from microorganisms and crop · vol. 28, 1964 phytotoxic substances fromsoil microorganisms...

BACTERIOLOGICAL REVIEWSVol. 28, No. 2, pp. 181-207 June, 1964Copyright @ 1964 by the American Society for Microbiology

Printed in U.S.A.

PHYTOTOXIC SUBSTANCES FROM SOIL MICROORGANISMS ANDCROP RESIDUES'

T. M. MCCALLA AND F. A. HASKINSSoil and Water Conservation Research Division, U.S. Department of Agriculture, and Nebraska

Agricultural Experiment Station, University of Nebraska, Lincoln, Nebraska

INTRODUCTION................................................................................. 181SOIL MICROBIAL PRODUCTS INHIBITORY TO MICROBIAL GROWTH ..................... 183PHYTOTOXIC SUBSTANCES IN PLANTS..............................................................184Germination Inhibitors....................................................................... 184Growth Inhibitors........................................................................... 186Plants containing growth inhibitors.......................................................... 186Nature of growth inhibitors in plants ........................................................ 187

EARLY WORK ON PHYTOTOXIC SUBSTANCES FOUND IN PLANT RESIDUES AND SOIL ............. 187Isolation of Specific Compounds From Soil.................................................... 188Effect of Straw on Plant Growth ................................... 188

RECENT WORK ON THE OCCURRENCE OF PHYTOTOXIC SUBSTANCES IN SOIL AND THE PRODUCTION OFTHESE SUBSTANCES BY MICROORGANISMS.................................................... 188

Influence of Certain Organic Compounds and Antibiotics on Plant Growth ....................... 188Organic Acids and Amino Acids in Soil....................................................... 190Influence of Carbon Dioxide, Bicarbonates, Nitrite, Hydrogen Sulfide, and Ammonia on Plant Growth. 191Production of Phytotoxins by Soil Microorganisms and Plant Pathogens........................... 192

Gibberella fujikuroi........................................................................ 193Penicillium ................................................................................ 193Aspergillus ................................................................................ 194Trichoderma and Fusarium ................................................................ 194Alternaria ................................................................................. 195Other microorganisms....................................................................... 195

Phytotoxic Substances in Decomposing Plant Residues, Soil Organic Matter, and Soil ..... .......... 195Phytotoxic Substances in Forest Soils ................................ 196Phytotoxic Effects Encountered in Stubble Mulching ............................................ 197

SOME SPECIFIC PLANT EFFECTS POSSIBLY CAUSED BY TOXINS FROM MICROORGANISMS ............ 198Legume Bacteria Factor in the Chlorosis of Soybeans............................................. 198Frenching Disease of Tobacco ................................................................ 198Replant Problem in Peaches. .......... ..................................................... 199Replant Problem in Citrus .................................................................... 199Sod-binding of Bromegrass.................................................................... 201Stunting of Corn Associated with Stubble Mulching ........................................... 201Seedling Disease of Rice ...................................................................... 202Inoculation Failure in Subterranean Clover .............................. 202

CONCLUSIONS................................................................................. 202OUTLOOK ............................................................................. 202LITERATURE CITED ............................................................................ 203

INTRODUCTION mary source of minerals for plant growth. In addi-The role of organic matter in the growth of tion, beneficial effects on soil structure and water-

plants has been a subject of much investigation holding capacity were attributed to the presenceand controversy since the sixteenth century. Dur- of organic matter. With the development of agri-ing the early era, decomposition of plant residues cultural chemistry culminating in the work ofand green and animal manures provided a pri- Liebig in the nineteenth century, it was dis-

covered that plants could make satisfactoryI Published with the approval of the Director as growth in the absence of added organic matter if

Paper no. 1354, Journal Series, Nebraska Agricul- suitable kinds and amounts of inorganic nutrientstural Experiment Station, Lincoln. were supplied. However, persistent efforts were

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MCCALLA AND HASKINS

made by various investigators and lay groups toemphasize the important role of organic matter,rather than inorganic fertilizers, in soil fertilityand plant growth. Because of the complexity ofthe problem and lack of suitable techniques, nosatisfactory solution to the controversy was ob-tained during Liebig's time.

Since the Liebig era, frequent attempts havebeen made to determine in detail the influence oforganic matter on plant growth. For example,

develop normally with adequate supplies of inor-ganic nutrients, but they may also take up manyorganic substances which influence growth. Inrecent years, a renewed interest has developed inthe role of these organic substances. This interesthas been greatly stimulated by the availability ofnew techniques such as paper, gas, and columnchromatography and infrared spectrophotometrywhich aid in the isolation and identification of or-ganic compounds.

FIG. 1. Influence of stubble mulching with 8 tons of straw per acre on growth of corn at Lincoln, Neb. Noteirregular growth of corn on mulched plot.

Schreiner and Shorey (113) attempted to explainthe low productivity of some soils on the basis ofcertain toxic organic substances present. Dihy-droxystearic acid and vanillin were isolated andshown to be toxic to plants in aqueous solution.However, it was shown that soil and fertilizerstend to neutralize the effect of these toxic sub-stances. Since adequate mineral fertilizationlargely overcame the toxic effects of such com-pounds, most investigators doubted that organicsubstances in the soil had any important directinfluence on plant growth. With the discovery andutilization of antibiotics and herbicides, therewas a re-examination of this position. Plants may

Recent field observations disclosed the occur-rence of reduced plant growth under the systemof farming known as stubble mulching (Fig. 1).McCalla and Army (80) and others suggested thatthis effect was the result of toxic decompositionproducts of microorganisms or substances fromplant residues. Similar observations of retardedplant growth encountered in citrus replant andpeach tree replant, seedling inhibition andfrenching of tobacco, sod-bound bromegrass, andreduced growth of forest tree seedlings in some in-stances add support to the belief that growth in-hibitors may accumulate in the soil (9, 72, 95,96, 128).

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VOL. 28, 1964 PHYTOTOXIC SUBSTANCES FROM SOIL MICROORGANISMS

Certainly under field conditions, with heavyfertilization, large quantities of plant and animalresidues are returned to the soil to decompose.Numerous organic and inorganic substances re-sult from this decomposition. The concentrationof any one constituent may vary greatly withtime and environmental conditions. Minutequantities of many organic substances have pro-nounced effects on plant growth, and it is highlyprobable that, under certain conditions, concen-trations of these substances become sufficientlyhigh to cause significant effects on plant growth.The purpose of this paper is to review research

(122), and others (17, 19, 38, 48, 142, 143, 144)have shown that many actinomycetes and fungican produce detectable quantities of antibioticsin sterile soils supplemented with organic matter.Grossbard (48) also found that antibiotics wereproduced in autoclaved soils enriched withreadily available sources of carbon and inocu-lated with Penicillium patulum or Aspergillusclavatus. Hessayon (52, 53) and Schmidt (110),in reviews of the literature, cited several reportsof toxicity of soil extracts to fungi and bacteria.Antibiotic production in sterile soil was also re-ported.

TABLE 1. Diameter of fungal colonies with and without alcohol extracts of surface soils

Avg diam of colonyb

Fungus Expt A Expt B Expt C

Cltr IExtract from Control Extract from Con Extract fromon I g of P55-i 3 g of P55-i1 oto 3 g of P54-i

mm mmn mm mm mm mm

Penicillium humicola .21. 019.. ** 23.5 19.8** 23.2 19.7**Aspergillus8sydowi.10.7 9.2* 13.7 10.8* 12.8 10.7*Zygorhynchus vuillemini. 78.5 | 78.7 56. 3c 48.5**c 53.Oc 55.5cSporotrichum pruninosum 20.2 18.7** 24.3 20.7** 22.0 19.5**Monilia humicola.22.0 19. ** 27.8 18.8** 28.0 22.5**

a This table appeared first in Soil Sci. 55:377-391, 1943. See reference 90. Published with permission ofThe Williams & Wilkins Co.

b Average of three plates after 5 days of incubation. Symbols: * = significant by t test in comparisonwith control, beyond 5% level; ** = highly significant by t test in comparison with control, beyond 1%level.

c Diameter measured on third day of incubation.

on microbial decomposition products and organicsubstances in crop residues and soil which mayinhibit plant growth. No attempt was made toreview all the literature available. Only typicalreferences are cited to illustrate salient points.

SOIL MICROBIAL PRODUCTS INHIBITORYTO MICROBIAL GROWTH

Numerous investigators (17, 44, 48, 69, 125,130) have shown that microorganisms produce awide variety of compounds (antibiotics) inhibi-tory to microbial growth. Some of these com-pounds are toxic to plant growth, as will be dis-cussed later. Many of the compounds are selec-tive in their action; others, such as penicillin, in-hibit the growth of a wide variety of micro-organisms.

Stevenson (127), Gottlieb et al. (45), Stallings

Newman and Norman (90) showed that driedethanol extracts of surface soil samples from Mar-shall silt loam and Clarion silt loam in Iowa con-tained toxic substances, probably of microbialorigin, that inhibited the growth of soil micro-organisms as shown in Table 1. The investigatorsfound that aqueous soil extracts did not reducethe growth of soil bacteria. However, the alco-holic extract of both surface and subsurface soilsamples reduced the activity of soil microor-ganisms.Hessayon (53) found that Trichothecium roseum

produced less trichothecin in nonsterile soils thanin sterile soils. Clay soils had a higher trichothe-cin-producing capacity than did sandy ones.Brian (17, 19) indicated that a number of fungi,actinomycetes, and bacteria are known to produceantibiotics in both sterile and normal soil if the

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soil is suitably supplemented with organicmatter.

Pinck et al. (101, 102) and Soulides et al. (121),in a study of the adsorption of antibiotics by clayminerals, showed that the antibiotics could beclassified into three groups: (i) strongly basic(streptomycin, dihydrostreptomycin, neomycin,and kanamycin); (ii) amphoteric (bacitracin,chlortetracycline, and oxytetracycline); and (iii)acidic (penicillin) or neutral (chloramphenicol andcycloheximide). The strongly basic antibioticswith one exception were not released from mont-morillonite, vermiculite, or illite. Streptomycinand dihydrostreptomycin were released to vary-ing degrees from kaolinite. The amphoteric anti-biotics were released from all the clay minerals.Bioassays indicated that, to be effective in in-hibiting microbial growth, the antibiotics had tobe released from the clays.

Brian (17, 19) showed that, while many anti-biotics, such as penicillin, viridin, gliotoxin, fre-quentin, and albidin, are labile substances andare rapidly decomposed in the soil, others likefradicin and cycloheximide are very stable in anumber of soils. Jefferys (56) tested the stabilityof the antibiotics albidin, frequentin, gladiolicacid, gliotoxin, griseofulvin, mycophenolic acid,patulin, viridin, penicillin, and streptomycin invarious soils. The antibiotics persisted in naturalsoils for varying lengths of time from a few daysto more than 20 days, depending on the kind ofsoil, pH, and microbial activity. Inactivation ofthe antibiotics was caused by adsorption by soil,microbial activity, or instability of antibiotic atcertain pH levels in the soil. The effect of pH wasimportant in the inactivation of albidin, frequen-tin, gliotoxin, penicillin, and viridin. Strepto-mycin was inactivated largely by adsorption.Biological inactivation of griseofulvin, myco-phenolic acid, and patulin was noted.Many factors appear to be involved in the pro-

duction as well as the persistence of antibiotics innonsterile soil. A microorganism that grows welland produces abundant yields of antibiotic insterile culture may not be able to survive, muchless to produce an antibiotic, in competition withthe entrenched flora in a nonsterile soil (57). How-ever, Brian (19) states that in certain circum-stances antibiotics are produced in appreciablequantities by microorganisms in soil. The chiefnutrient requirement for antibiotic productionappears to be a suitable carbon source. Thus, sub-

stantial amounts of antibiotics were produced ininoculated sterile soil supplemented with organicmatter, less in inoculated sterile unsupplementedsoil, and little or none in normal soil unless sup-plemented with large amounts of organic matter.In a normal soil there may be local areas whereconcentrations of organic matter are sufficientlyhigh to favor the production of antibiotics if theproper microorganisms are present.

PHYTOTOXIC SUBSTANCES IN PLANTSMany plants contain substances that adversely

affect seed germination and plant growth (39, 79,140). The return to the soil of residues from suchplants provides an obvious source of phytotoxicmaterials whose formation does not depend uponthe activity of microorganisms.

Germination InhibitorsCox et al. (29) found germination inhibitors in

the seed coat of certain varieties of cabbage. Theinhibitor could be removed with water, alcohol,or sulfuric acid. Smith (120) showed that chaffof small grains inhibited the germination of ce-reals. Siegel (117) found that water extracts of redkidney beans inhibited germination and growthof flax and wheat. Various experiments with theextracted material indicated that more than oneinhibitor was present. Barton and Solt (7) demon-strated germination inhibitors of wheat seeds inextracts of seeds of Sorbus aucuparia, Berberisthunbergii, of three varieties of Lactuca, and ofnondormant seeds of Phaseolus sp., Triticum sp.,Hordeum sp., Glycine sp., Solanum melongena, andNicotiana tabacum.

Evenari (39), in a review article, reported thatgermination inhibitors are widely distributed inmany plant species. Thus, inhibitors were foundin the pulp, juice, and coat of fruit, in seed coatand embryo, in the leaf sap, and in bulbs androots. Some of the materials responsible for ger-mination inhibition were: (i) hydrogen cyanidefrom amygdalin in many seeds; (ii) ammonia, re-leased by hydrolysis of nitrogenous compounds;(iii) ethylene, liberated from many fruits as wellas from whole plants; (iv) mustard oils contain-ing isothiocyanate derivatives; (v) organic acids,such as malic and citric acids formed by theguayule plant, or caffeic and ferulic acids fromtomato juice; (vi) unsaturated lactones, such asparasorbic acid, anemonin, and coumarin; (vii)aldehydes, such as benzaldehyde, salicylaldehyde,

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TABLE 2. Effect of aqueous extracts of plants on germination and growth of Mida wheat*

Coct aeroft congeoftl eg~Plant family Species germnna- oleopthltiont g

Asclepias syriaca 25

25

2

2525252

2

512345

2225

25

255

555

25

22

2

91

6451

7145

69

68406647645055

75

727369615356

81737169

5840

715560

608771

6557

7169

83

cm

3-5

2-41-3

2-41-2

2-3

1-30.5-1.5

2-41-21-2

0.5-1.51-3

2-3

1-22-41-31-31-2

MCCALLA AND HASKINS

TABLE 2-(cont.)

Plant family Species Peenageeoft- Range*opi | Range of rootCoctgermina- cleogthl lengthtiont entcm cm

Polygonaceae.Polygonum pennsylvanicum 2 76 2-3 1-2

Portulacaceae...... Portulaca oleracea 2 75 2-3

VOL. 28, 1964 PHYTOTOXIC SUBSTANCES FROM SOIL MIVROORGANISMS

LeFevre and Clagett (67) found that water ex-tracts of quackgrass rhizomes inhibited growth ofeight crop plants by 50 to 84%. Upon concentra-tion by paper chromatography and ion-exchangecolumn techniques, the growth inhibitor wasfound to be anionic, soluble in polar solvents,insoluble in nonpolar solvents, and unstable instrong acids and bases.Jameson (55) prepared water extracts and 50

and 100% ethyl alcohol extracts of leaves andherbaceous stems of plants representing 20 differ-ent species, including nine trees and shrubs, eightforbs and half shrubs, and three grasses. Extractsof all species tested inhibited the growth of wheatradicles. Extracts of Cordylanthus wrightii had thegreatest inhibitory effect; those of Erodium cicu-tarium and Agropyron smithii had the least effect.Although the greatest inhibitory effects werenoted in extracts of species that also displayed thestrongest toxic effects in the field, Jameson con-cluded that the existence of a growth inhibitor ina plant extract does not necessarily mean that theinhibitor is of ecological importance in the field.

Guenzi and McCalla (49) made water extractsof sweetclover stems, wheat straw, soybean hay,bromegrass hay, oat straw, sweetclover hay, andcorn and sorghum stalks, and observed theireffects on growth of sorghum, corn, and wheatseedlings. The toxicity of the extracts increasedin the order of their listing.

Cochrane (25) found that corn residue in-creased root rot and decreased yield of lettuce.The residue, added to soil in concentrations of 0,1, 2, and 5%, caused lettuce seedling mortalitiesof 0, 0, 41, and 44%, respectively. After the soil-residue mixture was incubated under warm, moistconditions for a period of 30 days, seedling mor-talities were reduced to 8, 3, 0, and 5%, respec-tively. The effect was not attributable to nutrientstarvation nor to simple pathogenesis by a nema-tode or microorganism. Cochrane suggested thatthe plant residues exerted a direct toxic effect onthe roots of susceptible plants, which was prob-ably complicated by action of secondary rot-producing microorganisms.

Other growth inhibitors have been reported inleaves and stems of sugarcane (35), immaturesoybean seeds (61), guayule roots (13), oxalisroots (12), in macadamia (Macadamia integrifolia,Al. tetraphylla, and their hybrids) (86), andinchoumoellier (Brassica oleraceae var.) (23). Obviously,

growth-inhibiting substances are widely dis-tributed among plant species.

Nature of growth inhibitors in plants. Borner(14), in a review dealing with liberation of organicsubstances from higher plants in relation to soilsickness, discussed the following compounds:(i) liberated from roots-amino acids, sugars,scopoletin and scopoletin glycoside, trans-cinna-mic acid, and enzymes; (ii) liberated from seedsand fruits-amino acids, sugars, flavones, phe-nolic compounds, and gaseous excretions (ethyl-ene, ammonia, hydrocyanic acid); (iii) liberatedfrom plant residues-phenolic compounds, 3-acetyl-6-methoxybenzaldehyde, amino acids,amygdalin, coumarin, and phlorizin; (iv) libe-rated from leaves-absinthin, amino acids, andjuglone.Woods (140), in a review of biological antagon-

ism due to phytotoxic root exudates, indicatedthat representatives of many plant families pro-duce phytotoxic root exudates containing oils,antibiotics, coumarin, alkaloids, thiamin, biotin,mineral salts, enzymes, aldehydes, nitrogenouscompounds, ammonia, amino acids, nucleotides,and nucleic acid derivatives.Garb (43) listed additional plant constituents

with growth-inhibiting properties as follows:protoanemonin from Anemone pulsatilla; byakan-gelicin from Thamnosma montana; parasorbicacid from Sorbus aucuparius; vanillin from vanillabean; arbutin from Bergenia crassifolia and Pyruscommunis; scopoletin from oats (Avena); andumbelliferone, plant not listed.

In other work on the production of growth in-hibitors by plants, Bonner (12) found that oxalicacid was secreted by the roots of oxalis; Bonnerand Galston (13) observed that trans-cinnamicacid was secreted by guayule roots; and Gray andBonner (46, 47) identified 3-acetyl-6-methoxy-benzaldehyde in extracts of Encelia farinosaleaves.

EARLY WORK ON PHYTOTOXIC SUBSTANCESFOUND IN PLANT RESIDUES AND SOIL

As indicated in the introduction, Liebig's workin the nineteenth century was followed by aperiod during which relatively little emphasis wasgiven to studies dealing with the role of organicmatter in plant nutrition. However, sporadicefforts have been made to demonstrate the im-portance of organic matter for plant growth. Sev-eral examples of such work, done between 1900

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and 1925, have been selected for inclusion in thisreview.

Isolation of Specific Compounds from SoilAbout 1910, Schreiner and Shorey (113) and

others (111, 112, 118), in attempting to explainthe poor growth of plants on some soils, isolatedspecific organic compounds from the soil. One ofthe compounds was dihydroxystearic acid, a sub-stance which reduced plant size and had the addi-tional effects of darkening the root tips, stuntingroot development, causing enlarged root endswhich were often turned upward like fishhooks,and strongly inhibiting the oxidizing power of theroots. When this compound was added to nutrientsolutions, it was toxic to plant growth in concen-trations of about 20 to 100 ppm (Table 3). How-ever, the effect of dihydroxystearic acid was

TABLE 3. Effect of dihydroxystearic acid on growthand transpiration of wheat seedlings in

solution culture*

Concn of dihydroxystearic Percentage of controlacid Transpiration Green wt

ppm

20 75 8750 56 78100 24 53200 20 54

* This table appeared first in U.S. Dept. Agr.Bur. Soils Bull. 53:1-53, 1909. See reference 113.

largely neutralized in soil. Fraps (41) found thatvanillin and coumarin were oxidized rapidly inthe soil; about 20 to 50% disappeared within 2weeks.

Schreiner and Skinner (114), Fraps (41), andFunchess (42) showed that the harmful effect ofdihydroxystearic acid could be largely overcomeby adequate mineral fertilization of the plant.However, toxic effects of coumarin and vanillinwere not prevented by the application of inor-ganic nutrients.

Skinner and Noll (119) found vanillin andsalicylic aldehyde in a silty clay loam soil atArlington Farms, Va., several months after thesecompounds were applied and after the end of thecrop season. On plots fertilized with sodiumnitrate, however, vanillin was quickly destroyed.Neither vanillin nor salicylic aldehyde persistedin limed soil. Truog and Sykora (135) found thatfinely divided soil materials such as lime, soil,

kaolin, or quartz largely neutralized the phyto-toxicity of vanillin or guanidine carbonate.

Effect of Straw on Plant GrowthOn the basis of qualitative chemical tests, Colli-

son and Conn (27) concluded that salicylic acid,dihydroxystearic acid, and vanillin were presentin straw. They suggested that these compoundsmight account for the detrimental effect of strawon plant growth, although they were also awareof the carbon-nitrogen relationships which hadbeen demonstrated by the classical works ofDoryland (34) and Murray (87). Collison (26)observed that water extracts of straw and alfalfahay were toxic to barley seedlings. The extractfrom alfalfa hay was more toxic than that fromwheat straw. Autoclaving the water extract didnot destroy the toxic factor. Later, Myers andAnderson (88) discounted the toxicity idea andconcluded from their experiments that the in-hibitory effect resulted simply from the tie-up ofavailable nitrogen by plant residues such asstraw.

RECENT WORK ON THE OCCURRENCE OFPHYTOTOXIC SUBSTANCES IN SOIL ANDTHE PRODUCTION OF THESE SUB-STANCES BY MICROORGANISMS

The discovery of antibiotics in 1929 by Flem-ing and the discovery of plant hormones or growthsubstances by Went and others (3, 11, 139) pro-vided new stimulation for studies concerned withthe influence of organic soil constituents onplants.

Organic substances in the soil are produced byor derived from animals, plants, and microorgan-isms. As Starkey (123) indicated, these organicmaterials in the rhizosphere may have a varietyof effects on plants. For example, plant growthmay be stimulated or inhibited, or the food valueof the plant may be affected. In this review,primary emphasis is placed on microorganisms asthe sources of these organic substances, althoughplant or animal sources will be discussed, particu-larly as they serve as substrates for microbialaction.

Influence of Certain Organic Compounds andAntibiotics on Plant Growth

Prill et al. (104) studied the growth of wheatroots in the presence of a large number of organicacids in concentrations ranging from 0.002 to 6.25

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TABLE 4. Lowest concentration of added organic compounds showing toxicity to ladino cloverseedlings in a complete nutrient culture, and the concentrations allowing optimal normal

growth in the absence of inorganic nitrogen*

Lowest Concn for Lowest Concn forOrgniccomoun toic onc optimal Organic compound toxic concn optimalOrganiccompound toxicgrowth growth

Amino AcidsL-Cystine .................D-Isoleucine...............DL-Lysine .................L-Tyrosine ................Cysteine-HCl .............HydroxyprolineL-Isoleucine...............DL-Isoleucine ..............L-Leucine .................DL-Serine .................Tryptophan...............DL-Alanine................D-Arginine HC1...........L-Asparagine ..............L-Aspartic acid............D-Glutamic acid...........Glycine ...................L-Histidine-HCI ...........L-Proline ..................DL-Threonine .............DL-Valine .................DL-Methionine ............DL-Norleucine .............L(+)-Glutamic acid .......

Nucleic Acid GroupNucleic acid..............Adenylic acid.............Guanine-HCl..............Uracil ....................Adenine sulfate...........Xanthine .................

AlkaloidsL-Hyoscyamine............Atropine..................Brucine...................Morphine .................Nicotine ..................Trigonelline...............Arecoline .................Caffeine...................Quinidine .................Nicotyrine ................Nornicotyrine.............Quinine-HlI ..............Theobromine..............

VitaminsCa panthothenate.........Pyridoxamine .............Ascorbic acid.............Thiamine .................

ppm

NTNTNTNT1,0001,0001,0001,0001,0001,0001,000500500500500500500500500500500200200

NT1,000500500200200

NTNT500500500500200505050505050

NT

1,0001,000

ppm

Si1,000500200Un500

1,0001,000

500500200

200-50050

200500500200500500500500UnSi200

1,0001,000

5020050Un

UnUnUnUnUn

200-500UnUnUnUnUnUnUn

UnUn

Un

Vitamins (cont.)Pyridoxine-HlI . .......p-Aminobenzoic acid.......Niacinamide...............Nicotinic acid.............Pyridoxal-HCl .............

AntibioticsPenicillin G-K.............Penicillin G-Na............Chlortetracycline-HCI.Oxytetracycline ............Streptomycin..............Bacitracin.................Chloramphenicol ...........

MiscellaneousFumaric acid..............Maleic anhydride..........Na Cu chlorophyllin.......Oxamide...................Rutin .....................Succinic acid..............Tannic acid...............Arbutin ...................Citric acid.................Malic acid.................Malonic acid..............Methylamine-HCI.Pancreatin ................Pepsin.....................Vanillin ...................Catechol ..................Cinnamic acid.............Creatinine.................Diethylphosphite ..........Phloroglucinol......a-Picoline .................Pyridine...................Dimethylhydroresor-

cinol ....................Na taurocholate...........Succinimide ...............Aniline nitrate.............Barbital...................Biuret .....................Coumarin .................Hydroxyquinone ...........m-Nitrobenzoic acid.......Quinone ...................Benzidine .................Quinoline ..................Stovaine...................Sulfapyridine..............

ppm

500200200200200

1,0001,000

50505055

NTNTNTNTNTNTNT1,0001,0001,0001,0001,0001,0001,0001,000500500500500500500500

200200200505050505050505555

ppm

Un200200UnUn

200

50

UnUn

Un50UnUnUnUn

Un

UnUn

Un

* This table appeared first in Agron. J. 52:317-319, 1960. See reference 108. NT = not toxic at 1,090ppm; Un = nitrogen unavailable; SI = nitrogen slightly available; - = not tested or non-nitrogenous.

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mM. At the 0.01 mm concentration, the followingaromatic acids were found to be approximately asinhibitory as coumarin: melilotic, phenylpropi-olic, hydrocinnamic, piperic, tetrahydropiperic,salicylic, and 2-furanacrylic. At the same concen-tration, trans-cinnamic, benzoic, and anthranilicacids were less inhibitory than was coumarin. Inthe fatty acid series, formic, propionic, butyric,caproic, capric, and crotonic acids were more in-hibitory than acetic acid at the 0.25 mm level.Succinic, fumaric, L-malic, and citric acids showedrelatively little toxicity.Audus and Quastel (4) tested the effect of a

number of amino acids and amines on the growthof cress roots. Glutamic acid had no toxic effect.Tryptophan and nicotinic acid were definitelytoxic at a concentration of 10 ppm in aqueoussolution. A number of amino acids and amineswere toxic to plant growth at a concentration of100 ppm. Riker and Gutsche (107) also found thata number of amino acids inhibited the growth ofsunflower tissue.

Routley and Sullivan (108) investigated a largenumber of organic substances for possible toxicityto ladino clover seedlings grown in sand culturefor 30 days (Table 4). In general, amino acids andcompounds related to nucleic acids were of lowtoxicity, and most of them were able to supplynitrogen to the seedlings. Several alkaloids, anti-biotics other than penicillin, and several of thecompounds included in the miscellaneous groupwere highly toxic.Norman (93) showed that elongation of cucum-

ber roots was inhibited by antibiotics. The follow-ing antibiotics, in concentrations of 0.2 to 8.5,4g/ml, produced 50% inhibition of root elonga-tion: L-cycloserine, azaserine, sulfocidin, magna-mycin, oligomycin, oxytetracycline, and D-cyclo-serine. For gramicidin, tyrothricin, thiolutin, andduramycin, concentrations of 14 to 39 Ag/ml wererequired, and for novobiocin, bacitracin, and van-comycin, concentrations of 140 to 510 ,ug/ml wereneeded to produce 50% inhibition in root elonga-tion.

Brian (18), in a review of the effects of 38 anti-biotics on plant growth, stated that most of theantibiotics tested exhibited some degree of phyto-toxicity. Four of the antibiotics-cycloheximideazaserine, alternaric acid, and polymyxin-in-hibited seed germination and root growth at con-centrations of 5 jAg/ml or less. One of the leasttoxic antibiotics was penicillin. Antibiotics may

inhibit germination, root growth, shoot growth,and production by plants of foliage, pigment, andplastid.

Barton and MacNab (6) determined the effectsof polymyxin, potassium penicillin G, strepto-mycin trihydrochloride, oxytetracycline hydro-chloride, and thiolutin on growth of wheat roots.The antibiotics were added to roots growing indistilled water, tap water, and nutrient solution.In concentrations of 1 to 0.1 units per ml, most ofthe antibiotics stimulated growth, but, in higherconcentrations of 1 to 32 units per ml, most ofthe antibiotics showed toxic properties.Wright (141) tested the effects of a large num-

ber of antibiotics on germination and root growthof wheat, white mustard, and red clover. The leasttoxic antibiotics were griseofulvin, penicillin, andstreptomycin. The more toxic ones were alternaricacid, glutinasin, mycophenolic acid, and glio-toxin. The range of effective concentration of anti-biotics was 1 to 25 ppm. In most cases, little or noeffect on germination was noted. However, anti-biotic concentrations that severely inhibited rootgrowth generally also reduced germination per-centage. None of the antibiotics was as toxic asindolylacetic acid, but most were more toxic thancoumarin. High concentrations (3,000 ppm) ofstreptomycin retarded chlorophyll formation andcotyledons were completely yellow. Alternaricacid at a concentration of 1 ppm inhibitedchlorophyll synthesis in mustard seedlings. Wheatwas less susceptible to the antibiotics than wasclover or mustard.

If conditions of light, temperature, and mois-ture are favorable and if mineral nutrients areavailable in sufficient quantities, plants of manyspecies can grow very well in the absence of addedorganic compounds. However, if these plants aregrown in the presence of vitamins, amino acids,antibiotics, alkaloids, or many other organic com-pounds, the organic substances may be absorbedand may influence plant growth, as indicated byKrasil'nikov (65, 66) and others (2, 21, 28, 91).

Organic Acids and Amino Acids in SoilSince many organic and amino acids influence

plant growth, it is interesting to know whetherthese compounds occur naturally in soil. Schwartzet al. (115), in a study of Ohio soils, found thatsignificant amounts of acetic and formic acidsand small quantities of other acids, includingcrotonic and lactic, could be separated from the

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soil. These compounds were probably derivedfrom a group of fatty acids of high molecularweight.Putnam and Schmidt (106) reported small but

detectable amounts of free amino acid in the soil.Soil amended with glucose and inorganic nitrogenyielded a wider variety and greater quantities offree amino acids than did untreated soil. How-ever, under usual soil conditions it is unlikely that

TABLE 5. Amino acids in acid hydrolysates of aHoytville (Ohio) soil*

Amt (mg/ Amt (mg/Amino acid 100 g of Amino acid 100 g of

soil) soil)

Cysteic acid. 11 a-Amino-n-N-acetyl gluco- butyricsamine......... P acid....... P

Glucosamine + Valine....... 81Galactosamine P MethionineAspartic acid 182 sulfoxide 6Serine ........... 182 Proline...... 48Threonine ....... 80 Methionine -Glutamic acid.... 137 Isoleucine ...33Methionine sul- Leucine. 58

fone.10 Ornithine 35Hydroxyproline 33 CystineGlycine.......... 80 Lysine.69Alanine.......... 109 Histidine.... 13fl-Alanine ........ 10 Tyrosine.. 17Ty-Aminobutyric Arginine 12

acid .......... 2 Phenylal-a,e-Diaminopi- anine...... 34

melic acid...... P

* This table appeared first in Ohio Agr. Expt.Sta. Circ. 61:1-18, 1958. See reference 145. Sym-bols: P = provisional identification; + = con-stituent identified but quantity not estimated;- = not detected.

free amino acid concentrations will become suffi-ciently high to be toxic to plant growth.A number of workers [Young and Mortensen

(145); Stevenson (127); Paul and Schmidt (98);Payne et al. (99); and others] have isolated aminoacids from the soil. The kinds and amounts ofamino acids found in an Ohio soil, as determinedby chromatographic methods, are shown inTable 5. Stevenson (127) found 29 ninhydrin-positive compounds in Illinois soils.

In view of the widely followed procedures ofusing strong acids and bases or other strongchemicals as extracting agents, the question might

be raised whether significant amounts of freeorganic or amino acids exist in the soil or whetherthese compounds are produced as a result of theextraction procedures used.

Influence of Carbon Dioxide, Bicarbonates, Nitrite,Hydrogen Sulfide, and Ammonia on Plant GrowthIn the decomposition of plant and animal resi-

dues in the soil, microorganisms produce largequantities of carbon dioxide. Stolwijk and Thi-mann (129) showed that root growth of Pisumsativum, Vicia faba, Phaseolus vulgaris, andHelianthus annuus was completely inhibited ifthe root medium was aerated with air containing6.5% carbon dioxide. Avena sativa and Hordeumvulgare roots were inhibited only slightly by this

TABLE 6. Carbon dioxide and oxygen concentrationsat 6-in. depth of soils sampled at

Cornell University, 1940*

Light silty clay loam Silty clay (under(under light sod) manure mulch)

Datet

C02 02 C02 02

% %0 % %12 July......... 3.5 17.2 6.1 11.615 July......... 3.9 16.4 5.1 13.622 July......... 2.4 18.3 4.1 16.45 August ....... 1.0 19.8 2.8 18.7

* This table first appeared in Cornell Univ. Agr.Expt. Sta. Bull. 763:1-43, 1941. See reference 15.

t Soil was wet at beginning of sampling periodand began to dry toward the end of the period.

treatment. Root growth of peas was slightly butconsistently stimulated in a root atmospherecontaining 0.5% carbon dioxide and was in-hibited at a carbon dioxide concentration of 1.5%.Chang and Loomis (24) found that water ab-

sorption by roots of wheat, corn, and rice plantsgrowing in water culture was reduced 14 to 50%by bubbling carbon dioxide through the solution.Plant growth was adversely affected by reducedwater absorption.

Taylor and Abrahams (134) of Ohio, and Boyn-ton (15) at Cornell University, found carbondioxide concentrations of 1 to 6%7o in the surface 6in. of soil (Table 6). Waksman and Starkey (137)reported carbon dioxide concentrations rangingfrom traces in aerobic soils to 20% in anaerobicsoils. Thus, in compacted soil with a high level ofenergy material and microbial decomposition,

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carbon dioxide concentrations probably approachlevels detrimental to plant growth. Bicarbonatesappear to be a factor contributing to the develop-ment of iron chlorosis (22).Although nitrites even at a low concentration

are toxic to plants, under most soil conditionsnitrite accumulation does not reach toxic levels.In Arizona, however, nitrite accumulation ap-parently causes "crazy top" of cotton. Scott (116)indicates that symptoms of this disorder are non-dehiscent anthers and apical branches which haveshortened internodes and varying degrees of ab-normal leaf size, shape, and arrangement. Thecondition was recognized in 1919 in the Salt RiverValley of Arizona, and it appears to be associatedwith soils having a high pH and an accumulationof nitrites. Applications of urea or choline chlo-ride temporarily corrected the effects of "crazytop" on anthers, so that pollen was released andnormal reproductive processes were completed.

Koike (63) showed that nitrite accumulation inHawaiian soils was negligible in acid soil (pH4.3). However, nitrites may accumulate undersome conditions, e.g., in alkaline soils and highammonium levels (1). In a neutral soil (pH 6.8)nitrite nitrogen accumulation exceeded 300 ppmwithin 4 weeks after the application of 1,500ppm of nitrogen in the form of NH40H. On theother hand, in an alkaline soil (pH 8.0), 300 ppmof nitrite nitrogen accumulated in 1 week from500 ppm of N added as (NH4)2SO4 and NH40H,and oxidation to nitrates was complete in 4 weeks.

In water-saturated soils (e.g., under conditionsof prolonged flooding) or in excessively compactedsoils, anaerobic decomposition of sulfates or pro-teins may result in the formation of hydrogensulfide which, even in low concentrations, is toxicto plants. Neilson-Jones (89) noted the presenceof hydrogen sulfide-producing bacteria in a heathsoil at Wareham Forest, and concluded thathydrogen sulfide was responsible for the difficultyencountered in growing trees. Alexander (1)indicated that many bacteria, fungi, and actino-mycetes can convert sulfates and partially re-duced sulfur compounds to sulfides.

Free ammonia is a cell toxin and, therefore,toxic effects on plants would be expected to resultif gaseous ammonia concentration became suffi-ciently high in the soil. In solution culture ofplants, toxic effects of ammonia may be found,but it is doubtful that the level of free ammoniain most soils is sufficiently high to cause toxicity.

Production of Phytotoxins by Soil Microorganismsand Plant Pathogens

Soil microorganisms produce a tremendousvariety of organic substances during the decom-position of plant and animal residues, and, asnumerous studies have shown, some of these sub-stances are phytotoxic. For example, Swaby (132)found that, when soil microorganisms were pres-ent in association with plant residues (lucerne andPhalaris tuberosa), substances inhibitory to plantgrowth were frequently produced. Hata (50)studied the influence of culture filtrates from3,300 isolates of soil microorganisms on thegrowth of Chlorella. He found that 81 isolates(2.5% of the total) produced growth-acceleratingfiltrates, 567 (17.2%) produced growth-inhibitingfiltrates, and filtrates from 2,652 (80.4%) hadno effect. Curtis (30), on the other hand, foundthat none of the culture filtrates from 500 actino-mycetes was inhibitory to plant growth.Krasil'nikov (66), in a study using 1,500 culturesof actinomycetes, indicated that 5 to 15% pro-duced substances inhibitory to plant growth.

Krasil'nikov (66) investigated more than 300cultures of nonsporeforming bacteria for theproduction of plant growth inhibitors. About100 of the cultures suppressed plant growth andgermination to some degree. Products fromcertain strains of Pseudomonas fluorescens, P.pyocyanea, and Bacterium sp. possessed strongherbicidal properties. More than 560 cultures ofsporeforming bacteria were tested for theirability to inhibit germination and plant growth.Of these cultures, 176 strongly suppressed thegermination of clover seeds and more than 200suppressed the germination of peas. Approxi-mately 20 to 30% of the sporeforming culturespossessed growth-inhibiting properties. Fourspecies, Bacillus mesentericus, B. subtilis, B.cereus, and B. brevis, accounted for most of theinhibitory spore formers. Among bacteria iso-lated from turfy podzol soils, 34 to 45% of theisolates produced plant growth inhibitors.

Krasil'nikov also observed that some strainsof actinomycetes caused chlorosis of plants suchas corn and that fungi of the genus Fusariumcaused chlorosis of grapevines (66). In otherstudies, Hodgson et al. (54) found that filtratesfrom crown gall bacteria produced wilting intomatoes.At Lincoln, Neb., McCalla et al. (83) found

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that, of 91 fungi isolated from stubble-mulchedplots where plant growth was reduced, 14 pro-duced culture filtrates that reduced by 50% ormore the germination of corn seeds soaked there-in. In another test, corn seedlings were grown onfilter paper soaked in the autoclaved culturefiltrates from fungi isolated at random fromplatings of soil representing stubble-mulchedplots at Lincoln, Neb. Of 318 filtrates tested, 52reduced shoot growth by 50% or more, and 167reduced root growth by a comparable percentage.

Gibberella fujikuroi. Stowe and Yamaki (131)and Stowe et al. (130) discussed the history of thegibberellins and the relationship of these com-pounds to the bakanae (foolish seedling) effect, aserious disease of rice. According to these authors,the Japanese plant pathologist, Sawada, in 1912suggested that a fungus might be the cause ofthe bakanae effect. More recently, it has beenestablished that this disease is caused by G.fujikuroi (San.) ver., and that this fungus pro-duces gibberellins which stimulate stem elonga-tion in the rice plant, resulting in the tall growththat is characteristic of the diseased plants (130,131). Severe infection with this fungus leads toadventitious root formation at the aerial nodes,stem curvature at the nodes, leaf curl, root rot,and death of the plants before flowering. Temper-ature is an important factor in the bakanae effecton rice. The optimal temperature for growth ofthe organism is 27 C, infection occurs best at 30 C,and the bakanae effect is most pronounced at35 C.

Brian et al. (20) indicated that wheat grownin weak nutrient solution and treated withgibberellic acid became chlorotic. This chlorosiscould be largely overcome by increasing theconcentration of the nutrient solution. Morganand Mees (85) reported similar results withpasture grass. In this case, the chlorosis could beovercome by the addition of nitrogen fertilizerwith the gibberellic acid. Although gibberellinscan induce stem elongation, very high concen-trations of these substances are inhibitory togrowth. "Overgrowth" in plants other than ricedoes not seem to occur naturally, but artificialinfection with the bakanae organism leads toovergrowth in corn, millet, wheat, and oats (70,131).Stowe et al. (130) found that G. fujikuroi also

produced a growth-deterring substance, fusaricacid, (5-n-butyl picolinic acid), which they ex-

tracted from culture solutions with benzene orpetroleum ether. They suggested that growthretardation sometimes observed in rice infectedwith bakanae organisms was due to fusaric acid.This compound, now a patented plant growthinhibitor and wilting agent, is effective in concen-trations as low as 0.1 mg per liter. Its effect seemsto depend upon its chelating properties-it is astrong inhibitor of heme-containing enzymes. Itsactivity may be partially reversed by the additionof minor elements.

Penicillium. A number of species of the genusPenicillium produce substances toxic to plantgrowth. Mirchink (84), in a study of soil toxicityat the Agrobiological Station in Chashnikov(Moscow area, Khimkin sector), isolated anumber of fungi, including numerous representa-

TABLE 7. Number of toxic fungal forms indifferent experimental variants*

No. of Percentageisolated of toxic

Experimental variants strains No. toxic strains

Control (without fer-tilizer) .............. 105 34 32

Nitrogen .............. 172 63 38Lime + manure ....... 85 21 24Lime + manure +NPK............... 68 10 15

* This table appeared first in MicrobiologyUSSR (Engl. Transl.) 26:83-90, 1957. See reference84.

tives of the genus Penicillium, which producedphytotoxic substances, as shown in Tables 7, 8,9, and 10. Liming of the soil and the addition ofmanure reduced the percentage of toxic forms.The most toxic fungi were P. cyclopium West.,P. nigricans Thom. (Bain.), P. paxilli Bain., andP. janthinellum Biour. The phytotoxic materialswere formed in the soil upon the addition ofsmall amounts of organic matter and inoculationwith an appropriate strain of Penicillium.

Curtis (31) found that, when P. thomii Maireculture filtrates were placed on the leaves ofbean and corn, the leaves became chlorotic,slightly necrotic, and were smaller than the un-treated leaves. Norstadt and McCalla (94), in astudy of factors causing reduction of crop yieldson stubble-mulched soil, isolated P. urticaeBainier from a stubble-mulched plot where plantgrowth had been poor. 'When sterile soil was

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TABLE 8. Action of culture fluid from highly toxicforms on wheat seeds*

Strain

C 1/8

D II/5C II/10

D II/4

II/14

II/35

C VIII/81

C VI/4

D IV/4

Control (nutrientmedium)

Control (water)Penicillium cyclo-pium Weste

P. paxilli Bain.P. janthinellum

Biour.P. martensii

Biour.P. nigricans Thom

(Bain.)P. nigricans Thom

(Bain.)P. nigricans Thom

(Bain.)Trichodermakoningii

Fusarium sp.

Germi-nated

seeds (%of con-trol)

100

1000

5474

74

100

87

90

68

68

2.61.5

3.0

1.0

0.6

1.0

3.6

4.5

* This table appeared first in MicrobiologyUSSR (Engl. Transl.) 26:83-90, 1957. See referer-ence 84.

TABLE 9. Action of soil containing different con-centrations of sucrose and infected with a culture

of Penicillium cyclopium on the germinationand growth of wheat seeds*

ScoeGerminatedLeghoVariants Sucroed seeds (9c of LenguthsoVari control)

% ~~~cm

Control (sterilized non-infected soil) ...... 100 5.5

P. cyclopium .......... 1 5 0.5P. cyclopium .......... 0.5 20 2.0P. cyclopium .......... 0.2 40 0.2

* This table appeared first in MicrobiologyUSSR (Engl. Transl.) 26:83-90,1957. See reference84.

supplemented with sterile plant residues, inocu-lated with P. urticae Bainier, and incubated for2 weeks, wheat seeds planted in this soil showedreduced growth in comparison with untreatedcontrols (8). The active growth inhibitor wasisolated and identified as patulin. Barnum (5)

demonstrated that P. expansum Link. producedsubstances that caused wilting of Malva rotundi-folia L., vetch (Vicia gigantea Hook), mint(Mentha sp.), cauliflower (Brassica oleraceae var.Botrytis), and alfalfa (Medicago sativa L.).

Aspergillus. Curtis (32, 33) observed curvatureof corn roots treated with A. niger culturefiltrates. Other malformations consisted of greatlythickened stems and petioles, and tumorlikestems. The active substance was isolated anddesignated as malformin. It is a neutral peptidecontaining four amino acids, valine, leucine,

TABLE 10. Distribution of highly toxicfungalformsamong field plots treated with fertilizers*

Experimental No. ofvariants Species of fungus strains

Control Penicilliumcyclopium Weste 2P. nigricans Thom (Bain.) 2P. paxilli Bain. 5P. janthinellum Biour. 10

Nitrogen P. nigricans Thom (Bain.) 3P. janthinellum Biour. 20P. paxilli Bain. 1P. martensii Biour. 6

Manure + P. janthinellum Biour. 1lime P. nigricans Thom (Bain.) 2

P. paxilli Bain. 1

Manure + P. janthinellum Biour. 1lime + P. nigricans Thom (Bain.) 1NPK P. paxilli Bain. 1

* This table appeared first in MicrobiologyUSSR (Engl. Transl.) 26:83-90, 1957. See reference84.

isoleucine, and one-half cystine. Takahashi andCurtis (133) suggested that malformin is a cyclicpeptide with the molecular formula C23H39N505S2.The infrared spectrum indicated strong adsorp-tion at 3.01, 6.06, and 6.54,u. Postlethwait andCurtis (103) found that bean seedlings treatedwith a culture filtrate of A. niger exhibited severecurvature, increased diameter, reduced length,and irregular protrusions of stem and leaf petioles.

Trichoderma and Fusarium. Mirchink (84)found that the culture filtrates of species of thegenera Fusarium and Trichoderma were lessphytotoxic than were species of the genus Peni-cillium. Fahmy (40) showed that F. solani pro-

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duced a toxic substance capable of causing wiltingof plants.

Alternaria. Ryan et al. (109) observed chloro-phyll deficiency in citrus seedlings grown fromseeds that had been in contact with cultureextracts of the A. tenuis during germination. Theactive material also inhibited chlorophyll forma-tion in the seedlings of various other plant species.

Other microorganisms. Phytotoxins are pro-

toxic substances were soluble in water and in-soluble in petroleum ether; stable to acid, heat,and storage at -3 to 1 C; and were precipitatedby alkali. The substances were most active in thepH range of 4.5 to 5.8, were nonspecific in actionon the plants tested, and were antifungal in someinstances. The inhibitory materials apparentlywere much more abundant in a water-saturatedsoil than in a soil at field capacity (Table 12).

TABLE 11. Effect of plant maturity and period of decomposition on relative toxicity of the resulting productson respiration of 6-day-old tobacco seedlings*

Decomposition period (days) at 60 to 70 FAge of plant material added to soil Type of plant

0 5 10 15 20 25 30

Young (5 to 6 weeks after Soil only -t - _ _ _ _planting) Timothy - + ++ +++ ++ _

Rye - - + ++ + _Corn - - + ++ + _Tobacco - - + + _ _

Intermediate (6 to 8 weeks after Soil only - - _ _ _ _planting) Timothy - + +++ +++ ++ +++ ++

Rye - + ++ +++ +++ ++ ++Corn - - ++ +++ ++ +++ ++Tobacco - - + +++ ++ _

Mature (10 to 14 weeks after Soil only - - - _ _ _planting) Timothy - - - ++ ++ +++ +++

Rye - - - + ++ +++ ++Corn - - - + ++ +++ +++Tobacco - - - + ++ +++

* This table appeared first in Can. J. Botany 36:621-647, 1958. See reference 96.t Toxicity rating: - = nontoxic (extracts which inhibit respiration of 6-day-old tobacco seedlings

by less than 19%) +, ++, +++ = mild, intermediate, and high toxicity, producing inhibition ofrespiration of 20-40%, 41-60%, and 61-95%, respectively.

duced by a large variety of microorganisms inaddition to those listed here. No attempt hasbeen made to present a complete listing of theseorganisms in this review.

Phytotoxic Substances in Decomposing PlantResidues, Soil Organic Matter, and Soil

Patrick and Koch (96) observed inhibition ofrespiration in tobacco seedlings caused by thedecomposition of timothy, corn, rye, and tobaccoplant residues in soil (Table 11). Germinationand growth also were adversely affected by thesesubstances. These toxic materials resulted whencrop residues were decomposed for 10 to 30 daysin wet soil at pH levels below 5.5. The highesttoxicity was obtained with residues of timothy,followed by corn, rye, and tobacco residues. The

Patrick et al. (97) conducted field studies in theSalinas Valley, Calif., which revealed that injuryto roots of lettuce and spinach seedlings was onthose parts of the roots in direct contact withdecomposing plant residues. Water-soluble sub-stances inhibiting the growth of seedlings wereextracted from plant residues decomposing infield soils. Phytotoxicity was most severe afterbarley, rye, wheat, sudangrass, vetch, broadbeam, and broccoli residues had decomposedfor 10 to 25 days. The toxicity decreased withincreasing periods of decomposition. After 30days of decomposition, stimulation of growth wasoften obtained.

Toxic substances may also be found in soilorganic matter. Kononova (64) and Krasil'nikov(65, 66) have shown that a number of growth-

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inhibiting substances are produced during thebreakdown of organic matter. Vanillin, benzoicacid, dihydroxystearic acid, and some aldehydesare phytotoxic substances of this sort.

Phytotoxic substances can be isolated from the

TABLE 12. Oxygen uptake of tobacco seedlings asaffected by the products of decomposition of

plant residues in soils held at satu-ration and at field capacitya

After 20-day decomposition period at

Plant material added Saturation Field capacityto s oiI_

pH rangeb2Os UP- pH range 2 up-prnetakec takep

Soil only ......... 6.4-6.6 128 6.4-6.6 131Timothy . . ..........4.8-5.2 18 5.9-6.4 90Rye ............ 5.3-5.8 45 6.0-6.6 104Corn ............ 4.9-5.4 28 6.0-6.5 99Tobacco .......... 5.5-5.9 68 6.9-7.5 112Control 125 (48)d

a This table appeared first in Can. J. Botany36:621-647, 1958. See reference 96.

bAfter the 20-day decomposition period; pH ofeach extract was adjusted to 5.3 prior to testing.

'r02 uptake, in ~Aiters of 02 (after 6 hr at 20 C),of 50 Harrow Velvet tobacco seedlings. In eachinstance, the tobacco seedlings were exposed for 16hr to the various extracts, then returned to phos-phate buffer (pH 5.3) before their 02 uptake wasdetermined. Each figure is based on the averageof four different decomposition series, for each ofwhich three determinations of four replicates weremade.

d 02 uptake, in jiliters Of 02 (after 6 hr at 20 C),of comparable seedlings pre-exposed (16 hr) to 0.01M phosphate buffer solution; based on the averageof 12 determinations.

soil. Mirchink (84), in a study of toxicity inturf-podzol soil, found that the water extract ofsoil had a depressing effect on germination andsprout length of wheat seedlings (Table 13).Extracts of soil to which lime and completemineral fertilization had been applied did notproduce this toxic effect.McCalla et al. (83) detected water-soluble soil

constituents which reduced root and shoot growthof wheat seedlings. Also, substances inhibitory tothe growth of roots and shoots of wheat seedlingswere isolated from methanol and acetone ex-tracts of stubble-mulched soil. Sodium pyro-phosphate extracts of soil from stubble-mulchedplots at Lincoln, Neb., contained a phytotoxicsubstance. The use of paper, column, andgas chromatography permitted a separation ofthe phytotoxic substance from the crude sodiumpyrophosphate extract. The substance was aneutral compound and had an RF value similarto that of patulin.The foregoing examples amply demonstrate

that, under appropriate conditions, phytotoxicsubstances are produced during decompositionof crop residues, and that phytotoxic substancescan be isolated from the soil.

Phytotoxic Substances in Forest Soils

Forest soils containing an abundance of organicmatter provide a favorable environment for rapiddecomposition. In such an environment, nu-merous organic substances are formed. Per-sidsky and Wilde (100) found that some of thevolatile substances released during decompositionof forest litter stimulated growth of excised rootsof blue lupine, but others were toxic.

Neilson-Jones (89), in a study of low produc-tivity of trees on Wareham Forest soils, observed

TABLE 13. Action of soil extracts on germination of wheat seeds*

7/VII 21/VII 7/VIII

Treatment Seed germi- Length of Germinated Germinated Length ofnation (% sprouts in seeds (% seeds (% sprouts inof control) cm (avg) of control) of control) cm (avg)

Control (water).100 5.6 100 100 3.2Control (without fertilizer).87 4.6 79 71 2.6Nitrogen.95 4.5 76 79 2.8Lime + manure.95 5.0 84 82 3.3Lime + manure + NPK.100 4.9 100 89 3.2

* This table appeared first in Microbiology USSR (Engl. Transl.) 26-83-90, 1957. See reference 84.

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the development of antifungal substances ofbiological origin. The effect on tree growth wasthought to result from inhibition of mycorrhizaformation and consequent restriction of rootgrowth and nutrient intake. In the laboratory,the toxic condition was developed in a soil witha high moisture content. Low temperature didnot seem to be a factor in the development of thetoxic condition. Autoclaving of the soil or treat-ment with alcohol, ether, or toluol removed theinhibitory factor.

mulching leads to (i) reduced nitrate production;(ii) lowered soil temperatures in the spring (thismay be detrimental to such crops as corn); and(iii) increased difficulty in the control of weedsand in other cultural operations requiring the useof machines. Even when measures are taken tooffset these difficulties (such as addition ofnitrogen, delayed planting of corn in the springto minimize the temperature effect, and goodcontrol of weeds), crop yields may still be inferiorunder the stubble-mulch system. An additional

FIG. 2. Stubble-mulchedfield in western Nebraska. Little wind and water erosion occur with this amount ofsoil cover.

Phytotoxic Effects Encountered in StubbleMulching

Stubble mulching, the use of crop residues onthe surface of the soil, is a very effective methodfor protecting the land against erosion by windand water (Fig. 2). However, plant growth andsubsequent crop yields obtained with this systemof farming are frequently reduced in comparisonwith growth and yields obtained by conventionalplow farming, particularly in the more humidareas of the United States (80). A number offactors are involved in this reduction. For ex-ample, in comparison with plowing, stubble

factor contributing to the observed yield re-duction may be the presence of phytotoxic sub-stances in residues, in soil, and among themetabolic products of soil microorganisms. As pre-viously indicated (49, 81, 82), most crop residuesappear to contain phytotoxic substances. Also,phytotoxic substances have been isolated fromstubble-mulched soil with extraction proceduresinvolving water, methanol, acetone, and sodiumpyrophosphate (83), and many soil microor-ganisms produce phytotoxic substances (e.g.,patulin) when grown in pure culture (94). Studiesare under way at Lincoln, Neb., to isolate andidentify the phytotoxic substances in the soil

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TABLE 14. Influence of organic compoundson the growth of tobacco seedlings*

Avg total Appear-Control or compound (200 mg/liter) yield ancet

mg

Control ........................ 20.1 S, 8DL-Alanine ..................... 0.4DL-a-Amino-n-butyric acid...... 0D-Arginine hydrochloride ....... 18.3 S,8L-Asparagine................... 0.5 R,0L-Aspartic acid ................. 2.0 R,6L-Cystine ....................... 15.6 S, 8D-Glutamic acid ................ 0.6 R,2Glutathione .................... 1.4 R, 2Glycine ........................ 0.5 R,1L-Jlistidine dihydrochloride..... 4.8 R,8L-Hydroxyproline .............. 0 .OL-Leucine ...................... 9.2 S,8DL-Isoleucinet.................. 1.8 R,4D-Lysine dihydrochloride ....... 2.0 R,4DL-Methionine.................. 4.8 S,8DL-Norleucine ................. 0.5 R,2DL-Ornithine hydrochloride. ... 13.9 S,8DL-4-Phenylalanine............. 10.1 S,8L-Proline ....................... 0.1 R,2DL-Serine....................... 0DL-Threonine ................... 0.2 R,2L-Tryptophan ................... 0.6 R,2L-Tyrosine..................... 10.4 S,8DL-Valine ...................... 0.3 R,2

* Maryland Medium Broadleaf, with 2% ofsucrose plus various amino acids in aseptic culturefor 28 days at 25 C and 500 ft-c of fluorescent whitelight for 8 hr daily. This table appeared first in J.Agr. Res. 75:81-92, 1947. See reference 125.

t S, shoot or stem; R, rosette. Rated from0 (white) to 10 (dark green).

t Seedlings grown with DL-isoleucine showed in-hibition in growth of apical bud, suckering, andstrap leaves; all are typical symptoms of french-ing.

obtained from stubble-mulched plots on whichreduced plant growth has been observed.

SOME SPECIFIC PLANT EFFECTS POSSIBLYCAUSED By TOXINS FROM

MICROORGANISMS

Legume Bacteria Factor in the Chlorosis ofSoybeans

Erdman et al. (36, 37) and Johnson et al. (58,59, 60) reported a bacterial-induced chlorosis ofsoybeans. Some soybean varieties, such as Lee,

were highly susceptible. Apparently, duringnodulation certain strains of rhizobium producedsubstances which inhibited growth and producedchlorosis of the susceptible varieties. There wassome indication that chlorosis and growth inhi-bition were caused by different substances. Ex-tracts of nodules from chlorotic soybean plantsinoculated with chlorosis-inducing Rhizobiumjaponicum strain 76 retained their inhibitoryactivity after autoclaving for 25 min at 120 C andstorage at room temperature for several months.In tests involving applications of the extract to37 species other than soybeans, severe chlorosiswas observed in 7 species.

Frenching Disease of TobaccoValleau and Johnson (136) described "french-

ing" of tobacco as primarily a chlorosis of thetissue between the larger veins in the leaves nearthe growing point of the plant. Because theaddition of nitrate nitrogen alleviated the symp-toms, these investigators believed that nitrogendeficiency accounted for the disease. The additionof lime or heating the soil to 65 C also caused thesymptoms to disappear.

Steinberg (124) grew Xanthi Turkish tobaccoseedlings aseptically on 50 ml of a mineral-agarsolution containing 200 ppm of Peptone (Difco)in 200-ml Erlenmeyer flasks at 25 C. About 60different species and strains of soil bacteria wereintroduced into tobacco cultures by stab inocula-tion near the seedling stems. Abnormal growthof various types resulted in several of the cultures.Included were chlorotic effects similar to thoseencountered in various mineral deficiencies;epinasty; cupped, narrow, and strap leaves; andleaves with lobes, hooked tips, and rim roll. Someof the effects were similar to typical frenchingsymptoms.

Steinberg (125, 126) also found that tobaccoplants grown in a nutrient solution supplementedwith various individual amino acids at a finalconcentration of 200 ppm exhibited stuntedgrowth and chlorosis to varying degrees (Table14). The effects produced by DL-isoleucine wereespecially similar to typical frenching symptoms.Steinberg suggested that "excesses of free aminoacid due to abnormalities in protein metabolismmay contribute to the formation of characteristicpatterns of symptoms in plants because of dis-turbances in mineral nutrition."

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Replant Problem in Peaches

The peach tree replant problem in Ontario,Canada, as described by Koch (62), is a complexone with probably no single dominant factorresponsible. When an old peach orchard is re-placed by young peach trees, many of the youngtrees become stunted in growth. The factorsinvolved probably include soil toxins, nematodes,fungi, bacteria, soil fertility, and possibly others(138). Replants that are most severely affectedare usually located at or very near the site of anold tree. For such replants, soil fumigation does

produced substances that inhibited the respira-tion of excised peach root tips (Tables 15 and16).

Replant Problem in Citrus

The citrus replant problem was studied ex-tensively by Martin and co-workers (71, 76, 77).In citrus groves affected by this condition,growth of replants is slow in comparison withgrowth of young trees planted in soil wherecitrus never has been grown. Martin (71, 72, 73,74) believes that the reduced growth is caused

TABLE 15. Influence of incubation time, soil type, and other factors on the relative toxicity of the resultingsoil leachates as determined by the intensity of their inhibiting effects on the

respiration of excised peach root tips*

Substances added to old peach orchard soil (site area)

Incubation time Peach root residue Amygdalin (0.25%) Peach soil autoclaved Peach root residue___h___at____25__ _(2%) added tobefore extracts Sour cherry "compost soil"were made root residue

2%t 5s Site area Intersite residue t Amygdalin (2%)soil area soil residu (0.25%)

60 48 - 51 --- -70 60 _ 55 8 10 ST 8 ST100 40 78 72 - 6 15 ST 18 ST150 68 82 7 ST 3 ST 7 ST 15 ST 25 ST200 - 59 73 19 ST 2300 - 74 14 12

* This table appeared first in Can. J. Botany 33:461-486, 1955. See reference 95. Results are expressedas per cent total inhibition (or stimulation) of respiration in 5 hr at 20 C of 15 excised peach root tipsplaced in the various soil-water leachates (containing varying amounts of the decomposition productsof the substances indicated). Per cent inhibition was calculated in the usual manner by using as check(or normal respiration rate) the respiration of identical root tip samples placed in soil-water leachatescontaining none of these products.

t Amount of peach root residues, amygdalin, or other root residues, calculated as per cent by weightof the total soil contents of the flask, added to each flask.

t If total 02 uptake was greater than that of the check, it was calculated as per cent total stimula-tion (ST) of respiration.

not completely eliminate stunted growth. Proeb-sting and Gilmore (105) demonstrated that atoxic factor is present in peach root bark. Later,Patrick (95) and Patrick and Koch (96) suggestedthat the growth inhibition is caused indirectly bythe cyanophoric ,3-glycoside, amygdalin, which ispresent in peach roots. Amygdalin itself is nottoxic, but when broken down it yields glucose,benzaldehyde, and hydrogen cyanide, andtoxicity results. The peach root and variousmicroorganisms contain enzymes which willbreak down amygdalin. Patrick (95) reportedthat microbial decomposition of peach roots

primarily by the development of plant rootparasites in the soil but that other factors, in-cluding nutritional deficiencies and imbalancesand deterioration of soil structure, influence theextent of injury caused by the detrimentalorganisms. Citrus seedlings in old citrus soilssupporting very poor growth of replants did notrespond to fertilization with P, K, Mg, Cu, B,Zn, or Mn. The poor growth occurred even whengood practices of fertilizer application, pestcontrol, and general management were used.Partial soil sterilization with such fumigants asD-D (a mixture of dichloropropane and dichloro-

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TABLE 16. Comparison between the intensity of the inhibiting effects on the respiration of peach root tipsproduced by decomposition products of peach root residues, amygdalin, and

other root residues as influenced by incubation time and soil type*

Substances added to old peach orchard soil (site area)

Incubation Peach root Pahrotime (hr) at Peachduoeot Peach soil autoclaved rSourrridue25 C before residue Sourr Tobacco Pepper (5%)extracts were Am~ygdai roo root root Calcium added tomade (0.25%) Peach root Am d residue residue cyanide muck soil1% 5% residue ygda5hn (2 (0% (0%(5%) (.5) u

40 63 66 10 ST 27 ST 36 ST 40 -60 68 76 12 4 ST 4 ST 670 - 84 47 5 12 ST 28 ST - 380 55 81 14 - 7 30 ST -90 43 90 65 3 ST 5 10 ST 29 7110 - 83 68 13 5 _

* This table appeared first in Can. J. Bot. 33:461-486, 1955. See reference 95. Results are expressedas per cent total inhibition (or stimulation) of respiration in 5 hr at 30 C. The method used to comparethe inhibiting or stimulating (ST) effects on the respiration of peach root tips produced by the differentleachates is identical to that in Table 15, the only difference being that the respiration studies werecarried out at 30 C on eight excised peach root radicals.

propylene), chloropicrin, carbon disulfide, andethylene dichloride usually resulted in improvedgrowth of citrus seedlings, but in early tests theseedlings were still inferior to those grown onnoncitrus soils. Some fumigation treatmentschanged the fungus populations drasticallywithout improving plant growth. Results ofexperiments in which fumigated soil was inocu-lated with various fungi indicated that fungalspecies differed considerably in their influence ongrowth of citrus seedlings (Table 17).Martin (74) used various leaching treatments

on old citrus soil and observed the growth ofcitrus seedlings subsequently planted in the soil.Leaching the soil with large amounts of distilledwater did not improve growth of the seedlings.However, complete recovery of seedling growthwas observed in soil leached with 2% H2SO4 or2% KOH followed by saturation of the soilcolloids with a favorable combination of Ca, Mg,K, and H. As a result of these observations,Martin (72, 74) suggested that toxic substancesoriginating from roots or from organisms de-composing roots might possibly be a contributingfactor to the replant problem. Certain fungi,primarily Fusarium solani, Pyrenochaeta sp., andFungus D1 were more numerous in old citrussoil than in noncitrus soil. The chemical com-position of citrus seedlings grown in some oldcitrus soil did not differ significantly from that of

TABLE 17. Effect offumigation and inoculation withspecies of soil fungi, separately and in various

combinations, on growth of sweet orangeseedlings in Yolo sandy loam (test 2)*

Soil treatment Dry wt of topst

gCheck (none).17Fumigated.24Fumigated and inoculated withPyrenochaeta sp.25Thielaviopsis basicola. 5Fusarium solani.24Pythium ultimum.21Pyrenochaeta sp., T. basicola, F.

solani, and P. ultimm.13

* Only old citrus soil used. This table appearedfirst in Soil Sci. 81:259-267, 1956. See reference 78.Published with permission of The Williams & Wil-kins Co.

t Average per 3-gal pot of soil. LSD 5% = 4 g.

seedlings grown in noncitrus soil. Old citrus soilswere not toxic to numerous other crops.Martin et al. (75) found an occasional

temporary growth inhibition of citrus seedlingsafter heat or fumigation treatment of soils in thegreenhouse or in field nursery beds. The toxiceffect lasted from a few weeks to 1 year. A sug-gested explanation for the phenomenon was thatthe inhibitory substance was produced by an

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organism which became dominant after theexisting population was destroyed by the treat-ment. The toxic substance continued to be pro-duced until the organism involved was reduced

dead roots added. Benedict suggested that inhib-itory substances accumulate in the soil or othermedium where bromegrass is grown. He believedthat such substances are excreted from living

.Iv

INK

N..

FIG. 3. Influence of stubble mulching on corn growth at Lincoln, Neb., May, 1958. (left) Typical stalkfromplowed plot; (right) stunted stalkfrom stubble-mulched plot.

in numbers and activity by antagonistic organ-isms.

Sod-Binding of BromegrassBenedict (9) observed growth depression of

bromegrass (Bromus inermis) grown in pure ormixed stands in sand culture. The degree ofdepression was related to the proportion ofbromegrass plants in the stand. Also, the additionof dead bromegrass roots to bromegrass growingin sand culture caused a depression in growth, theextent of which was related to the amount of

roots, are liberated from dead cells sloughed offfrom living roots, or are formed by the decom-position of these dead cells.

Stunting of Corn Associated wih StubbleMulching

In the stubble-mulch system of farming, corngrown in the humid northern areas of the UnitedStates seems to be particularly retarded in growth(80). At Lincoln, Neb., corn in a corn, oats, andwheat rotation is severely retarded about every3 or 4 years. The retardation is evident at the

I

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seedling stage, and the plants seem never torecover fully. The toxic effect is not uniform overthe entire stubble-mulched plot (Fig. 1). Only afew plants seem to be severely affected (Fig. 3).Such stalks at maturity may be only a foot talland may bear a small ear. The plants generallydisplay a chlorosis similar to that resulting froma deficiency of nitrogen or of some minorelements.

Seedling Disease of RiceThe bakanae effect, a seedling disease of rice

which results in abnormal rapid elongation ofstem, has long been known by the Japanese. Thisdisease is discussed in an earlier section of thisreview.

Inoculation Failure in Subterranean Clover

Hely et al. (51) found that in certain yellowpodzolic soils of granite origin in New SouthWales, Australia, it was difficult to obtain nodula-tion of subterranean clover. Evidence was ob-tained for the existence of a nodulation-inhibitingfactor that could be transferred to other soils.This factor was believed to have a microbiologicalorigin, and to prevent nodulation by antagonizingthe multiplication of the nodule bacteria in therhizosphere. The inhibitory effect could beeliminated by autoclaving the soil. The additionof charcoal with the seed at planting time im-proved nodulation.

CONCLUSIONS

A wide variety of phytotoxic substances existin plant residues and seeds. Included are suchcompounds as amino acids, alkaloids, certainessential oils, coumarin, unsaturated lactonessuch as parasorbic acid, phenols and aldehydes,organic acids, ammonia, hydrogen cyanide,mustard oils, scopoletin and scopoletin glyco-sides, and ethylene. Soil microorganisms alsoproduce many organic substances which aretoxic to plant growth. Examples are organic andamino acids, numerous antibiotics, and sub-stances such as gibberellin. Toxic inorganiccompounds, such as nitrites, carbon dioxide, andhydrogen sulfide, are also produced by soilmicroorganisms.Although green plants can live autotrophically,

it is apparent that. under suitable conditions theycan also live heterotrophically, absorbing organiccompounds from their surroundings and me-

tabolizing these compounds. Plants grown in thesoil are normally exposed to a tremendousvariety of organic compounds which have comedirectly from plant and animal residues in thesoil, or indirectly from these residues through theaction of soil microorganisms. Depending upontheir nature and concentration, and on the kindof the plant being grown, these substances maybe innocuous, stimulatory, or inhibitory to plantgrowth. Thus, the role of soil organic constituentsin plant growth is extremely complex, and presentunderstanding of this role is at best fragmentary,particularly for plants grown under field con-ditions. Nevertheless, it appears that under somechemical, physical, and biological conditions inthe field, phytotoxic substances are producedand can accumulate to such an extent that re-duced plant growth results.The presence of phytotoxic substances in plant

residues and soil, and the production of suchsubstances by microorganisms, may account inpart for adverse effects of a particular crop on asucceeding crop in crop rotations, yield reductionsencountered in stubble mulching, citrus and peachtree replant problems, sod binding in bromegrass,frenching of tobacco, seedling disease of tobacco,the legume bacteria factor in chlorosis of soy-beans, and inoculation failure with subterraneanclover.

OUTLOOK

Although some information is available on theoccurrence and nature of phytotoxic substancesin microorganisms and crop residues, muchadditional knowledge is needed. Apparently,phytotoxic substances of many different kindsare produced by a myriad of microorganisms andhigher plants. A knowledge of the chemicalnature of these phytotoxic substances is neededin order that the substances may be preciselytested, individually and in combination, for theireffects on various kinds of plants at all stages ofgrowth. The information obtained in such testswould be useful agronomically, for example, incontributing to an understanding of the resultsobtained with various crop rotations. Further-more, such information would be importantecologically, inasmuch as naturally occurringphytotoxic substances no doubt influence plantsuccession.

Phytotoxic substances from higher plants andmicroorganisms also would be expected to affect

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microbial ecology. Taxonomic information on thesoil microorganisms that produce phytotoxicsubstances and on those that are affected by suchsubstances is badly needed.The isolation and identification of new phyto-

toxic substances will undoubtedly result in thediscovery of some new compounds suitable foruse in the control and management of bothplants and microorganisms. Quite possibly someof these compounds may be at least as effectiveas presently used herbicides and fungicides, andthey may be free of some of the undesirableattributes of currently popular compounds. Thisarea of research is complex, and much work willbe needed before all aspects of the interaction ofthe phytotoxic substances with plants and micro-organisms can be understood.

LITERATURE CITED

1. ALEXANDER, M. 1961. Introduction to soilmicrobiology, p. 285 and 370. John Wiley &Sons, Inc., New York.

2. ANONYMOUS. 1940. Plant growth-substancesin soil and manure. Soils Fertilizers 3:243-244.

3. AUDUS, L. J. 1959. Plant growth substances,2nd ed. Interscience Publishers, Inc., NewYork.

4. AUDUS, L. J., AND J. H. QUASTEL. 1947. Toxiceffects of amino acids and amines on seed-ling growth. Nature 160:222-223.

5. BARNUM, C. C. 1924. The production of sub-stances toxic to plants by Penicillium ex-pansum link. Phytopathology 14:238-243.

6. BARTON, L. V., AND J. MAcNAB. 1954. Effectof antibiotics on plant growth. Contrib.Boyce Thompson Inst. 17:419-434.

7. BARTON, L. V., AND M. L. SOLT. 1948. Growthinhibitors in seeds. Contrib. Boyce Thomp-son Inst. 15:259-278.

8. BEHMER, D. E., AND T. M. MCCALLA. 1963.The inhibition of seedling growth by cropresidues in soil inoculated with Penicilliumurticae Bainer. Plant Soil 18:199-206.

9. BENEDICT, H. M. 1941. The inhibiting effectof dead roots on the growth of bromegrass.Agron. J. 33:1108-1109.

10. BENNETT, E. L., AND J. BONNER. 1953. Isola-tion of plant growth inhibitors fromThamnosma montana. Am. J. Botany 40:29-33.

11. BENTLEY, J. A. 1958. The naturally-occurringauxins and inhibitors. Ann. Rev. PlantPhysiol. 9:47-80.

12. BONNER, J. 1950. The role of toxic substances

in the interactions of higher plants. Botan.Rev. 16:51-65.

13. BONNER, J., AND A. W. GALSTON. 1944. Toxicsubstances from the culture media of gua-yule which may inhibit growth. Botan. Gaz.106:185-198.

14. BORNER, H. 1960. Liberation of organic sub-stances from higher plants and their role inthe soil sickness problem. Botan. Rev. 26:393-424.

15. BOYNTON, D. 1941. Soils in relation to fruit-growing in New York. XV. Seasonal andsoil influences on oxygen and carbon-dioxide levels of New York orchard soils.Cornell Univ. Agr. Expt. Sta. Bull. 763:1-43.

16. BREAZEALE, J. F. 1924. The injurious after-effects of sorghum. Agron. J. 16:689-700.

17. BRIAN, P. W. 1949. The production of anti-biotics by microorganisms in relation tobiological equilibria in soil. Symp. Soc.Exptl. Biol. 3:357-372.

18. BRIAN, P. W. 1957. Effects of antibiotics onplants. Ann. Rev. Plant Physiol. 8:413-426.

19. BRIAN, P. W. 1957. The ecological significanceof antibiotic production, p. 168-188. InR. E. 0. Williams and C. C. Spicer [ed.],Microbial ecology. Cambridge UniversityPress, New York.

20. BRIAN, P. W., G. W. ELSON, H. G. HEMMING,AND M. RADLEY. 1954. The plant-growth-promoting properties of gibberellic acid, ametabolic product of the fungus Gibberellafujikuroi. J. Sci. Food Agr. 5:602-612.

21. BRIAN, P. W., J. M. WRIGHT, J. STUBBS, ANDA. M. WAY. 1951. Uptake of antibioticmetabolites of soil microorganisms byplants. Nature 167:347-349.

22. BROWN, J. C. 1961. Iron chlorosis in plants.Advan. Agron. 13: 329-369.

23. CAMPBELL, A. G. 1959. A germination inhibi-tor and root-growth retarder in chou moel-lier (Brassica oleracea var.). Nature 183:1263-1264.

24. CHANG, H. T., AND W. E. LooMIs. 1945. Effectof carbon dioxide on absorption of waterand nutrients by roots. Plant Physiol. 20:221-232.

25. COCHRANE, V. W. 1949. Crop residues as caus-ative agents of root rots of vegetables.Conn. Agr. Expt. Sta. (New Haven) Bull.526:1-34.

26. COLLISON, R. C. 1925. The presence of certainorganic compounds in plants and their rela-tion to the growth of other plants. Agron. J.17:58-68.

27. COLLISON, R. C., AND H. J. CONN. 1925. Theeffect of straw on plant growth. N.Y. State

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Agr. Expt. Sta. (Geneva, N.Y.) Tech.Bull. 114:1-35.

28. COOPER, R. 1959. Bacterial fertilizers in theSoviet Union. Soils Fertilizers 22 :327-333.

29. Cox, L. G., H. M. MUNGER, AND E. A. SMITH.1945. A germination inhibitor in the seedcoats of certain varieties of cabbage. PlantPhysiol. 20:289-294.

30. CURTIS, R. W. 1957. Survey of fungi and acti-nomycetes for compounds possessing gib-berellinlike activity. Science 125:646.

31. CURTIS, R. W. 1957. Translocatable plantgrowth inhibitors produced by Penicilliurnthomii and Arachniotus trisporus. PlantPhysiol. 32:56-59.

32. CURTIS, R. W. 1958. Root curvatures inducedby culture filtrates of Aspergillus niger.Science 128:661-662.

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34. DORYLAND, C. J. T. 1916. The influence ofenergy material upon the relation of soilmicroorganisms to soluble plant food. N.Dakota Agr. Expt. Sta. Bull. 116:319-401.

35. ENGARD, C. J., AND A. H. NAKATA. 1947. Agrowth inhibitor and a growth promotor insugar cane. Science 105:577-580.

36. ERDMAN, L. W., H. W. JOHNSON, AND F.CLARK. 1956. A bacterial-induced chlorosisin the Lee soybean. Plant Disease Reptr.40:646.

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44. GOLDBERG, H. S. 1959. Antibiotics: theirchemistry and nonmedical uses, p. 356-365.D. Van Nostrand Co., Inc., Princeton,N.J.

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from microorganisms and crop · vol. 28, 1964 phytotoxic substances fromsoil microorganisms...

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