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I N D U S T R I A L A N D ENGINEERING C H E M I S T R Y
Vol. 15, No. 4
Recent Investigations on t h e Oxidation of Sulfur by Microorganisms'vz By J. G. Lipman STATE OF N E W JERSEYAGRICULTURAL EXPERIMENT STATION, NEWBRUNSWICK, N. J.
HIOBACILLUS medium the bacteria could The following paper is a brief discussion of some inuestigations TH I o o x I DAN8 apparently no longer on the microzrganism Thiobacillus thiooxidans, isorated under the 29318,12B*is the Sulfurwriter's direction at the New Jersey Agricultural Station, The tion and no sulfur was oxidizing organism Par exoxidized. properties and action of this organism and its use a s a sulfurcellence. It is one of the oxidizing agent to solue uarious soil problems have been described. autotrophic bacteria that USE AGAINST POTATO SCAB are able to function in mineral media and to derive their carbon from carbon dioxide. Sulfur oxidation by Thiobacillus thiooxidans has been Growth readily occurs at 20" to 25" C. in suitable media with utilized as a means for solving certain problems of agricula pH of 4.4to 4.6. Sulfur is oxidized to siilfuric acid, and the tural production. It is well known that far-reaching damage organism, because of the energy set free, is able to utilize the is done t o our potato crop in certain localities by Actinomyces carbon of carbon dioxide for the building of its cell substance. scabies. This organism will grow best in soils that are only It has been found in our investigations that for every very slightly acid or not a t all acid. For many years thig 32 parts of sulfur oxidized there is 1part of carbon assimi- fact has been recognized by soil investigators and by plant lated. This would represent a utilization of about 6.5 per pathologists, and such methods of soil treatment have been cent of the energy liberated in the oxidation of sulfur to sul- recommended as would tend to increase the acidity of the furic acid. In case df the bacteria capable of oxidizing soil. The use of fertilizers tending to increase the hydrogenammonia to nitrites the ratio of nitrogen transformation to ion concentration of the soil solution is advocated. Ammocarbon assimilated is 35 : 1. In case of the organisms capa- nium sulfate and other ammonium salts are more or less effecble of oxidizing nitrites to nitrates there is 1 part of carbon tive for this purpose. Potassium chloride and potassium assimilated for each 135 parts of nitrogen transformed. In sulfate, as well as acid phosphate, are helpful in a limited view of the fact, however, that the oxidation of nitrites to way. Organic acids derived from the decomposition of rye nitrates makes available only one-fourth of the amount of and of other crops used as green manures may also serve to energy that is made available in the oxidation of equivalent lessen the damage done by the potato-scab organism. Invesquantities of ammonia nitrogen to nitrites, the differences tigations covering a period of more than twenty-five years are, after all, slight. It is evident, therefore, that in the have indicated the effectiveness of sulfur for the same purpose. oxidation of ammonia to nitrites and of nitrites to nitrates The New Jersey Experiment Station was one of the pioneers about 5 to 6 per cent of the energy available is utilized for in the study of sulfur as a means of controlling the damage the assimilation of carbon by the bacteria. This does not done by the potato-scab organism. Until recent years, howdiffer very greatIy from the amount of energy used for the ever, consistent results with sulfur were not obtained by same purpose by Thiobacillus thiooxidans. In the case of different investigators. It was finally shown by the New higher plants only 1 per cent of the total energy of the sun's Jersey Stationg that sulfur often fails to increase the acidity of radiation is used for the assimilation of carbon from carbon the soil promptly and effectively to a point where the activities dioxide. It may be added here that aztobacter, representing of the potato-scab organism are more or less effectively the most efficient of the nonsymbiotic nitrogen-fixing bac- suppressed. Two of the important factors that have apparteria, will assimilate, under favorable conditions, as much ently been overlooked by earlier investigations are the need as 2 parts of nitrogen for every 100 parts of sugar used. In of uniform distribution of the sulfur and the need of supplythis case, however, the utilization of the carbohydrate is ing an organism or organisms that would rapidly oxidize not complete, since organic acids and other organic products sulfur to sulfuric acid. Within the past three years the writer and his associates have compared inoculated with uninocuappear in the culture medium. Thiobacillus thiooxidans has a remarkable tolerance for lated sulfur on many different soils and under varying condisulfuric acid. I n one of our experiments sulfuric acid was tions. The results secured in these experiments have been added to the medium in varying amounts and the rate of published in part. Other data are to be published in the oxidation of sulfur was determined. The increasing con- near future. On the basis of these numerous experiments, centration of the sulfuric acid did not seriously retard the covering a period of three years, it is safe to state that sulfur activities of Thiobacillus thiooxidans until the concentration inoculated with Thiobacillus thiooxidans is much more effective than equivalent amounts of sulfur not inoculated with of 5 per cent of acid was reached. these It appears that 300 lbs. of inoculated With 0.025 per cent of acid there were oxidized in 100 CC. sulfur organisms. are practically as effective as 600 lbs. of uninoculated of the culture medium containing 1 g. of sulfur an equivalent sulfur. Methods have been developed for inoculating SUIof 249.3 mg. of sulfur. When the concentration of the acid reached 5 per cent there was still oxidized 75.55 mg. fur with Thiobacillus thiooxidans on a commercial scale. With a concentration of 10 per cent of acid in the T o PRODUCE SOLUBLE PHOSPHATE
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1 Presented before the Division of Fertilizer Chemistry at the 64th Meeting of the American Chemical Society, Pittsburgh, Pa., September 4 to 8, 1922. 2 Paper No. 118 of the Journal Series, New Jersey Agricultural Experiment Station, Department of Soil Chemistry and Bacteriology. Numbers in text refer to bibliogtaphy at end of article.
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The writer suggested some years ago6~I0 that the biological oxidation of sulfur might be employed as a means of producing soluble phosphates. The method as proposed is quite simple. Quantities of ground phosphate rock and
April, 1923
I N D U S T R I A L A N D ENGINEERING C H E M I S T R Y
ground sulfur were intimately mixed with soil and inoculated with Thiobacillus thiooxidans. The sulfuric acid resulting from the oxidation of the sulfur reacted with the tricalcium phosphate and led to the formation of dicalcic and monocalcic phosphate. Our more recent 4* studies have dealt with more effective methods for the production of available phosphates by the process just described. It has been found that when mixtures of sulfur and ground phosphate rock are broadcasted on any soil area conveniently located, and worked into the surface soil by means of ordinary tillage implements, the formation of available phosphates goes on fairly rapidly. If successive additions of mixtures of sulfur and ground phosphate rock are made from time to time and similarly worked into the surface soil, a concentrated material is finally obtained which contains 16 per cent of total phosphoric acid, of which 50 to 60 per cent is citrate-soluble. The time necessary for reaching this result is about 15 to 16 wks. It is expected by the writer and his associates that further improvements in the technic will reduce the time to 12 to 15 wks. Under such conditions it may become practicable to produce any desired quantity of acid phosphate a t a very slight cost. This could best be done a t points from which supplies of both phosphate rock and sulfur may be readily reached. Portions of Florida, Louisiana, and Texas offer such locations and possess the further advantage of being situated under climatic conditions suitable for the functioning of the sulfur-oxidizing organisms throughout the year. 31
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USE ON LANDDEFICIENTIN SULFUR The oxidation of sulfur by microorganisms makes this substance suitable for use on land that is deficient in sulfur. It has been demonstrated that in sections of Oregon, Washington, and Idaho much of the land responds to applications of sulfur-carrying materials. Gypsum was a t first used by farmers in these localities for increasing the yields of alfalfa. More recently it has been found that sulfur is practically as effective, and, from the standpoint of actual concentration in sulfur, much more satisfactory. Experiments5 carried on at the writer’s suggestion with samples of soil from Oregon show that the addition of Thiobacillus thiooxidans intensifies the rate of oxidation of the sulfur and makes the latter much more effective in supplying this element to the crop. Many hundreds of tons of sulfur are already used in the regions referred to and the consumption will materially increase. The greater effectiveness of inoculated sulfur should make the results from its use much more certain. BLACKALKALIPROBLEM Another agricultural problem that may be solved through the oxidation of sulfur by microorganisms is the so-called black alkali problem. I n certain of the soils on the Pacific Coast an accumulation of soluble salts containing substantial proportions of sodium carbonate and bicarbonate leads to the deterioration of the texture of the soil. The land becomes impervious and ordinary methods of reclamation by the washing out of the excessive salts fail to produce the desired results. The writer was, therefore, led to suggest that the use of sulfur might offer a satisfactory solution. The sulfuric acid formed from the oxidation of sulfur would react with sodium carbonate and bicarbonate and lead to the formation of sodium sulfate. The texture of the soil would improve to a point’ where the soluble salts could be more or less effectively washed out. Experiments were accordingly initiated with soil derived from black-alkali spots in California. Inoculated sulfur was employed a t the rate of 2000 to 6000 lbs. per acre. Two weeks after the inoculated sulfur had been added to the soil it was found that 20 to 25 per cent of the
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sulfur had been oxidized, all carbonates had disappeared, and the bicarbonates were reduced to about 40 per cent of the original amount. The p H of the soil had changed from 8.8 to 7.9. Where sulfur was added a t the rate of 6000 lbs. per acre, the permeability of the soil to water improved to the extent of 30 to 40 per cent. It was evident that the colloids present in the soil had largely been coagulated by the sulfuric acid formed in the oxidation of the sulfur. The experiments are being continued and*itis expected that further improvement of the texture of the soil and increased permeability will be brought about. CONTROL OF OTHERPLANT DISEASES Among the other soil problems capable of being solved through the biological oxidation of sulfur, there may be noted the control of other plant diseases, such as scurf and soil rot of sweet potatoes, the scab of sugar beets, and certain diseases of flax and of tobacco. It is quite probable that the biological oxidation of sulfur may also serve as an effective means for destroying soil-infesting insects and likewise nematodes. Among the soil-infesting insects wire worms are particularly destructive. Experiments are being undertaken at the New Jersey Station on the control of these and of white grubs by means of sulfur oxidation. Experiments are also being undertaken on the control of nematodes by means of the oxidation of inoculated sulfur. These are one of the most serious and destructive of the known plant parasites. Should these experiments yield the results expected from them we shall have found an economical means for enhancing crop production. OTHERUSES Other uses of inoculated sulfur have been suggested. At this time mention may be made of the production of potassium sulfate from the mineral glauconite; the control of intestinal parasites of sheep, swine, and poultry; and the eradication of objectionable vegetation. These and other possible uses of inoculated sulfur would seem to indicate a wide usefulness for a material that is both cheap and abundant. BIBLIOGRAPHY I-Joffe, “Studies of Sulfur Oxidation in Sulfur-Floats-Soil Mixtures,” Soil Science, 13 (1922), 107. 2-Joffe, “Preliminary Studies on the Isolation of Sulfur-Oxidizing Bacteria from Sulfur-Floats-Soil Composts,” Soil Science, 13 (1922), 161. 3-Joffe, “Some Agricultural Relations of Sulfur.” Thesis, R u t g e n College (soon t o appear as a technical bulletin). 1922. G J o f f e , “Acid Phosphate Production by the Lipman Process,” a series of three articles appearing in Soil Science, beginning December, 1922. &Joffe and McLean, “A Note on the Oxidation of Sulfur in Oregon Soils,” Soil Science, 14 (1922), 217. 6-Lipman, McLean, and Lint, “Sulfur Oxidation in Soils and Its Effect on the Availability of Mineral Phosphates,” Soil Science, 2 (1916), 499. 7-Lipman and Joffe, “The Influence of Initial Reaction on the Oxidation of Sulfur and the Formation of Available Phosphates,” Soil Science, 10 (1920), 327. 8-Lipman, Waksman, and Joffe, “The Oxidation of Sulfur by Soil Microorganisms,” Soil Science, 12 (1921), 475. 9-Martin, “The Relation of Sulfur t o Soil Acidity and the Control of Potato Scab,” Soil Science, 9 (1920), 393. l&-McLean, “The Oxidation of Sulfur by Microorganisms in Its R e lation t o the Availability of Phosphates,” Soil Science, 6 (1918), 251. 11-Rudolfs, “Composting Rock Phosphate with Sulfur in Slightly Alkaline Calcareous Soils,” Soil Science, 14 (1922). 37. 12-Waksman and Joffe, “Microorganisms Concerned in the Oxidation of Sulfur in Soil,” J . Bact., 7 (1922), 239. 13-Waksman and Starkey, “Carbon Assimilation and Respiration o! Autotrophic Bacteria,” Proc. Soc. E x p . Biol. M e d . , 20 (1922), 9.
