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Biosafety of genetically-modified baculoviruses for insect control

Objective

The objective is the genetic engineering and field testing of biosafe baculovirus insecticides for control of insect pests in agriculture. This objective will be achieved by constructing viruses with increased virulence, but reduced persistence and survival in the environment.
Baculoviruses are insect pathogens which are successfully used as biological control agents of insect pests in agriculture and forestry. Although baculoviruses are effective, there is a necessary period when disease must develop in the pest. During this time severe damage to the crop may occur. This slow speed of action is a drawback to a much wider commercial application of these viruses and is most relevant for crops with low damage thresholds. Genetically modified baculoviruses with improved insecticidal properties could overcome this problem and may lead to the wider use of this ecorational and more environmentally sound control approach.

The objective of the project is the construction of baculoviruses with enhanced insecticidal activity by with reduced survival in the field. This will be achieved by introduction of a 'built-in' suicide mechanism or gene deletions into baculoviruses, that would impair their survival but not their short-term efficacy.

Work to date has been to start the construction of the various recombinant viruses; to determine their biological activity and host range; and to preliminarily test those recombinants already available in a microcosm. Experiments have been initiated to produce a 'suicide' Autographa californica nucleo-polyhedrovirus (AcNPV) insecticide. Baculovirus transfer vectors have been constructed based on the polyhedrin gene region (containing the hybrid polyhedrin gene promoter lacO) and the p10 gene region (containing the lacI coding sequences). These have been characterized at the sequence level to confirm authenticity. The construction of baculovirus recombinants with deletions of genes that affect the virus persistence in the field has been focused on three genes which have been described to be not essential for viral replication, to increase virulence, and to reduce persistence of the virus. These genes are the p10, the polyhedron-envelope (pp34) and the ecdysteroid UDP-glycosylk transferase (egt)gene.

The biological and behavioural properties of an AcNPV recombinant AS3 (p10-minus) and wild type AcNPV were analyzed in bio-assays and in a microcosm. The biological activity of recominant AcNPV-AS3 and wild type AcNPV expressed as LC50-value was compared in standard bioassays. Five concentrations were used against first instar larvae of Spodoptera exigua and Autographa gamma. No statistically significant differences in infectivity was determined between wild type and recombinant. A field release of the genetically modified AcNPV-AS3 and wild type AcNPV was simulated in a contained model-ecosystem, or microcosm. Equal quantities of the wild type AcNPV and AcNPV-AS3 (p10-minus) applied to 200 larvae of S exigua showed no difference in response to infection (82 and 81% mortality).
Baculoviruses are insect pathogens which are successfully used as biological control agents of insect pests in agriculture and forestry as alternatives to chemical insecticides. Safety testing has confirmed that these natural viruses are insect specific and cause no hazards to other animals and to plants. A major drawback to a much wider commercial application of these viruses is their slow speed of action, which is most relevant for crops with low damage thresholds. A number of potentially useful baculoviruses with enhanced insecticidal activity have been engineered. Within the BAP program genetically marked viruses were constructed to aid in ecological studies, such as on host range and persistence of wild type viruses. In addition, a microcosm has been designed and tested, and limited field releases using recombinant baculoviruses, notably Autographa californica nuclear polyhedrosis virus (AcNPV) have been executed.

The current research builds on this knowledge and expertise as well as on developments elsewhere and intends to evaluate the biosafety of engineered baculoviruses which are more effective, but also possess reduced capacity for survival in the field. In this project baculoviruses, in particular AcNPV and possibly other baculoviruses, will be marked and provided with a built-in suicide mechanism. In addition, deletions will be introduced in the virus which reduce its persistence and survival in the environment. The recombinant viruses will undergo extensive laboratory and biosafety testing (host range testing and microcosms evaluations) before, in a second phase, experimental releases in the environment are considered.

Ultimately, this project will increase our understanding of the ecology of genetically modified baculoviruses and also allow predictions to be made about their behaviour in the environment.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

Landbouwuniversiteit Wageningen
Address
11,Binnenhaven
6709 PD Wageningen
Netherlands

Participants (2)

FEDERAL BIOLOGICAL RESEARCH CENTRE FOR AGRICULTURE AND FORESTRY (BBA)
Germany
Address
Heinrichstrasse 243
64287 Darmstadt
NERC Institute of Virology and Environmental Microbiology
United Kingdom
Address
Mansfield Road
OX1 3SR Oxford