Impacts of the cultivation of Diabrotica-resistant Bt maize on the maize ecosystem

(2005 - 2008) RWTH Aachen University, Institute of Environmental Research (Biology V) Chair of Ecology, Ecotoxicology and Ecochemistry

Topic

The aim of the project is to compare the impact of Bt maize, which is resistant to the European corn borer, and conventional maize on various organisms in the maize ecosystem.

Key research points are the effects of the Bt protein (Cry3Bb1) on organisms living in the soil which are involved in breaking down the maize plant and are therefore exposed to the Bt toxin.

Experiment description

A total of 32 plots are arranged on the trial field. Eight plots are planted with Bt maize (MON88017), eight with an isogenic lin e and eight each with two other conventional varieties. The plots with the different varieties are distributed at random, but in compliance with two conditions: no two adjoining plots contain the same variety and there must be the same number of plots of each variety at the edge of the field and in the middle. Bt maize is grown on the same plots in all years so that any accumulation effects in the soil can be observed.

Effects of Bt maize cultivation on soil-dwelling organisms

Maize litter is decomposed directly by wood mites and earthworms among other organisms. These primary decomposers are in turn eaten by other organisms like springtails and mites.

The decomposition process is studied both in the field and in mesocosms.

Bait strips with maize litter for measuring the feeding activity of the soil fauna at different soil depths

Mesocosms as field model systems for investigating the degradation rates of different maize varieties

Field

Feeding activity: To determine the feeding activity of the soil fauna, bait strip tests are carried out after the harvest. A bait substance is inserted into the bait strips and placed in the soil. Two variants are used:

Feeding activity on one bait type: 16 bait strips (each with 16 holes) are placed in each plot of each maize variety. The strips are baited with isogenic maize and cellulose.
Feeding activity on different bait types: 16 bait strips containing the different maize variants (transgenic, isogenic and two other conventional varieties, each with cellulose) are placed in each of the isogenic maize plots.

Species spectrum: The species spectrum of the soil fauna in the field is determined by extracting ‘soil cores’ (diameter 5 cm, depth 5 cm). Five soil cores are taken from each plot of the trial field (=160 samples).

The organisms are extracted from the samples and sorted into groups, such as wood mites and springtails.

Mesocosms

In line with the field experiments, in 2005 and 2006 a comparative study was made of the feeding activity of the soil fauna in mesocosms. In August 2005 32 soil samples (diameter 30 cm, height 40 cm) were taken from a field close to the trial field. In the experiment, 76 grams of chopped maize litter of each of the different varieties were mixed into the top ten centimetres. Eight bait strips (bait substance: isogenic maize litter + cellulose) were placed in each mesocosm. From September to December, and in April and May of the following year, one bait strip was examined each month in terms of feeding activity and species spectrum.

Impacts on epigeous (above-ground) predatory arthropods

Ground beetles (Carabidae) and spiders (Araneae) are important predators in the species community of a maize field.

Activity densities: To measure the activity density of predatory arthropods in Bt maize compared with the conventional varieties, a pit trap was placed in each plot. The traps were emptied once a week from July 2005 (11 times). The samples were sorted into animal groups and identified down to species level.

ELISA: At the same time, live traps were set in order to study the ground beetles for Cry3Bb1 using the ELISA method. The ground beetles could have ingested the Bt protein via their prey (e.g. springtails, mites, aphids), or from plant food sources such as maize litter.

On three dates in August 2006 ground beetles that had been caught alive were frozen at –50 degrees Celsius. These beetles were later tested for their Cry3Bb1 content.

Biotest: In complementary biotests in the laboratory, ground beetles were fed larvae from the European corn borer. The ground beetles were given the choice between corn borers that had previously been fed on Bt maize and corn borers that had been fed on isogenic maize (choice trials). In additional feeding experiments, the four most common species (Calathus fuscipes, Pseudophonus rufipes, Calathus ambiguus and Pterostichus melanarius) were fed with the Bt protein, either directly or indirectly.

Results

Diag.1 Evaluation of the bait strip experiments showing the number of holes eaten (averaged out) for 16 bait strips per plot. The bait was isogenic maize + cellulose.

Diag.2 Evaluation of bait strip experiments, showing the number of holes eaten (averaged out) for 16 bait strips per bait type. The bait strips were placed in eight isogenic plots.

Diag.3 Numbers of springtails and predatory mites per m2 in the different maize variants 2005

Diag.4 Numbers of wood mites per m2 in the different maize variants 2005

Diag.5 Comparison of year-begin figures of epigeous predatory arthropods in the different maize variants (with standard deviations)

Impacts of Bt maize cultivation on soil-dwelling organisms

Field experiment

Feeding activity: Feeding activity on one bait type: No differences were found when comparing the feeding activity in the different maize variants in 2005 and 2006 (see Diag. 1).

Feeding activity on different bait types: A comparison of the feeding activity on different bait types found significant differences in both years (see Diag. 2). The Bt maize and the conventional variety DK315 were eaten significantly less in 2005 than the isogenic maize and the conventional variety Benicia. In 2006 there was no difference between the transgenic maize and the isogenic variety, but both were eaten significantly less than DK315 and Benicia.

Species spectrum: A correlation was found in 2005 between feeding activity and the number of wood mites (Oribatidae). By contrast, no correlation was found between feeding activity and the presence of springtails (collembolans). No significant differences between the different maize variants were found in 2005, either in terms of wood mites or springtails and predatory mites.

The time-consuming analyses for 2006 are not yet complete.

Mesocosms

Feeding activity: No significant differences in feeding activity were found between the variants in either year.

Species spectrum: In 2005 there was no significant difference in the numbers of wood mites and springtails before the maize litter was placed in the mesocosms. Once the maize litter had been added, the number of springtails in the isogenic variety was significantly different from all other varieties studied, including the Bt maize. The number of wood mites in the Bt maize was significantly different from the numbers in the conventional varieties. The isogenic variety was not significantly different from the other varieties.

The samples from 2006 are still being assessed.

Effects on epigeous (above-ground) predatory arthropods

Activity densities: The dominant species among the ground beetles was found to be the Calathus fuscipes (52 per cent of ground beetles found), but the Pseudoophones rufipes and Pterostichus melanarius were also common. A total of 3660 ground beetles of 33 different species were found.

Here too, the data point to differences between the conventional maize varieties on the one hand, and the Bt maize variety and isogenic maize variety on the other. So far, no differences have been found between the transgenic and isogenic varieties. The greatest differences can be attributed to soil conditions.

The assessment of the activity densities of spiders is not yet complete.

ELISA: In initial ELISA tests in 2005, the Bt protein was found in approx. 28 per cent of ground beetles from Bt maize plots. The food chain investigation found on average seven micrograms of Bt protein per gram in the fresh Bt maize leaves, over two micrograms per gram in the corn borer larvae, and 0.2 micrograms per gram in the ground beetles.

Biotest: The biotests did not find any preference for one maize variety over the others. Furthermore, the Bt protein was not shown to have any effect on ground beetles.