Next generation biopesticides for environmentally benign control of crop pests

Lead Research Organisation: Durham University
Department Name: Biosciences

Abstract

Pesticides are an essential tool for increasing agricultural productivity and meeting the challenge of providing increased food supplies from diminishing land areas available for cultivation. Invertebrate pests of crops, which include both insects and other arthropods, such as caterpillars, aphids, beetles, flies and mites, and molluscs, such as slugs and snails, can cause total crop losses if not treated. However, the negative effects of unregulated use of indiscriminate chemical pesticides on the environment, and human health, are well-documented. In the search for new environmentally-friendly
pesticides, many investigators have identified the possibility of using examples from the wide range of protein and peptide toxins produced naturally by organisms such as spiders, scorpions and other predators to control insect and other invertebrate pests. These toxins have many advantages - they are effective, can be highly specific in their action, with no effects on higher animals; they are also susceptible to biodegradation in the environment. However, delivery of these toxins to their sites of action - usually in the central nervous system - has remained a consistent problem which has prevented their adoption as pesticides; whereas a predatory organism can sting or bite its prey to introduce toxins into its circulatory system, delivery through contact or feeding by the target organism is usually ineffective.
Fusion protein technology enables the conversion of a protein or peptide toxin which normally requires access to the circulatory system of an organism to be effective, into a pesticide which is effective when orally delivered. This is achieved by fusion of the toxin to a protein "carrier", which is able to transport across the gut wall of the target organism into the circulatory system. The fusion protein, containing both the toxin and the carrier in a single polypeptide, is produced from a synthetic gene in a recombinant expression system, either for exogenous application or produced endogenously in the crop. When the target organism ingests material containing the fusion protein, the toxin component of the fusion is transported by the "carrier" into the circulatory system; it is now able to access its targets. Fusion proteins are thus effective pesticides in baits or sprays. Fusion protein technology has been developed to produce novel pesticides which are more specific towards target species, with minimal side effects on beneficial organisms, such as bees. Since fusion proteins are biodegradable, they do not persist in the environment, which is timely in view of current concerns about molluscicide residues in water supplies.
The proposed project aims to develop two prototype fusion proteins, which have been shown to have insecticidal activity towards caterpillars, beetles, flies and aphids, and molluscicidal activity towards slugs, to the point where a prototype biopesticide is ready for evaluation in a commercial setting, in collaboration with potential customers. This will involve optimising the production process, and carrying out formulation and product development. At the same time the necessary background testing for preparing data for pesticide registration will be accumulated; this will include evaluation of both efficacy towards target pests, and ecotoxicological and environmental studies. The project will also aim to develop further fusion protein biopesticides, to allow a portfolio of products suitable for different applications to be built up. It will also look at development of resistance towards fusion protein biopesticides in potential targets, to develop a strategy for managing use of these products to minimise loss of effectiveness; the availability of a range of fusion proteins with differing modes of action will also be a valuable resource in resistance management.

Technical Summary

The proposed project aims to develop two prototype fusion proteins, which have been shown to have insecticidal activity towards caterpillars, beetles, flies and aphids, and molluscicidal activity towards slugs, to the point where a prototype biopesticide is ready for evaluation in a commercial setting, in collaboration with potential customers. This will involve optimising the production process, and carrying out formulation and product development. At the same time the necessary background testing for preparing data for pesticide registration will be accumulated; this will include evaluation of both efficacy towards target pests, and ecotoxicological and environmental studies. The project will also aim to develop further fusion protein biopesticides, to allow a portfolio of products suitable for different applications to be built up. It will also look at development of resistance towards fusion protein biopesticides in potential targets, to develop a strategy for managing use of these products to minimise loss of effectiveness; the availability of a range of fusion proteins with differing modes of action will also be a valuable resource in resistance management.

Planned Impact

Besides the anticipated commercial benefit to the industrial partner Lonza, in bringing a new class of biopesticides to market and developing a range of products for different applications in agriculture and horticulture, the beneficiaries of the proposed programme will include:
1. Farmers and growers in the agricultural and horticultural sectors, through the availability of new pesticides with improved efficacy, which can substitute for pesticides being withdrawn due to negative environmental effects.
2. Consumers, through reduced exposure to pesticide residues as a result of replacement of pesticides which are persistent in the environment, and difficult to remove, such as the molluscicide metaldehyde in water supplies, with biodegradable biopesticides.
3. The wider public, through improvements in biodiversity and environmental quality resulting from reduction in effects of pesticides on non-target organisms, as a consequence of the introduction of biopesticides which do not show harmful effects on non-target organisms.
4. The UK as a whole, as a result of the economic benefits gained by developing better strategies for crop protection and pest management, and by developing technology that can be used as a basis for a range of future pesticidal products.
Withdrawal of pesticides under regulations such as EC directive 91/414, and the more recent ban on the use of
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neonicotinoids to protect bees, has serious implications for the productivity of UK agriculture and horticulture, and consequently for economic costs for a range of products derived from these industries. New environmentally benign pesticides are needed to meet these growing gaps in product ranges. The proposed programme attempts to address some of these potential increased costs in agricultural and horticultural production, and thus represents a positive economic advantage; the commercial benefits resulting from the programme envisaged by the industrial partner Lonza are based on the underlying economic benefits to society as a whole. These benefits will start to be realised in the medium term (i.e. approx. 5 years after project start) as products incorporating fusion protein biopesticides start to become available to farmers and growers.
Benefits to the broader research community are described under "Academic Beneficiaries" above. Benefits to the participants in the programme will include professional development, principally in the area of working with industrial partners and being involved in bringing an idea from the laboratory to the marketplace, a progression where UK science has been found wanting. Expertise in areas such as large-scale recombinant protein production, and carrying out efficacy and toxicology assays involving field studies will provide additional advantages.
 
Description The main challenges addressed by the project were as follows:
(1) Translation of laboratory based process (using commercially available vectors and expression host) for the production of recombinant prototype fusion protein to the lead partners pilot production process using "in house" expression vectors and expression host. This was succesfully achieved although further optimisation to enhance expression levels is desirable. In addition a downstream processing process suitable for commercial scale production was developed by the industry lead.
(2) Proof of concept to demonstrate that the commercially produced prototype has comparable biopesticidal efficacy to protein produced by the academic partners (Durham/Fera) was also demonstrated in slug ingestion trials and Colorado potato beetle glasshouse trials conducted by i2L.
(3) Preliminary ecotox studies suggested that the prototype fusion protein would have minimal toxicity towards non-target pests. Further studies are required for regulatory approval. Similarly preliminary environmental fate studies suggest that fusion proteins are prone to fairly rapid (2 days) degradation in soils. Whilst experimental methods gave qualitative results further development is required to enable quantitative assessment of fusion perotein degradation/persistence in the soil environment.
(4) Further extension of the range of target pests was demonstrated; a variant of the protype fusion protein under development was shown to have good efficacy against small hive beetle, a serious pest of honeybees.
(5) Research to develop follow-on candidates has provided proof of concept for a new approach (through protein engineering) that has potential for the production of further fusion protein prototypes with enhanced efficacy as compared to the current prototype under development.
(6) Whilst preliminary studies showed proof of concept for the use of Lonza produced fusion protein to control Colorado potato beetle, the level of expression achieved was too low to enable appropriate bait formulation for efficacy against slugs to be determined.
Exploitation Route Exploitation as commercial biopesticides - development in progress.
There has been a change in emphasis away from the initial wish of the industrial partner to use fusion proteins biopesticides as a molluscicide targeting slugs, to a more general use as an insecticide. Issues encountered with bait formulation, together with preliminary evidence for contact activity have driven a change in focus for Lonza who now wish to place more focus, in the short term, on the opportunity to explore further insecticidal applications of this platform technology.
Sectors Agriculture, Food and Drink

 
Description Lonza 
Organisation Lonza Group
Country Global 
Sector Private 
PI Contribution Proof of concept research
Collaborator Contribution Pump-priming funding
Impact Results confidential
Start Year 2011
 
Title Spider toxin 
Description Improved production methods for recombinant spider toxins. 
IP Reference WO2012131302 
Protection Patent granted
Year Protection Granted 2017
Licensed No
Impact Protection of IP for spider toxin fusion protein programme.