Differential gene expression in chicken primary B cells infected ex vivo with attenuated and very virulent strains of infectious bursal disease virus (IBDV)

Abstract

Infectious bursal disease virus (IBDV) belongs to the family Birnaviridae and is economically important to the poultry industry worldwide. IBDV infects B cells in the bursa of Fabricius (BF), causing immunosuppression and morbidity in young chickens. In addition to strains that cause classical Gumboro disease, the so-called 'very virulent' (vv) strain, also in circulation, causes more severe disease and increased mortality. IBDV has traditionally been controlled through the use of live attenuated vaccines, with attenuation resulting from serial passage in non-lymphoid cells. However, the factors that contribute to the vv or attenuated phenotypes are poorly understood. In order to address this, we aimed to investigate host cell-IBDV interactions using a recently described chicken primary B-cell model, where chicken B cells are harvested from the BF and cultured ex vivo in the presence of chicken CD40L. We demonstrated that these cells could support the replication of IBDV when infected ex vivo in the laboratory. Furthermore, we evaluated the gene expression profiles of B cells infected with an attenuated strain (D78) and a very virulent strain (UK661) by microarray. We found that key genes involved in B-cell activation and signalling (TNFSF13B, CD72 and GRAP) were down-regulated following infection relative to mock, which we speculate could contribute to IBDV-mediated immunosuppression. Moreover, cells responded to infection by expressing antiviral type I IFNs and IFNstimulated genes, but the induction was far less pronounced upon infection with UK661, which we speculate could contribute to its virulence.

An <em>Ex Vivo</em> Chicken Primary Bursal-cell Culture Model to Study Infectious Bursal Disease Virus Pathogenesis

“…IBDV VP4 expression increased to 16,603 copies at 48 h postinfection with D78 and 38,632 copies at 48 h postinfection with UK661. Taken together, these data demonstrate that the chicken primary bursal cells could support the replication of cell-culture-adapted and very virulent IBDV strains 13 .…”

“…When chicken primary bursal cells were cultured in the presence of soluble chCD40L, the number of cells increased fourfold from 9.02 x 10 5 to 3.63 x 10 6 per mL over a period of 6 days, in contrast to when it was absent ( p <0.05) ( Figure 1A ). Cell viability was also significantly improved, for example from 25% at day 3 post-culture in the absence of chCD40L to 48% in the presence of chCD40L ( p <0.05) ( Figure 1B ) 13 .…”

“…Following identification of the gene encoding chicken CD40L (chCD40L), a soluble fusion protein was engineered that, when added to the culture media, induced the proliferation of chicken primary B cells ex vivo 12 . In 2015, B cells cultured in this fashion were found to support the replication of MDV 2 and in 2017, we and others found that primary bursal cells stimulated with chCD40L could be used as a model to study IBDV replication 13 14 . Here, we describe the isolation and culture of chicken primary bursal cells, the infection of the cells with IBDV, and the quantification of viral replication.…”

The stronger downregulation ofin vitroandin vivoinnate antiviral responses by a very virulent strain of infectious bursal disease virus (IBDV), compared to a classical strain, is mediated, in part, by the VP4 protein

“…However, the molecular basis for this increased virulence remains poorly understood [24, 25]. We have previously shown that the vvIBDV UK661 was able to down-regulate type I IFN and a selection of ISGs to a greater extent than a vaccine strain, D78, in primary B cells cultured and infected ex vivo [26]. Here, we extend these observations by demonstrating that UK661 is also able to down-regulate type I IFN and pro-inflammatory cytokine responses compared to a classical field strain, and we confirm that this occurs not only in vitro , but also in vivo (Figures 2 and 3).…”

Chicken eEF1α is a Critical Factor for the Polymerase Complex Activity of Very Virulent Infectious Bursal Disease Virus

“…Virus infection induced alterations in expression of some important signal transduction host genes including BCR signaling and antigen presentation [13]. Other studies used microarray to investigate gene expression profiles of chicken B cells infected with vvIBDV compared with aIBDV and found that key genes involved in B cell activation and signaling were down-regulated in infected compared to mock-infected cells, which may contribute to IBDV-mediated immunosuppression [14]. These studies provided important insights into host responses to vvIBDV infection and virus pathogenesis.…”