Assembly of Recombinant Israeli Acute Paralysis Virus Capsids

Abstract

The dicistrovirus Israeli Acute Paralysis Virus (IAPV) has been implicated in the worldwide decline of honey bees. Studies of IAPV and many other bee viruses in pure culture are restricted by available isolates and permissive cell culture. Here we show that coupling the IAPV major structural precursor protein ORF2 to its cognate 3C-like processing enzyme results in processing of the precursor to the individual structural proteins in a number of insect cell lines following expression by a recombinant baculovirus. The efficiency of expression is influenced by the level of IAPV 3C protein and moderation of its activity is required for optimal expression. The mature IAPV structural proteins assembled into empty capsids that migrated as particles on sucrose velocity gradients and showed typical dicistrovirus like morphology when examined by electron microscopy. Monoclonal antibodies raised to recombinant capsids were configured into a diagnostic test specific for the presence of IAPV. Recombinant capsids for each of the many bee viruses within the picornavirus family may provide virus specific reagents for the on-going investigation of the causes of honeybee loss.

In-depth proteomic analysis of Varroa destructor: Detection of DWV-complex, ABPV, VdMLV and honeybee proteins in the mite

“…Proteomic identification is an alternative strategy to detect viruses at the protein level and is more accurate than traditional immunochemical methods such as the application of a polyclonal antibody against the viral capsid protein VP1 of DWV and VDV-1 to confirm viral presence in Varroa 86 . However, antibody-based diagnostic assays are an alternative tool that has good applications in honeybee field conditions, as proposed for the monoclonal antibody against VP2 of IAPV 87 , and is essential in histological localization studies 56 . The advantage of MSP is that it enables the detection of a range of virus/pathogen proteins in one run from a heterogeneous sample rather than detecting only the epitope against which the antibody reacts.…”

Plant and insect virus‐like particles: emerging nanoparticles for agricultural pest management

“…For instance, significant upregulation of immune-and RNAi-related genes is observed in the extracted midgut tissues of silkworm larvae when incubated with CPV-based VLPs in vitro, 96,192 providing alternative antiviral agents for the silkworm. In addition, IAPV-based VLPs can stimulate the production of anti-IAPV monoclonal antibodies in mice, 113 which indicates that IAPV VLPs could act as immunitystimulating antigens and might also trigger antiviral responses in bees. The first bee vaccines that are being developed are based on inactivated microbial pathogens 193 but the use of VLPs may become adopted because of biosafety or specificity against viral pathogens.…”

“…Israeli acute paralysis virus (IAPV) ( Aparavirus ) can infect honeybees and cause severe losses in bee colonies. Similar to TrV‐based VLPs, IAPV‐based VLPs of the desired morphology and size require the correct cleavage of IAPV structural polyprotein (P1, 100 kDa) into mature coat protein (VP2, 39 kDa) by a 3C‐like protease 113 . More importantly, IAPV‐based VLPs can stimulate the production of monoclonal antibodies in mice, indicating their feasibility for displaying antigens and stimulating the immune response.…”

Viral Delivery of dsRNA for Control of Insect Agricultural Pests and Vectors of Human Disease: Prospects and Challenges

“…Also the baculovirus expression system has been used for production of VLPs based on insect viruses [e.g. nodavirus (Krishna et al, 2003 ); bidensovirus (Pan et al, 2014 ); dicistrovirus (Ren et al, 2014 ); tetravirus (Mendes et al, 2015 ); and alphavirus (Hikke et al, 2016 )] and these could be directly used as specific delivery vehicles when loading with specific cargo is achieved. After assembly of the VLPs and their cargo in vitro , they could be further purified and directly applied to insects to investigate insecticidal effects.…”