Propagation and titration of infectious bursal disease virus, including non-cell-culture-adapted strains, usingex vivo-stimulated chicken bursal cells

Abstract

Infectious bursal disease virus (IBDV) is a Birnaviridae family member of economic importance for poultry. This virus infects and destroys developing B lymphocytes in the cloacal bursa, resulting in a potentially fatal or immunosuppressive disease in chickens. Naturally occurring viruses and many vaccine strains are not able to grow in in vitro systems without prior adaptation, which often affects viral properties such as virulence. Primary bursal cells, which are the main target cells of lymphotropic IBDV in vivo, may represent an attractive system for the study of IBDV. Unfortunately, bursal cells isolated from bursal follicles undergo apoptosis within hours following their isolation. Here, we demonstrate that ex vivo stimulation of bursal cells with phorbol 12-myristate 13-acetate maintains their viability long enough to allow IBDV replication to high titres. A wide range of field-derived or vaccine serotype 1 IBDV strains could be titrated in these phorbol 12-myristate 13-acetate -stimulated bursal cells and furthermore were permissive for replication of non-cell-culture-adapted viruses. These cells also supported multistep replication experiments and flow cytometry analysis of infection. Ex vivo-stimulated bursal cells therefore offer a promising tool in the study of IBDV.

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

“…Our work complements that of S ebastien Soubies et al in the lab of Nicolas Eterradossi, who have recently demonstrated that phorbol 12-myristate 13-acetate (PMA)-stimulated B cells can also support IBDV replication [40]. Ex vivo models of IBDV infection allow virus-host cell interactions to be studied in the relevant host cells, retaining the advantages of in vivo studies without the limitations of multiple cell types that might confound the data.…”

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

“…In addition, it is important to isolate the primary cells as soon as possible after the organ harvest to limit cell death. The need to use chCD40L is a limitation of the technique; however, work conducted by Soubies et al shows that the use of phorbol 12-myristate 13-acetate (PMA) to prolong bursal cell viability instead of chCD40L 14 may enable the model to be adopted by a greater number of laboratories. The protocol outlined above determines the optimal concentration of chCD40L empirically, by culturing primary B cells in serially diluted concentrations of the molecule and observing cell proliferation and viability.…”

“…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.…”

Analysis of computer network attack based on the virus propagation model

“…The reason for the buffer overflow is that the parameters entered by the user are not carefully checked in the program. For example, it is as the following program [15]: void function(char *str) { char buffe r [16]; strcpy(buffer,str); } The strcpy () above will directly transform the content from str to copy into the buffer. So long as the length of str is greater than 16, it will cause buffer overflow and cause the program to run wrong.…”

“…The most common way is to create a buffer overflow to enable the program to run a user shell and execute other commands through shell. If the program belongs to root and has suid permissions, the attacker gets a shell with root permissions and can operate any on the system [16].…”