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Biocontrol of important soil dwelling pests by improving the efficacy of insect pathogenic fungi

Contract nr: FAIR-CT98-4105
Project nr: 4105
Project type: SC
Starting date: 01/02/1999
Duration: 36 months
Total cost: 2,443,297 EUR
EC Contribution: 1,574,000 EUR
Scientific Officer: Massimo BURIONI
Research topic: Plant health
Acronym: BIPESCO

Larvae of scarab beetles and weevils feed on the roots of a wide range of economically important plants such as soft and top fruit (e.g. vines, apples, raspberries, blackcurrants, strawberries), ornamentals (e.g. cyclarnen, nursery stock), pasture, arable crops (e.g. cereals, potatoes), and forest trees (e.g. firs, Christmas trees). The damage throughout Europe is estimated at several billion euros. Current control measures are dependent on toxic chemical pesticides some of which are ineffective in the soil, while others have been - or will soon be - banned (e.g. methylbromide). Fungal pathogens are endemic in pest populations and efforts are being made to develop these for pest control. There is, however, a need to develop economical methods for the production of large quantities of stable, virulent microbial pesticides, and appropriate targeting strategies to:
1) maximise the impact of these agents for pest control in different soil habitats:
2) minimise the impact on the environment.

This project set out to develop and to study environmentally friendly biological control agents which should replace or reduce the input of chemical pesticides in European agriculture, forestry and horticulture. This is in accord with the reformed Common Agriculture Policy (CAP) guidelines and is the objective of area four of the FAIR programme. The aims of this project will be to:
1) control scarabs and weevils with virulent, ecologically competent strains of insect-pathogenic fungi;
2) improve production and formulation technologies;
3) develop biochemical methods to monitor fungal virulence and nutritional (carbon) requirements;
4) use molecular techniques to characterise strains to monitor the pathogen in the field (spatial-temporal distribution, genetic stability, interactions with autochthonous, conspecific strains) and for the detection of instability factors (e.g. transposons, mycoviruses);
5) test new application systems for effective targeting of the pathogen;
6) study the impact of the pathogens on target and non-target insects;
7) conduct field trials to demonstrate/evaluate the efficacy of the fungal biological control agents;
8) address some of the criteria for the registration of insect pathogenic fungi.

These goals will be achieved through the use of new formulated fungal biopreparations. The fungal pesticides have to be evaluated for their environmental benefits and efficacy, to demonstrate that these promising products will be an alternative to chemical insecticides. This project links partners with different - but complementary - expertise from several countries, both from research institutions and SMEs. It should accelerate the progress in achieving the proposed objectives. Successful implementation of the expected results should substantially reduce the need for chemical control of soil-dwelling pests like scarabs and weevils. Most of the results from this programme have universal applicability and could be utilised in both field and protected crop ecosystems.
Key words: biocontrol agent, insect pathogenic fungi, scarabs, weevils, soil pests, virulence, biotechnology, formulation, ecological fitness, sustainable agriculture, organic farming.

Current situation/results:
The first milestone - establishing protocols to maintain and assay selected pest insects - has now been accomplished. The BIPESCO-team has defined appropriate methods for conducting assays against target and non-target pests and rearing of selected test insects. This has enabled the partners to evaluate, on the basis of the dose-mortality and time-dose response analyses, the virulence of their respective isolates. Furthermore, standardised bioassays were conducted in soil habitats to evaluate:
1) the ecological fitness of the fungal BCAs;
2) the effect of edaphic factors (soil type, moisture, temperature) on the efficacy of introduced pathogens. Eight virulent, ecological fit Beauveria and Metahrizium strains have been provided to each partner, to compare specific isolates with the reference strains already characterised.
The industrial partners will continue liquid and solid state fermentation studies to prepare the foundation for the scale-up of the production process. Particular effort will be concentrated on the development of liquid and granular formulations which enhance shelf life, sporulation and growth after application.
The sequencing of the entire rDNA gene complex of M. anisopliae was set as a target and completed. Currently, the sequencing of parts of functional genes of M. anisopliae mtDNA has been initiated. It is expected that within the first quarter of 2000, a full screening of all isolates will take place. Emphasis will be given to the sequencing of rDNA variable regions from our best isolates. A few of the primer amplicons have been cloned, and the others will be cloned within the next few months. They will then be sequenced, in order to provide homologous sequences for mtDNA gene amplification. Further implementation and development of molecular methods is scheduled, to characterise M. anisopliae, enabling the separation of released strains from indigenous strains. Additional primer pairs will be included in the studies. Furthermore, a larger collection of different Beauveria isolates will be tested with SSR-markers to assess the variability at the various microsatellite loci.
The evaluation of the survival and distribution of B. brongniartii at sites where the fungus was applied periodically, will be intensified in the second year, using additional test sites in Italy, Austria, Germany, Denmark and Switzerland. Periodically, the sites will be evaluated for the quantity and virulence of B. brongniartii-spores, infection of Melolontha-larvae and detectable damage of crops. Further studies are planned to study the shelf life of the fungus in pest-free soil areas. This will contribute to a better understanding of the relationship between the fungal BCAs and pest densities.
The impact of entomogenous fungi on target and non-target insects and on the behaviour of pest insects in soil where M. anisopliae was applied have been initiated and will be continued.


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