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Synthesis/Regeneration 18   (Winter 1999)

Ecological Balance and Biological Integrity

Good Intentions and Engineering Organisms that Kill Wheat

by Elaine Ingham, Oregon State University

A genetically engineered Klebsiella-planticola had devastating effects on wheat plants while in the same kind of units, same incubator, the parent bacteria did not result in the death of the wheat plants.

Consider that the parent species of bacteria grows in the root systems of every plant that has been assessed for Klebsiella's presence. The bacterium also grows on and decomposes plant litter material. It is a very common soil organism. It is a fairly aggressive soil organism that lives on exudates produced by the roots of every plant that grows in soil. This bacterium was chosen for those very reasons to be engineered: aggressive growth on plant residues.

Field burning of plant residues to prevent disease is a serious cause of air pollution throughout the US. In Oregon, people have been killed because the cloud from burning fields drifted across the highways and caused massive multi-car crashes. A different way was needed to get rid of crop residues. If we had an organism that could decompose the plant material and produce alcohol from it; then we'd have a win-win situation. A sellable product and get rid of plant residues without burning. We could add it to gasoline. We could cook with it. We could drink grass wine-although whether that would taste very good is anyone's guess. Regardless, there are many uses for alcohol.

So, genes were taken out of another bacterium, and put into Klebsiella-planticola in the right place to result in alcohol production. Once that was done, the plan was to rake the plant residue from the fields, gather it into containers, and allow it to be decomposed by Klebsiella-planticola. But, Klebsiella would produce alcohol, which it normally does not do. The alcohol production would be performed in a bucket in the barn. But what would you do with the sludge left at the bottom of the bucket once the plant material was decomposed? Think about a wine barrel or beer barrel after the wine or beer has been produced? There is a good thick layer of sludge left at the bottom. After Klebsiella-planticola has decomposed plant material, the sludge left at the bottom would be high in nitrogen and phosphorus and sulfur and magnesium and calcium-all of those materials that make a perfectly wonderful fertilizer. This material could be spread as a fertilizer then, and there wouldn't be a waste product in this system at all. A win-win-win situation.

But my colleagues and I asked the question: What is the effect of the sludge when put on fields? Would it contain live Klebsiella-planticola engineered to produce alcohol? Yes, it would. Once the sludge was spread it onto fields in the form of fertilizer, would the Klebsiella-planticola get into root systems? Would it have an effect on ecological balance; on the biological integrity of the ecosystem; or on the agricultural soil that the fertilizer would be spread on?

Would it have an effect on ecological balance; on the biological integrity of the ecosystem; or on the agricultural soil that the fertilizer would be spread on?

One of the experiments that Michael Holmes did for his Ph.D. work was to bring typical agricultural soil into the lab, sieve it so it was nice and uniform, and place it in small containers. We tested it to make sure it had not lost any of the typical soil organisms, and indeed, we found a very typical soil food web present in the soil. We divided up the soil into pint-size Mason jars, added a sterile wheat seedling in every jar, and made certain that each jar was the same as all the jars.

Into a third of the jars we just added water. Into another third of the jars, the not-engineered Klebsiella-planticola, the parent organism, was added. Into a final third of the jars, the genetically engineered microorganism was added.

The wheat plants grew quite well in the Mason jars in the laboratory incubator, until about a week after we started the experiment. We came into the laboratory one morning, opened up the incubator and went, "Oh my God, some of the plants are dead. What's gone wrong? What did we do wrong?" We started removing the Mason jars from the incubator. When we were done splitting up the Mason jars, we found that every one of the genetically engineered plants in the Mason jars was dead. Wheat with the parent bacterium, the normal bacterium, was alive and growing well. Wheat plants in the water-only treatment were alive and growing well.

...we found that every one of the genetically engineered plants in the Mason jars was dead.

From that experiment, we might suspect that there's a problem with this genetically engineered microorganism. The logical extrapolation from this experiment is to suggest that it is possible to make a genetically engineered microorganism that would kill all terrestrial plants. Since Klebsiella-planticola is in the root system of all terrestrial plants, presumably all terrestrial plants would be at risk.

So what does Klebsiella-planticola do in root systems? The parent bacterium makes a slime layer that helps it stick to the plant's roots. The engineered bacterium makes about 17 parts per million alcohol. What is the level of alcohol that is toxic to roots? About one part per million. The engineered bacterium makes the plants drunk, and kills them.

But I am not trying to say that all genetically engineered organisms are technological terrors. What I am saying is that we have to test each and every genetically engineered organism and make sure that it really does not have unexpected, unpredicted effects.

They have to be tested in something that approximates a real world situation. I've worked with folks in the Environmental Protection Agency (EPA) and I know the tests the EPA performs on organisms. They often begin their tests with "sterile soil." But if it's sterile, then it's not really soil. Soil implies living organisms present. If you use "sterile soil" and add a genetically engineered organism to that sterile material, are you likely to see the effects of that organism on the way nutrients are cycled, or on the other organisms in that system? No, you're not likely to. So it it's probably no surprise that no ecological effects are found when they test genetically engineered organisms in sterile soil. They really need to put together testing systems, which assess the effects of the test organism on all of the organisms present in soil

...is to suggest that it is possible to make a genetically engineered microorganism that would kill all terrestrial plants.

What do we mean, organism-wise, when we talk about soil? Agricultural soil should have 600 million bacteria in a teaspoon. There should be approximately three miles of fungal hyphae in a teaspoon of soil. There should be 10,000 protozoa and 20 to 30 beneficial nematodes in a teaspoon of soil. No root-feeding nematodes. If there are root feeding nematodes, that's an indicator of a sick soil.

There should be roughly 200,000 microarthropods in a square meter of soil to a 10-inch depth. All these organisms should be there in a healthy soil. If those conditions are present in an agricultural soil, there will be adequate disease suppression so that it is not necessary to apply fungicides, bactericides, or nematicides. There should be 40 to 80% of the root system of the plants colonized by mycorrhizal fungi, which will protect those roots against disease.

What happens when you apply the most fungicides and pesticides to soil? In every single case where we have looked at foodweb effects of pesticides, there are non-target organism effects, and usually very detrimental effects. The sets of beneficial organisms that suppress disease are reduced. Organisms that cycle nitrogen from plant-not-available forms into plant-available forms are killed. Organisms that retain nitrogen, phosphorus, sulfur, magnesium, calcium, etc. are killed. Organisms that retain nutrients in the soil are killed. Once retention is destroyed, where do those nutrients go? They end up in our drinking water; or end up in our ground water. You and I as taxpayers have to pay in order to clean up that water so we can drink it.

Wouldn't it be much wiser to keep those organisms present in the soil so those nutrients would be retained and become available to the next crop of plants instead of ending up in our drinking water where we have to pay in order to have clean drinking water? How do you do that? You get the organisms back into the soil. If you grow the proper number and types of bacteria, fungi, protozoa, nematodes and microarthropods, mycorrhizal fungi in the root systems of the plants, you can do away with pesticides. It's been done. We can reduce significantly the amount of fertilizer that goes into that soil. In experiments that have been done all over the country, all over the world, inorganic fertilizer inputs have been reduced, or are not added at all, without reduction in plant growth. Where green manure or legumes are not available, approximately 40 pounds of nitrogen fertilizer, once every four years, are still necessary.

Let's talk about why today's conventional agricultural systems require such massive inputs of pesticides and fertilizers. When a healthy soil is first plowed out of native grassland, for example, the disease-suppressive bacteria and fungi, protozoa and nematodes are present. For the first 5 to 15 years after plowing native grassland you don't have to use any pesticides. No fertilizers are required because there is natural nutrient cycling, natural nitrogen retention, and disease suppression. As you plow that soil, you start to kill the beneficial organisms, you lose the organic matter, and you lose the food to feed the beneficial organisms. After about 10 to 15 years, if you're not adding back adequate plant residue to feed those organisms, you lose them, and start having significant disease problems. Then you either leave that land and farm elsewhere, or in the US, we used fertilizers to keep yields high. As more and more of the organisms were killed by the salt effect of the fertilizers, and the constant plowing mined out more and more of the organic matter, starving the beneficial organisms to death, disease became a serious problem. And we started using more and more pesticide to knock the disease back.

In California, around 1955, those disease problems became so severe that they thought they would lose agricultural production. So the University of California came up with a better way to kill those disease-causing organisms. It's called methyl bromide. This chemical kills disease-causing organisms-but it also kills everything else. There is very little natural disease suppression going on in agricultural soils in California.

How many organisms are left in strawberry fields that have been methyl-bromided 2 to 3 times a year for the last 14 years? There are no microarthropods left. There are no beneficial nematodes left; only root feeding nematodes. And there is nobody to control root-feeding nematodes in those soils. How many protozoa are left in that soil? None. You cannot cycle nutrients. There is nobody home to make nitrogen plant-available. So what do you have to do? You have to add fertilizer. We force ourselves to have to add fertilizer. We have no other choice if you're going to grow those plants in those soils.

How many fungi do you have left in that soil? No beneficial fungi-they're all disease-causing. How many bacteria are left? All are gone, except for 100 per gram of soil. We should have 600 million per teaspoon in that soil; we have 100 left. There is nothing left to retain nitrogen in those soils, nothing. So you apply fertilizer. What happens to the fertilizer? Whatever fertilizer contacts the roots of the plants is indeed taken up; the rest of it flushes through the soil into the ground water, into the river. Take Santa Maria River in California as an example. This land has had methyl bromide applied 2 to 3 times a year for the last 14 years or more. Fertilizer is applied as sidedress when strawberries are planted. About two weeks later, the river goes up to around 150 parts per million nitrates. What is the toxic level for nitrate for humans? Ten parts per million nitrates is what the EPA tells us. It used to be three parts million but that level was increased. Can you drink that water in the river in the Santa Maria valley? Not unless you'd want to die. You would destroy your kidneys pretty fast if you drank that water. It is high in nitrate. It is so toxic that you can't even put that water back on the plants. The high nitrate burns the plants.

We have a simple solution for this problem. Get the right kind of organisms, the right numbers of organisms, back in the soil and let them start performing their functions again. Put food for the organisms back into the soil; put the organisms back into the soil. It's that simple. Send us your soil samples and we can tell you whether you have that food web in your soil.

How are you going to fix that set of organisms it if you don't have a healthy foodweb? We can help you with that question. We can indeed move towards that time when we really don't need pesticides anymore; where you only apply fertilizer once every four years and in very small amounts. We can move to a sustainable agriculture. It takes time and effort, but it is possible.

This article is adapted from the presentation the author gave on July 18, 1998 at the First Grassroots Gathering on Biodevastation: Genetic Engineering.

See also: Holmes, M.T., Ingham, E.R., Doyle, J.D., & Hendricks, C.W. (1998). Effects of Klebsiella-planticola SDF20 on soil biota and wheat growth in sandy soil. Applied Soil Ecology, 326, 1-12.

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