ON THE ROLE OF SOIL BACTERIA IN. N. KRASILNIKOV Institute of Microbiology of the USRR Academy of Sciences Received for publication, Sept.

Transcription

1 CONTRIBUTION J. Gen. V Appl. Microb;ol. ol. 7, No. 2, 1961 ON THE ROLE OF SOIL BACTERIA IN PLANT NUTRITION N. KRASILNIKOV Institute of Microbiology of the USRR Academy of Sciences Received for publication, Sept. 1, 1960 The part played by microorganisms in plant nutrition is one of the most important problems in soil microbiology. It has lately been established that microbes produce various biologically active substances which exert considerable influence on the growth and development of plants. The microbial metabolites evoke various reactions in the tissues depending on the specific properties and peculiarities of the plants. The metabolites of some species of microorganisms produce a stimulating effect; under their influence the plants grow faster and yield higher crops. These microbes are activators. Other species of microbes depress the growth of plants and their various functions. These organisms are inhibitors. The effect of either of these species may be quite sharply pronounced. Gibberelin and gibberelinlike substances may serve as an example of the strongly acting metabolites formed by the cultures of the first group of microbes. Many toxins and some antibiotics are known as strongly depressing substances synthesized by the other group of microbes. The soil contains many antagonistic microorganisms whose metabolites (antibiotics) produce a protective and curative effect and raise the immunobiological properties of plants (See KRASILNIKOV, 1944, 1952, 1954, 1958). When abundantly developing in the soil, numerous species of bacteria, actinomycetes, fungi and yeasts form various biologically active compoundsvitamins, auxins, substances of the gibberelin type, antibiotics, toxins, amino acids and many other growth substances. All these metabolites may be found in the soil. From the soil they pass into the plants through the roots and are distributed evenly or unevenly in the tissues (KRASILNIKOV and BEZZUBENKOVA, 1955). It has been established experimentally that feeding vitamins to plants increases the content of mineral and organic forms of nitrogen in the body fluid. (RATNER and DOBROKHOTOVA, 1956). A similar effect is produced by activating microbes (KRASILNIKOV, 1941, 1944, 1952, 1954; GEBHARDT, 1957; KoLosov and UKHINA, 1954, et at). Regrettably little attention is as yet devoted to this question in physiology and agrobiology. In the present report we are citing experimental data on the influence exerted by some soil bacteria on the synthesis and accumulation of amino acids and organic phosphorus containing compounds in plant tissues. 128

2 1961 On the Role of Soil Bacteria in Plant Nutrition 129 MATERIALS AND METHODS The experiments were performed under sterile, semi-sterile and nonsterile conditions. The sterile experiments were conducted in glass vessels, in an agarized medium, on sand or in sterilized soil. The non-sterile experiments were conducted in vegetational vessels filled with soil. The first series of experiments were conducted on the accumulation of amino acids in plants, the second series on the accumulation of organic phosphorus. The following vegetable objects were used : leguminous-beans, vetch, china, lucerne and blue lupine; non-leguminous-maize, barley and winter wheat. In the first series of experiments the plants were inoculated with rootnodule and non-root-nodule bacteria. The leguminous plants were inoculated with specific and non-specific root-nodule bacteria and with non-root-nodule bacteria. The grown plants were analyzed for total nitrogen and for the quantitative and qualitative composition of free amino acids (for the methods of analysis see KRASILNIKOV and ASEYEVA, 1959). Under field conditions the grown plants were removed from the ground and sorted into groups. The plants with many nodules on the roots, as well as those with no nodules on the roots were segregated and studied. Methods of radioactive tracers in combination with methods of chromatographic adsorption (see below) were used in the second series of experiments with phosphorus. RESULTS The data obtained by us in the experiments of the first series conducted with leguminous and non-leguminous plants grown under laboratory conditions and on experimental plots are shown in corresponding tables. Table 1 shows the indices of total nitrogen in the plants grown under conditions of sterile experiments in vessels with sand. These data show that the amount of total nitrogen increases in plant tissues in the presence of certain species of bacteria (activators), the overground parts of the plants becoming nitrogenized to a greater extent than does the root system. Different species of bacteria produce a different stimulating effect. The greatest acquisition of nitrogen by the plant tissues was observed in almost all the experiments using the culture of Ps. fluorescens. In the experiments with leguminous plants a great effect is observed from specific nodule bacteria cultures: in lucerne from Rh. meliloti, in clover from Rh. trifolii, and in beans from Rh, phaseoli. Azotobacter exerted no influence or but a weak influence on the accumulation of nitrogen in leguminous plants. Analogous results were obtained in experiments on open field plots. Table 2 shows the data obtained in the study of lupine and beans, both

3 130 N. KRASILNIKOV VOL. 7 Table 1. Total nitrogen content in the in sand under conditions of sterile the 20 days of growth). tissues of leguminous experiments (in % to plants grown dry weight on Table 2. Total nitrogen content in ments on field plots on 51 days to dry weight). the tissues of grown lupine lupine and beans in experiand 32 days grown beans (in with and without nodule on the roots. As was to be expected (see KRASILNIKOV and KoRENYAKO 1946), the plants with numerous nodules on their roots contain more total nitrogen than do the plants which have no nodules. As a rule, the nodules themselves contain more total nitrogen; in some cases (lupine) they contain per cent more than do the plant tissues. For example, in the nodules of the annual blue lupine we discovered in the 29th day of growth 4.950, in the 51st day 6.57% and in the 65th day 7.86% to the absolutely dry weight; in the vegetative mass of the same plants we discovered 3.00%, 5.07 o and 3.06% respectively. Analyses show that as the total nitrogen in plant tissues increases the

4 1961 On the Role of Soil Bacteria in Plant Nutrition 131 content of nitrogen in the amino acids also increases. Table 3 shows results of the analysis of beans and lucerne. The analysis of other leguminous plants-lupine, clover lucerne, etc., furnishes similar data. Table 3. Nitrogen of the free amino acids in the tissues of beans and lucerne. Experiments in sterilized sand. Grown for 32 days. (in % to dry weight) Table 3 shows that the specific active nodule bacteria produce a greater effect than do the non-specific nodule bacteria. Plants inoculated with certain active bacteria-ps. fluorescens, and some others-show that they contain high percentage of free amino acid nitrogen. The data of the field experiment are shown in Table 4. The tissues of beans and lupine were analyzed because it is possible to obtain these plants without nodules on their roots. A comparison of the figures in the Table shows that under field conditions, root-nodule bacteria form numerous nodules on the roots, the content of nitrogen in the free amino acids may also rise perceptibly. However, this is not always the case. Table 4. Nitrogen grown on of an the free amino acids in experimental field plot the (in tissues of beans and lupine to dry weight). both In one active series of experiments and inactive. As a we rule, inoculated lucerne with various strains, the active strains produced a more

5 132 N. KRASILNIKOV VOL. 7 perceptible increase in total nitrogen than did the inactive strains. Thus, inoculation with active strain IP-D showed 4.54% nitrogen, while inoculation with inactive strain 12-G showed 3.79 o in over-ground parts; the roots contained respectively 3.35% and 2.80% of the weight of the dry mass. The effectiveness of the bacteria manifests itself in different degrees at the different stages in the development of the plants. In one of the experiments we analyzed lupine plants at three stages of growth-during powerful vegetative growth (29th day), during flowering (51st day) and during fruiting (65th day). The results are shown in Table 5. The table shows that during the early growth the roots contain the maximum of the total nitrogen which then decreases as the growth proceeds. The over-ground parts contain somewhat more total nitrogen during flowering. During ripening, the nitrogen seems to flow to the over-ground organs. Table 5. Accumulation different stages of its of total growth nitrogen (in % to in the tissues dry weight). of lupine during the Field experiment The nitrogen of the free amino acids in the roots of the experimental plants which have numerous nodules increases with the age of the plants; 0.299% on the 29th day, 0.361% on the 51st day, and on the 65th day. The same fact was observed in the nodules: x, 0.657% and 0.786% respectively. In analyzing the roots and over-ground parts of plants we discovered by the method of paper chromatography 17 to 22 free amino acids in different quantitative relations. In Table 6 one may see the amino acids found in the tissues of blue lupine grown on the experimental field plot in the flowering phase. In the root of the control plants without nodules 18 free amino acids were found (2 of them were not identified) and in the roots of plants with nodules 19 amino acids with one unidentified. The nodules contained the greatest number of 21 amino acids of which 4 were not identified. Analogous data were furnished by the analyses of the tissues of other plants. The over-ground mass of beans (experimental plants) was found to contain 21 amino acids with 3 unidentified on the 32nd day of growth; the control plants had 19 amino acids with 3 unidentified; no proline or tryptophan were found.

6 1961 On the Role of Soil Bacteria in Plant Nutrition 133 Table 6. Composition of periment) depending dry weight). amino on the acids in the tissues of presence of nodules blue lupine on the roots (field (in exto In the overground parts of lucerne inoculated with root-nodule bacteria in a laboratory experiment 17 amino acids were found, while the control plants (not bacterized) had 16; proline and one unidentified amino acid were not found (Table 7). The scarcely active or inactive root-nodule bacteria produce a less perceptible effect or do not stimulate the processes of growth and of amino acid accumulation. The activity of root-nodule bacteria is of essential importance in these processes. The influence of the root-nodule bacteria manifests itself particularly well when the amino acids are determined quantitatively. The inoculated plants have perceptibly more amino acids than do the control plants. The

7 134 N. KRASILNIKOV VoL. 7 Table 7. Influence of active and inactive root-nodule bacteria on the accumulation of free amino acids in the tissues of lucerne. Experiment conducted under sterile conditions on 65 days growth (in % to dry weight). tissues of experimental plants accumulate 2 times or more quantities of such amino acids as aspartic acid, glycine, glutamic acid, threonine, alanine, etc., than do those of the control plants. The diagram in Fig. 1 shows the quantitative correlation of some of these amino acids obtained by us in analyzing the roots and green mass of blue lupine grown on the experimental field plot, at different periods of its growth. The diagram shows that the roots of inoculated plants accumulate large amounts (even more than do the over-ground parts) of aspartic acid and serine together with glycine; the control plants which do not have nodules on the roots contain hardly any of these compounds, except the early stage of growth when a notable amount of alaine and serine together with glycine are formd. Incidentally, the roots of control plants have very little of any other amino acids and some completely fail to be revealed by our methods of analysis. Root-nodule bacteria exert a perceptible influence on the accumulation of glutamic acid, threonine, alanine, tyrosine, tryptophan and certain other amino acids in the roots of the plants.

8 1961 On the Role of Soil Bacterian in Plant Nutrition 135 Fig. 1. Accumulation of released (Field J r~ amino acids in experiment). An analysis of other species of leguminous plants-beans (Fig. 2), clover and vetch-reveals analogous results. There is apparently no strict constancy in the quantitative composition of amino acids in the tissues of the plants studied. Their quantitative correlation perceptibly changes according to the conditions under which the plant grows; the plant may contain a minimum or maximum of the same amino acid. In the experiments conducted with lucerne on the field plots near KISHINEV (MoLDAVIAN SSR) inoculation with active strains of root-nodule bacteria yielded the following maximums in the green parts of experimental plants: cystine-1.00 mg/g (the control plants showed traces of it), lysine-0.50 mg/g (control-traces), histidine-1.00 mg/g (control-traces), arginine-0.62 mg/g (control-traces), proline-0.75 mg/g (controltraces), valine-2.2 mg/g (control-traces). In the roots the quantitative differences in the amino acids are still more clearly pronounced, the following amounts being found in the experimental plants : cystine-2.50 mg/g (control-traces), proline-5.00 mg/g (control-traces), tryptophan-5.62 mg/g (control-none), valine-2.12 mg/g (control-0.87 mg/g). The foregoing data show how important the symbiotic bacteria are in the nutrition of leguminous plants or in the accumulation of free amino acids and total nitrogen, to be exact. We cannot exactly say whether these bacteria affect the synthetic processes with their metabolites or synthesize the amino acids themselves and transfer them to the host plant. We suppose that the processes of amino acid accumulation operate in both ways. It is well known that the microbial metabolites-vitamins, amino acids, antibiotics, toxins, etc.-pass from the substratum into the roots and accumulate in the plants (see KRASILNixov, 1958). We also know that some microbial the UJ tissues r of blue lupine substances

9 136 N. KRASILNIKOV VOL. 7 Fig. (beans), 2. Chromatogramms of amino acids in the which have been cultured in the sterilized over-ground sand. tissues of haricot

10 1961 On the Role of Soil Bacteria in Plant Nutrition 137 activate the process of growth and other functions in plants (gibberelin and gibberellin-like compounds). Not only symbiotic microbes-root-nodule bacteria-but also many species of freely living soil microorganisms possess the ability to activate the processes of amino acid accumulation in plants. In our research we used simultaneously root-nodule and non-nodule bacteria (Pseudomonas, Bacterium, Azotobacter, etc.). All in all we have tested more than two dozen species. Table 8. Influence tissues of of non-nodule bacteria on lucerne and beans (in amino acid accumulation to dry weight). in the Some of them produced as great an effect as did the active, specific root nodule bacteria. The experiments with lucerne were conducted under sterile condition and with beans under semisterile condiation, in vegetative vessels, and in sand. The results shown in Table 8 indicate that the cultures of the bacteria Ps. l uorescens and Az. chroococcum are no less effective than the specific root-nodule bacteria. Under their influences free amino acids accumulate in plant tissues much more than in the tissues of control plants

11 138 N. KRASILNIKOV VOL. 7 and even more when inoculated with specific root-nodule bacteria. For example, 0.939% aspartic acid was found in the experimental plants of lucerne under the influence of Ps. fluorescens, 0.784% under that of Rh. meliloti, and 0.572% in the control plants. INFLUENCE OF BACTERIA ON AMINO ACID ACCUMULATION IN THE TISSUES OF CEREALS After the establishment of the positive effect produced by bacteria on amino acid accumulation in the tissues of leguminous plants the question naturally arose as to whether all plants reacted analogously or this ability was characteristic only of leguminous plants. In our former studies we repeatedly observed a positive stimulating effect on leguminous plants when suitable species of activating microbes were chosen. A greater crop increase due to the influence of activators is observed in cereals (wheat, maize, rye, cotton, etc.). In our subsequent investigations we therefore conducted experiments on amino acid accumulation in wheat. The plants were grown under sterile conditions in sand or in an agarized medium, in glass vessels. They were inoculated with cultures of root nodule bacteria, Azotobacter and activators of the genus Pseudomonas. Within days of growth the plants were analyzed for their free amino acids and total nitrogen content but we could not detect any essential differences between the control and experimental plants. The control plants had 4.69% total nitrogen in the over-ground parts and 2.66% in the roots. The following picture was revealed under the influence of bacteria: Ps. fluorescens over-ground parts 4.81%, roots 2.91% Az. chroococcum 4.77%, 2.73% Mixture of rhizospheric bacteria 4.83%, 2.80% Rh. meliloti 4.75%, 2.70% Only under the influence of Ps. fluorescens and the mixture of rhizospheric bacteria there was a small increase in nitrogen compared with the control plants. We discovered no differences in the qualitative composition of amino acids in these plants in any variant of the experiment. Their quantitative composition differed within small limits, but naturally the bacterized plants having more free amino acids. Under the influence of Ps. fluorescens and the mixture of rhizospheric bacteria the increase in amino acids was greater than in the other experiment. In Table 9 we show data dependent only on four experiments including the control test. A comparison of these data with the data furnished by the experiments conducted with leguminous plants warrants the assumption that activating bacteria exert a positive influence on amino acid accumulation both in cereals and leguminous plants, but in different quantitative correlations. If we add ammonium sulfate to the substratum and grow plants of

12 1961 On the Role of Soil Bacteria in Plant Nutrition 139 Table 9. Influence of activating bacteria tissues. Experiment conducted under dry weight). on amino acid accumulation in maize sterile conditions in sand (in % to maize and wheat in it in the presence of activating microbes, the effectiveness of these microbes will manifest itself more perceptibly. For example, in the experiment with wheat grown in sterile sand we found the following amounts of amino acids in the over-ground parts after 30 days cultivation: % to dry matter. Control with Ps. luorescens Histidine Arginine Aapartic acid Alanine Tyrosine Tryptophan Leucine Activating bacteria stimulate the formation and accumulation of amino acids from ammonium salt. The foregoing data show that in the presence of microorganisms the processes of synthesis and accumulation of free amino acids operate more

13 140 N. KRASILNIKOV VOL. 7 intensively. Part of the amino acids are formed by the microbes themselves and are liberated into the substratum whence they are absorbed by the plants. This was established by some authors experimentally (see SHAVLOVSKY, 1955: KRASILNIKOV, 1958; KRASILNIKOV and ASEYEVA, 1959). INFLUENCE OF SOIL BACTERIA ON THE ASSIMILATION AND SYNTHESIS OF PHOSPHORIC COMPOUNDS BY PLANTS The significance of the soil microflora in nourishing plants with phosphorus was formerly judged by its ability to transform phosphoric compounds into a soluble and assimilable state. As regards organic phosphorus it was believed that the microbes changed it to inorganic compounds accessible to plants. In the ideas of specialists this was essentially due to the role of microorganisms in nourishing plants with phosphorus. Recent studies shown, however, that soil microorganisms not only transform and change the composition of phosphoric compounds but also exert a certain influence on their assimilation, synthesis and accumulation in plant tissues (KOTELEV, 1958, KRASILNIKOV and KOTELEV, 1958). In our experiments we introduced phosphoric salts labelled with radio active phosphorus p32 into the medium and in this substrate barley, wheat and other plants were grown in the presence of certain species of bacteria and without them (sterilely). At certain intervals analyses were made to determine the content of organic and inorganic phosphorus in the tissues of the roots and over-ground parts. Since we introduced radioactive phosphorus its presence in the tissues was detected and determined by a counter. The organic and inorganic phosphoric compounds were separated from each other by the method of paper chromatography. The first series of experiments were conducted with superphosphate containing p32. The sprouts of barley were grown in sterile sand with tracer superphosphate. Some of the vessels were inoculated with a rhizospheric microflora or pure cultures of bacteria, others remained sterile. The analyses of the grown plants have shown that in the presence of microorganisms the assimilation and accumulation of phosphorus in the tissues was greater than in the control plants grown without bacteria. For example, in the experimental plants we had counts per minute for 10 mg of the dry mass (an average of 1156 c.p.m.) and in the control plants counts (an average of 430 c.p.m.). Investigations have shown that phosphorus is assimilated by the plants in inorganic and organic compounds. Both are separated by paper chromatography. Organic phosphorus is well seen on the paper in the form of circumscribed size (Figs. 3 and 4). In one of the series of experiments barley sprouts and organic phosphorus lecithin with tracer p32 were introduced into the medium. At the same time the vessels were inoculated with cultures of microorganisms. The experiments were conducted under sterile conditions. The results are shown

14 1961 On the Role of Soil Bacteria in Plant Nutrition 141 in Table 10. Fig. 3. Radioautograms (a) of inorganic and (b) organic phosphoric compounds in the sprouts of corn. 1) Plants, which grew up in Eshby's liquid media, inoculated with Azotobacter. 2) Plants, which grew up on the liquid sterilized Eshby's media. Fig. 4. 1) Radioautograms of (a) inorganic and (b) organic phosphoric compounds in soil (sterilized). 2) Radioautograms of (a) inorganic and (b) organic phosphates in plants (barley), which were grown up in sterilized soil conditions. 3) Radioautograms of (a) inorganic and (b) organic phosphates in plants (barley), which were grown up in non-sterilized soil (seeds were bacterized by soil suspension). The foregoing table shows that in the presence of bacteria the tissues of barley assimilate and accumulate S-9 times as much organic phosphorus as do the plants grown under sterile conditions (without bacteria). We conducted special experiments to prove that the bacteria synthesize organic phosporus themselves and pass it on to plants. At first it was shown that, in the nutrient medium containing inorganic phosphorus in the form of potassium phosphate, organic compounds of phosphorus were formed during

15 142 N. KRASILNIKOV VOL. 7 Table 10. ganic porus Influence of bacteria on the phosphorus in the tissues of P32 in lecithin). assimilation and barley (in c.p.m. accumulation of radioactive of or- phos- the development sence is shown Fig 5. Radioautograms of (a) inorganic and (b) organic phosphates in Eshby's media and cul tured filtrate inoculated with Azotobacter. 1) Sterilized Eshby's media (organic phosphates were not contained). 2) Cultured filtrate inoculated with Azotobacter (formation of organic phosphates is visible). of suitable bacteria (Bacterium sp., Azotobacter). Its prein the radioautograms (Fig. 5). After these experiments we added to the medium organic phosphorus with tracer p33. The latter was obtained from the cultures of bacteria which had synthesized it. The plants grown in the medium containing such phosphorus were analyzed by the counter method. (See KOTELEV, 1955; KRASILINIKOV and KOTELEV 1957). The organic phosphorus compounds were concentrated by means of chromatographic adsorption. The results have shown that the plants accumulate organic phosphorus synthesized by the bacteria. In our experiments about 9% of the tracer phosphorus in the organic compounds synthesized by Azotobacter passed into the tissues of the vetch; about 8% by the culture of Bacterium sp. and about 10% by the culture of Ps. fluorescens passed into the plants. As was to be expected, the highest concentration of these compounds was found in the root system. GENERAL CONCLUSIONS 1. Soil microorganisms play an important role in plant nutrition. Many species of microbes, especially those that abundantly develop in the rhizosphere, exert a direct influence on plant nutrition. These microbes synthesize various

16 1951 On the Role of Soil Bacteria in Plant Nutrition 143 biologically very active organic compounds and transmit them to the plants. The latter absorb through their roots the microbial metabolites and use them as sources of additional nourishment. 2. The present paper shows that some bacteria aid in the accumulation of amino acids in plant tissues. In the presence of certain species of activating bacteria the amount of amino acids in the plant tissues increases per cent or more. In leguminous plants more amino acids are accumulated under the influence of active nodule bacteria. As a rule, the more nodules on the roots, the more free amino acids in the tissues. Greater concentration of amino acids is also observed under the influence of certain non-nodule bacteria-bacterium sp., Ps. fluorescens, etc.. A particularly high increase in amino acids in leguminous plants is observed when they grow in the presence of Ps. fluorescens. 3. A similar effect is produced by certain activating bacteria on nonleguminous plants. The tissues of cereals (maize, wheat, barley, etc.) also show an increased accumulation of free amino acids in the presence of bacteria although to a lesser extent than those of the leguminous plants. 4. Certain activating bacteria influence the acumulation of phosphorus containing organic compounds in plant tissues. These compounds are in some extent synthesized by bacteria and are transmitted to the plants through the roots. This was shown in the experiments with tracer phosphorus p33 with the simultaneous use of the method of paper chromatography. REFERENCES (1) A. GEBHARDT and S. KOVALCHUK : Influence of the Azotobacter on the Vitamin Content in the Soil and Oat Sprouts. Microbiology, 27, No 3 (1958). (2) N. KoLosov and S. UKHINA : On the Role of the Root System in the Assimilation of Mineral Substances by Plants. Plant Physiology, 1, No. 37 (1954). (3) V. KOTELEV and A. GARKOVENKO: Acquisition of Phosphorus by the Cells of Microorganisms and Its Transmission to Plants (by the P32 Method). Herald of the Moldavian Branch of the USSR Academy of Sciences, No. 17, p. 15 (1954). (4) V. KOTELEV: Significance of the Soil Micro~lora in the Movement and Assimlation of Phosphorus by Plants When Introduced Focally (by the P32 Method). Herald of the Moldavian Branch of the USSR Academy of Sciences, No. 1, p 9 (1955). (5) N. KRASILNIKOV: Phytohormonal Effect of Soil Bacteria. Reports of the USSR Academy of Sciences, II No. 2 (1944). (6) N. KRASILNIKOV: Microorganisms in Additional Plant Nutrition. Achievements of Modern Biology, 33, Book 3 (1952). (7) N. KRASILNIKOV: Microorganisms and Soil Fertility. Herald of the USSR Academy of Sciences, Ser. Biology, No. 2 (1954). (8) N. KRASILNIKOV: Assimilation of Microbial Metabolites by the Plant Roots. Reports of the USSR Academy of Sciences, 110, No. 5 (1955). (9) N. KRASILNIKOV: Soil Microorganisms and Higher Plants, Moscow (1958.) (10) N. KRASILNIKOV and I. ASEYEVA: The Influence of Soil Bacteria on the. Free

17 144 N. KRASILNIKOV VOL. 7 (11) (12) (13) (14) (15) (16) Amino Acid Content of Papilionaceous Plants. Folia ticrobiologica 4, p. 45 (1959). N. KRASILNIKOV and N. BEZZUBENKOVA: Influence of Bacteria on the Assimilation of Organic Substances by Plants. Reports of the USSR Academy of Sciences, 101, No. 6 (1955). N. KRASILNIKOV and A. KORENYAKO: Influence of Non-nodule Bacteria on the Growth and Nitrogen Fixation of Leguminous Plants. Microbiology, 15, No. 5 (1946). N. KRASILNIKOV and A. KORENYAKO: Influence of Non-nodule Bacteria on the Nitrogen Fixation of Clover under Conditions of Sterile Cultures. Microbiology, 15, No. 4 (1946). N. KRASILNIKOV and V. KOTELEV: Qualitative Determination of the Phosphatase Activity of Certain Groups of Soil Microorganisms. Reports of the USSR Academy of Sciences, 117, No. 6 (1957). Y. RATNER and I. DOBROKHOTOVA: On the Possible Rloe of Vitamins, Produced by Soil Microorganisms, in the Root Nutrition of Plants. Plant Physiology, 3, No. 2 (1956). G. SHAVLOVSKY: Role of Rhizospheric Microorganisms in the Vitamin and Amino Acid Nutrition of Plants. Isotopes in Microbiology, Moscow, (1955). Translated by David A. Myshne