of different strains. Results showed that compared with Gt, the very virulent Gx replicate rapidly in B lymphocytes in vitro, and higher viral protein load and viral RNA load were detected in Gx inoculated B cells.To sum up, very virulent infectious bursal disease virus cause acute death of infected chicken, the main reason is that vv IBDV replicate and proliferate in the bursa of Fabricius rapidly, and causes a number of B lymphocytes rapid apoptosis and necrosis, which cause lymphoid tissue rapid lesions, resulting in death of infected chickens rapidly and acutely. In this study, we compared the differences in replication characteristics between very virulent and attenuated infectious bursal disease virus in vivo and in vitro. Primary B cells in vitro were used to evaluate the replication characteristics of different strains in target cells.