Sep 22, 2006

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SiFo Project: The development of resistance by the Western corn rootworm to Bt maize through alternative host plants

What the pest likes to eat

Late August 2006. Figures clad in protective suits stand in front of a row of miniature tents teeming with life in a small room at the University of Göttingen. Strict safety measures are in force because a pest which does not occur at all yet in Germany is being bred here. Joachim Moeser and his team need eggs and larvae from the Western corn rootworm for their experiments. For a year now they have been observing how mobile the larvae are and what else they eat apart from maize.

In their project, Joachim Moeser and his colleagues are working on a distinctive feature of the Western corn rootworm (Diabrotica virgifera). Although the pest lives mainly on a diet of maize, it can also survive on other plants. “The theory is”, explains Joachim Moeser, “that cucurbits (squashes), not maize, were originally the primary food source of the Western corn rootworm. In Mexico, the homeland of maize, maize was grown alongside squashes and consequently the pest eventually specialised in maize. Even today the beetles are still drawn to the attractant given off by squashes, even though the squash is no longer a suitable fodder for the larvae.

“However, the fact that the Western corn rootworm can also survive on other plants could become a problem. Scientists suspect that this ability could favour the development of resistance to Bt maize. Bt maize, which is effective in controlling the Western corn rootworm, produces comparatively little Bt toxin; it mainly attacks the first larval stage (L1 larvae) of the pest. Subsequent larval stages (L2 and L3 larvae) are less susceptible. If L1 larvae were to feed on alternative host plants initially and only then migrate to Bt maize, this could increase the likelihood of survival in the Bt maize and with it the population of “partially resistant” beetles.

Beetle breeding

The researchers need the pest for their research. But since the Western corn rootworm is currently not found in Germany, it is classed as a quarantine pest, which means that special safety precautions must be taken and corresponding codes of behaviour must be adhered to.

This explains why there is a small anteroom outside the actual breeding room. To ensure that the two doors are never open at the same time, a warning light goes on outside when the door to the breeding room is open. As soon as the outer door is opened, the inner door locks magnetically.

The anteroom is lit by a UV lamp. Joachim Moeser takes protective suits out of a freezer. “All items from the breeding room, including rubbish, have to spend three weeks in here, before they can be removed”, he explains. The protective suits should prevent any escaping beetles from being carried out in clothing or hair. He switches the light off to prevent any insect flying to it, as the door now opens on a room containing several miniature tents teeming with beetles.

Each tent contains around 1,500 beetles. Joachim Moeser takes a few young maize leaves and pushes his hand through a sealed sleeve into the inner tent. The beetles are fed on pieces of maize plant and artificial feed, which, amongst other things, contains a pollen substitute also used by beekeepers. The insects for breeding were collected from infested areas in Austria and Italy. Eggs from American populations were also used. The main aim of the breeding programme is to produce eggs, which will be used in the trials.

Suitable plants

A bioassay test has been developed to find out which plants, if any, make suitable alternative host plants for the Western corn rootworm larvae. Each larva is placed in a glass test tube along with a piece of root from the plants that are being tested. The bottom half is filled with plaster, which acts as a hydrator and the other half with vermiculite, an aluminium silicate which is used to aerate the soil. A selection of wild grasses and monocotyledonous cultivated grasses that can grow as volunteers in maize fields, such as winter wheat for example, were tested. After six days researchers checked to see how much the larvae had eaten and the extent to which they had been able to convert the feed into measurable body mass. Wild oat (Avena fatua), yellow foxtail (Setaria glauca), rye and winter wheat were the most suitable alternative fodder plants. These plants are now undergoing further tests.

Microhabitat in a box

There are several large 550 litre containers standing in a greenhouse, half filled with earth and planted up. Here, semi-field conditions are simulated in so-called microhabitats. Bt maize is growing alongside various alternative host plants. The trial is designed so that between the two outer rows of Bt maize, there is a row containing one of the different plant species being tested. At present yellow foxtail is growing in one container and in another winter wheat. The wheat and foxtail, which were sown four weeks ahead of the maize, as is customary, are taller than the maize at the moment, but it will soon catch up. When the maize is six weeks old, around 120 eggs per plant from the Diabrotica breeding programme will be placed in the soil between the rows. The larvae will hatch out and immediately feast upon the roots of the maize, foxtail or wheat. After twenty days the plants, together with their root balls, will be removed from the soil.

“The remaining soil must be decontaminated of course, i.e. any residual larvae must be killed off.” Joachim Moeser points to a piece of equipment on wheels, which basically consists of a large metal boiler. The soil is steamed inside it at a temperature of plus eighty degrees and can then be reused.

Further experiments are planned; trials with different plants (definitely rye and possibly wild oats) and also different set-ups, e.g. with the alternative host plants in among the Bt maize, instead of between the rows. There will also be containers with only Bt maize as a control.

Counting, measuring and weighing larvae

There is an ingenious device known as the Kempson extractor, which is used to remove the larvae from the root balls. The root balls are placed in containers with wide-mesh grid floors covered by a finer mesh. Similar containers with a solid floor and containing water are placed underneath. The two containers are placed together in the apparatus, where they are heated from above by lamps and cooled from below with cold water. As the soil dries out in the heat, the larvae migrate towards the cool damp atmosphere and then fall through the mesh. Then they are counted and the larval stage is determined based on the diameter of the head capsule. A maximum of ten larvae from each larval stage then undergo further tests. The total weight and the head capsule width of the larvae are measured. In this way it is possible to determine for each individual plant how many larvae have hatched out on it, how many have survived and how good a source of nourishment the plant is.