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Optimisation of biological control through disruption of insect immune defences - PS2115

Description
The overall objective of the work is to devise improved and/or new crop protection strategies as effective alternatives to the use of broad-spectrum, neurotoxic pesticides. We aim to do this by selectively suppressing the key immune responses of target pest insects which ordinarily protect them from potential pathogens in their environment, and those used in biocontrol strategies.

Insects, including agricultural and horticultural pests, possess an impressive armoury of immune responses (both cellular and humoral) which help to protect them against potential pathogens, including fungi, bacteria and/or protozoa which might be employed in biocontrol strategies. We have already determined that adult female Pimpla hypochondriaca (an endoparasitic wasp which lays its eggs in the haemocoel of its host), injects a venom into the host prior to ovipostion, in order to suppress host immune responses which would otherwise kill the wasp progeny. A number of the active venom factors have been biochemically identified and in the current project we aim to further isolate and characterize these, and others, and then clone their genes from a venom gland cDNA library. The genes for selected immunosuppressive factors will be expressed in bacteria, insect cells and/or yeast, and recombinant protein produced. Ultimately, these proteins will be utilise in concept-proving experiments to demonstrate practical application i.e. by determining that suppression of key immune responses in a target pest insect increases the biopesticidal properties and efficacy of commercially available biocontrol agents.

This approach has the potential to lead to the development of novel biocontrol strategies that are target-specific, effective, and environmentally friendly and sustainable. These aims address Defra policy objectives relating to the minimization or eradication of broad spectrum, toxic pesticides (including organophosphates), which in turn responds to professional and public concern about their current and future use. It is anticipated that the results generated from this work will enable us to forge alliances with relevant companies so that they can be further developed into novel, environmentally sensitive and sustainable pest management strategies, which will offer the grower realistic levels of efficacy with negligible chance of adverse impact on the environment.
Objective
The primary aim of the proposed work is to generate the ‘tools’ we need (i.e. isolated venom factors, and their genes and recombinant protein) to prove the concept that suppression of key immune responses in a target pest insect increases the bio-pesticidal properties and efficacy of commercially available bio-control agents.

The specific objectives of the work are as follows:

1. To extend the characterization of factors present in Pimpla hypochondriaca venom which manipulate (suppress) immune defences in our model pest insect, Lacanobia oleracea. In particular, to optimise a PCR procedure for cloning genes for anti-haemocyte factors, and their insertion into a suitable expression vector(s). In the first instance, these will be used to transform bacteria, and pre-tests performed to optimise a procedure for obtaining recombinant protein. If recombinant protein expression in bacteria is not feasible for the particular genes tested, yeast and/or insect cells will be tried as an alternative. This work will be performed initially with the gene for one (proven) immunosuppressive factor (VP3), and later on will be extended to the genes for other candidate immunosuppressive factors.

2. To use a variety of biochemical techniques and assays (most already optimized), in order to ascertain that the recombinant proteins produced in bacteria (see 1 above) are bio-chemically and functionally identical to their ‘native’ counterparts in venom. This is important to control for any ‘mutant’ proteins which may be produced, for example as a result of gene splicing, mistakes in transcription, misfolding of the protein, etc.

3. To develop procedures for isolating/harvesting relatively large amounts of the recombinant proteins produced in bacteria or, if necessary, insect cells or yeast. This is a prerequisite to performing functional (concept-proving) experiments that will require more recombinant protein than necessary for the initial (screening) experiments.

4. To utilise recombinant immunosuppressive proteins in concept-proving experiments to determine if suppression of key immune responses in selected target pest insects increases their susceptibility to commercially available bio-control agents, thereby effectively increasing the bio-pesticidal properties of such agents.

The project is based on a two year work plan, with a break point at the end of the first year to review scientific progress and likelihood of being taken up by LINK or other matched funding at the end of the second year.
Project Documents
• Final Report : Manipulation of Insect Immune Defences to Optimise Biological Control   (8955k)
Time-Scale and Cost
From: 2006

To: 2008

Cost: £335,158
Contractor / Funded Organisations
Central Science Laboratory
Keywords
Biopesticide              
Chemicals              
Crop Pests              
Minimisation              
Natural              
Pest and Weed Control              
Pest Control              
Pesticide use              
Pesticides              
Plant              
Plants and Animals              
Fields of Study
Pesticide Safety