The use of infectious bursal disease virus (IBDV) reverse genetics to engineer tagged reporter viruses has revealed that the virus factories (VFs) of the Birnaviridae family are biomolecular condensates that show properties consistent with liquid–liquid phase separation (LLPS). Although the VFs are not bound by membranes, it is currently thought that viral protein 3 (VP3) initially nucleates the formation of the VF on the cytoplasmic leaflet of early endosomal membranes, and likely drives LLPS. In addition to VP3, IBDV VFs contain VP1 (the viral polymerase) and the dsRNA genome, and they are the sites of de novo viral RNA synthesis. Cellular proteins are also recruited to the VFs, which are likely to provide an optimal environment for viral replication; the VFs grow due to the synthesis of the viral components, the recruitment of other proteins, and the coalescence of multiple VFs in the cytoplasm. Here, we review what is currently known about the formation, properties, composition, and processes of these structures. Many open questions remain regarding the biophysical nature of the VFs, as well as the roles they play in replication, translation, virion assembly, viral genome partitioning, and in modulating cellular processes.

Infectious bursal disease virus (IBDV) vaccines do not induce sterilizing immunity, and vaccinated birds can become infected with field strains. Vaccine-induced immune selection pressure drives the evolution of antigenic drift variants that accumulate amino acid changes in the hypervariable region (HVR) of the VP2 capsid, which may lead to vaccine failures. However, there is a lack of information regarding how quickly mutations arise, and the relative contribution different residues make to immune escape. To model IBDV antigenic drift in vitro, we serially passaged a classical field strain belonging to genogroup A1 (F52/70) ten times, in triplicate, in the immortalized chicken B cell line, DT40, in the presence of sub-neutralizing concentrations of sera from birds inoculated with IBDV vaccine strain 2512, to generate escape mutants. This assay simulated a situation where classical strains may infect birds that have suboptimal vaccine-induced antibody responses. We then sequenced the HVR of the VP2 capsid at passage (P) 5 and 10 and compared the sequences to the parental virus (P0), and to the virus passaged in the presence of negative control chicken serum that lacked IBDV antibodies. Two escape mutants at P10 had the same mutations, D279Y and G281R, and a third had mutations S251I and D279N. Furthermore, at P5, the D279Y mutation was detectable, but the G281R mutation was not, indicating the mutations arose with different kinetics.

Infectious bursal disease virus (IBDV) causes an endemic immunosuppressive disease in chickens. Prior exposure to IBDV influences the pathogenesis and shedding of chicken strains of influenza A virus (IAV), but its effect on waterfowl strains is poorly understood. To address this, we inoculated 14-day-old specific pathogen-free chickens with low pathogenicity avian influenza strain A/Mallard/Alberta/156/01 (H3N8) and compared the replication, shedding, pathogenesis, transmission and intra-host evolution between immunocompetent chickens and chickens that had IBDV-mediated immune dysregulation due to a prior infection with strain F52/70 at 2 days of age. The IAV replicated in the upper respiratory tract, and the virus was shed from the oropharyngeal cavity, but there was no shedding from the cloaca and no transmission to sentinel chickens. IAV replication in chickens was associated with amino acid substitutions in the polymerase complex and HA. Prior IBDV infection had no significant effect on IAV pathogenicity, replication or shedding and had a modest effect on IAV diversity, increasing the number of amino acid substitutions from an average of 2.50 substitutions per sample ( sd ±1.83) in the Mock/IAV group to 4.75 ( sd ±1.81) in the IBDV/IAV group ( P <0.01). Taken together, our data suggest that IBDV is unlikely to play a major role in the spillover or spread of waterfowl IAV strains in chicken flocks, although it could expand IAV diversity. This information is useful for informing preventative measures for controlling IAV in poultry flocks.