Science
Nitrogen fixation research advances Grasses such as rice, corn, and wheat plant's photosynthesizing system for are among the most important food their own energy needs. Thus, the plants that acquire fixed plant actually provides the energy crops. Reports that certain forage grasses that the bacteria use to break nitronitrogen from soil bacteria and maize could fix limited amounts gen's triple bond and make nitrogen of nitrogen have been made for about available for the plant to use in makand to improve efficiency of five years, van Berkum points out. ing proteins. This symbiotic association has Some of the most detailed early process start to bear fruit studies came from Brazilian workers been particularly well studied in the Johanna Dobereiner and John M. case of soybeans and the bacteria Day, who reported in 1975 experi- Rhizobium japonicum. Not all R. Agricultural researchers are making ments in which grasses fixed nitrogen japonicum strains are equally effisome headway in efforts to expand at rates of up to 2 kg of nitrogen per cient at fixing nitrogen, and one goal the range of plants that can utilize hectare per day. The bacterium of such research is to find ways to asatmospheric nitrogen and thus grow thought to be responsible for the sociate the most efficient bacteria with agriculturally important soylipoferum. without the aid of nitrogen fertil- fixation was Spirillum "There's no question that the levels bean strains. izer. Since the early 1900's, soybean Ordinarily plants require nitrogen of nitrogen fixation first reported for in a reduced form, usually as ammo- these grasses are too high," van Ber- seeds have been treated with nitronia, for protein synthesis. In fact, kum says. He points to problems in gen-fixing bacteria before they are certain strains of bacteria and algae the method used to measure fixed planted to be sure there will be plenty seem to be the only biological systems nitrogen that resulted in these in- of these bacteria in the soil to satisfy the plant's nitrogen requirements. that are able to convert atmospheric flated values. nitrogen to the reduced form that Nevertheless, there are enough However, researchers are just now plants can use. "Nitrogen-fixing" careful studies of nitrogen withdrawal developing techniques to study the bacteria are anaerobic and usually from grass croplands that receive no interaction between these introduced live in the soil either independently or external nitrogen fertilizer to make it bacteria and the soybean plants. in association with the roots of certain clear that bacteria associated with And the results are surprising. USDA microbiologist L. David plants, particularly legumes. these grasses are fixing nitrogen in the Thus, research to improve nitrogen soil, van Berkum says. In tropical Kuykendall of the Cell Culture & fixation for plants takes two direc- environments, as much as 100 kg of tions. In one, researchers use recom- reduced nitrogen per hectare is probinant DNA techniques to attempt to duced annually in the soil where these transfer the nitrogen-reducing ability grasses grow. of certain bacteria into plant cells. So In temperate zones, one of the best far, this transfer has been achieved nitrogen-fixing grasses is salt-marsh successfully from a bacterium to cordgrass, Spartina alterniflora, yeast, the first step toward transfer- which has a Spirillum bacterium asring such genes to higher plants sociated with it that can fix about 70 (C&EN, Sept. 29, page 35). kg of nitrogen per hectare per year. The other direction of research Rice and * other marshland grasses uses more classical plant breeding that are transplanted into marsh soils techniques and biochemistry to make of the type where Spartina alterninaturally occurring associations be- flora flourishes also develop small tween nitrogen-fixing bacteria and amounts of nitrogen-fixing activity, plants that utilize the nitrogen more van Berkum finds. useful for the plants. Here, too, reAlthough the evidence is still search is yielding promising find- spotty, van Berkum suggests that the ings. Spirillum bacterium that fixes niFor instance, the range of plants trogen for these grasses is a free-living that acquire fixed nitrogen from soil soil bacterium that is somehow stimbacteria is almost certainly greater ulated by the presence of the grass than had been thought as recently as plant. The bacteria that fix nitrogen a decade ago. Of particular interest in for legumes behave differently. These crop production is the growing evi- bacteria, of the family Rhizobium, dence that this range includes at least irritate the root hairs of the develsome grasses, says Peter van Berkum, oping legume plant, causing the roots a biochemist at the University of to develop nodules in which the bacMaryland who works at the Depart- teria live. The vascular system of the USDA's Thomas Devine and Barbara ment of Agriculture's Beltsville, Md., plant is tied directly to these nodules, Breithaupt examine soybean plants grown Agricultural Research Center. so that the bacteria can tap the in nitrogen-free medium
Efforts to increase range of
Dec. 8, 1980 C&EN
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Nitrogen Fixation Laboratory, Beltsville, Md., has developed a method that genetically labels the bacteria introduced with the soy beans. He finds that certain strains are preferentially picked up by the growing roots of the soybean. Simply increasing the amount of bacteria introduced into the soil along with the soybean seeds does not increase the uptake of the bacterium by the plant. In fact, when specially developed, highly efficient strains of Rhizobium are introduced with the soybeans, only about 20% of the plants actually incorporate the improved strain into their root nodules, Kuykendall finds. The rest pick up instead one of the nitrogen-fixing bacteria already present in the soil. Thus, there are factors more com plicated than mere concentration that influence competition between Rhi zobium strains for pickup by the soybean roots. And the soybean does not necessarily take up the bacterial strain that is most efficient at fixing nitrogen. Although Kuykendall has few clues, so far, as to what the com petitive factors are, he is ready to try to take advantage of them. "I'm sure you can breed Rhizobium for com petitive advantage," he says. Tackling the same problem from
the soybean end, Thomas E. Devine, a plant geneticist at USDA's Belts ville lab, is trying to develop a soy bean strain that will not associate with indigeneous soil Rhizobium but can be infected with selected strains. Devine begins with a mutant soy bean that doesn't ordinarily produce bacteria-containing nodules. This soybean contains a gene for nonnodulation called rji. In 1977, Devine planted rj χ-containing soybeans in a field known to contain many strains of Rhizobium and examined the re sulting plants for nodule formation. He found 34 plants, out of 30,000, that had developed some nodules. These nodules were removed and the bacteria isolated from them. These bacteria, Devine reasoned, have the ability to overcome the plant's resistance to bacterial infec tion. He infected young rjx-containing plants with the bacteria isolated from the nodules and found this strain does produce nodules. When grown in sand culture, these plants were able to fix enough nitrogen to grow into healthy plants without external ni trogen fertilizer, but in field studies they did not do so well. More work will be needed to get adequate ac ceptance of selected strains. Rebecca Rawls, Washington
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C&ENDec. 8, 1980
NIH recombinant-DNA guidelines revised The National Institutes of Health Guidelines for Research Involving Recombinant DNA Molecules have been revised again, and already a clamor has been raised to revise them further. The major change in the guidelines concerns the role NIH takes in su pervising recombinant DNA research throughout the U.S. In particular, the change shifts much of that supervi sory responsibility away from NIH to local biological safety committees at institutions where such research is being conducted. Central review by NIH is "unnec essary, inhibitory, expensive, and counterproductive," said NIH scien tist Maxine Singer in proposing the current changes. "Confidence in the efficacy of the guidelines has been based primarily on compliance by individual investigators within their own laboratories/' The new changes streamline procedures by putting more responsibility on local com mittees, eliminating the need for sci entists to submit plans of their ex periments to NIH for evaluation. "Seven years have passed since the scientific community first raised questions regarding recombinant DNA experiments," Singer says. "The ignorance of those early years has been supplanted with a wealth of experience and information. Just as the early stringent guidelines were promptly adopted in response to the ignorance, we must now respond just as promptly to the current more re alistic appraisals." Representatives of biosafety com mittees across the U.S. are calling for still further changes in the guidelines. They voiced their wishes during a two-day conference sponsored by NIH and held recently in Washing ton, D.C. For example, committee repre sentatives question whether routine physical exams of scientists, students, and technicians doing recombinant DNA research serve a useful purpose. Representatives argued that current general health surveillance probably would not indicate any "subtle" or "low-level" health risks, if they exist, in doing such research. Thus it was recommended that "specific ques tions" must be posed and "specific data must be collected" to be able to evaluate any such health risks. Un fortunately, the committee members seemed unable to define just what kind of "specific" data ought to be sought. But far bolder issues are at stake. Many representatives now say that
