2010 Annual Report
1a.Objectives (from AD-416)
Genetically engineer specific fluorescent protein genes into baculoviruses that will be expressed in the envelopes of budded virus and occlusion bodies to serve as barriers against ultraviolet inactivation. Incorporate specific proteins into polyhedra (occlusion bodies) of baculoviruses by a genetic fusion process that would lead to early mortality or inhibit feeding of pest insects. Examine the basis of baculovirus resistance at the cellular level employing cell culture methodology in order to better understand the nature of baculovirus host range.
1b.Approach (from AD-416)
This proposal employs a molecular approach whereby baculoviruses will be stabilized against inactivation by ultraviolet light (UV) as well as to make them faster acting. To achieve these goals, several fluorescent protein genes will be engineered into baculovirus genomes which when expressed in the envelopes of the viral particles and occlusion bodies will provide protection against inactivation by UV light. This methodology is designed to reduce damage to the viral DNA of baculoviruses. To make the baculoviruses faster acting, specific proteins will be incorporated into occlusion bodies of baculoviruses by a genetic fusion process that will lead to early mortality or inhibit feeding by pest insects since viral replication in the host is not a pre requisite for elaboration of the active proteins. This research project will also examine the nature of baculovirus resistance at the cellular level employing insect cell lines that are refractile to baculoviruses. Recombinants with fluorescent markers under various promoters will elucidate where the block occurs and thus provide knowledge that could be valuable in understanding viral host range.
This project was terminated on March 25, 2010 and replaced with project 3622-22000-032-00D. Significant progress was made in two of the three project objectives, both of which focused on pest control in Component 2B. Under Objective 1, progress was made in developing baculoviruses that demonstrated protection against UV light which is a significant drawback to the field use of baculoviruses. Under Objective 3, significant progress was made in identifying host proteins that are in involved in resistance to baculovirus infection of cells. This work significantly moved forward the understanding of how the host range of viruses is determined. Because little progress was made on the second objective, a new objective was substituted two years ago. The new objective was to characterize new populations of insect viruses for development as control measures in niche markets and/or control of invasive species. Substantial progress was made on this objective including the characterization of three different viruses with the potential for use as biological control agents.
Host proteins and viral host range. Identifying how an insect blocks a baculovirus infection is important in developing tools to undermine the insect immune system for greater control of specific pest insects. By separating host proteins, ARS researchers in Columbia, MO discovered 25 proteins in corn earworm and budworm cell lines that were either present in greater or less abundance when cells were infected with baculovirus. These proteins and this information can be used to identify how the insect host range of a baculovirus can be expanded to control a wider range of pest insects. Expanding the host range will allow baculoviruses to be used as biopesticides in a wider array of crops.
Popham, H.J., Grasela, J.J., Goodman, C.L., Mcintosh, A.H. 2010. Baculovirus Infection Influences Host Protein Expression in Two Established Insect Cell Lines. Journal of Insect Physiology. Available: http://dx.doi.org/10.1016/j.jinsphys.2010.03.024.