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Part IV, continued:

 

Significance of antagonists in plant immunity

  Microbial antagonists, an noted above, suppress theircompetitors with their metabolic products, among which a special place is occupiedby the antibiotics. The latter may accumulate not only in artificial nutrient mediabut also directly in the soil, being concentrated there in smaller or greater amounts.

  It is also known that plants can absorb various organicsubstances from the medium, including antibiotics, through their roots. If the antibioticsubstances enter the plants, it should be assumed that they may produce a certaineffect in the tissues, namely, suppress the activity of microbes that have penetratedthe cells and increase the toxicity of the cell sap and, consequently, also increasethe resistance of the plants to infections. In other words, an assumption is madethat the entry of antibiotics is reflected in the immunobiological properties ofthe plants.

  It should be noted that the problem of immunity inplants, regardless of the numerous studies done on the subject, is still very littleknown.

  Literature data show that immunity, nonsusceptibilityto infections in plants is determined by various, quite complicated internal andexternal factors.

  One of such internal factors is the toxicity to bacteriaof the call sap. It was noted long ago that the sap of plants possesses the abilityto suppress growth of bacteria and fungi. Wagner (1915) observed death of bacteriain sap from the tubers and stems of potatoes. Precipitating these juices with ammoniumsulfate and washing the precipitate with water, he obtained a substance of a proteinnature with strong antibacterial properties. The substance is thermolabile, decomposesquickly in light and under the influence of oxidases and peroxidases. The toxicityof potato juice was observed by Cholodnyi (1939), he found that the toxicity of thejuice increases upon sprouting of potato tubers.

  Antimicrobial properties were observed in the juiceof onions, corn, tomatoes, cotton, orchids and other plants. In the literature thereare descriptions of the inactivation of fungal toxins by the sap of plants resistantto the diseases (Yachovskii, 1935, Vavilov, 1919, Naumov, 1940, Carbone and Arnaudi,1937). There are indications, that the cell sap from varieties which are resistantto the infection is more toxic than that of susceptible varieties (Kramarenko, 1949;Gorlenko, 1950).

  The antimicrobial properties of vegetative juice areexplained by the presence of various substances in the cells.

  An opinion was voiced that plant immunity is causedby the presence of elements of mineral nutrition in the sap. Some of them, potassiumcopper, cadmium, etc create a nonfavorable milleau in the plant tissues for growthof microbes. According to some authors, they are favorable to the accumulation oforganic acid in the cells. Other investigators tried to explain the resistance toinfections by the osmotic pressure of the sap, by the presence of alkaloids, enzymes,agglutinins, and lysis, dissolving the microbial cells, etc.

  Israil'skii (1952) expresses the opinion, that thenonsusceptibility of plants to certain infections is caused by bacteriophages whichsaturate the tissues of the plants.

  Winter and Ruemker (1952) concluded that the nonsusceptibilityof plants is caused by the presence of a special "resistance factor" inthe roots.

  Many investigators ascribe considerable importancein plant immunity to the redox system (Rubin, 1050, et al.) or to the permeabilityof the protoplasm of the cells Kuprevich, 194 7, Kokin, 1948; Sukhorukov, 1952),Recently, a connection was noted between plant resistance to infections and the productionof pigments of the anthocyanin group.

  On the basis of him extensive studies Tokin (1951)attaches great importance to the phytocides in the immunity of plants. In phytocidesare included different chemical antimicrobial substances which are formed by plants.Essential oils and organic acids, aldehydes, alcohols, phenols and also various specificcompounds may be included here.

  One would assume that during the course of the historyof their development plants acquired various means of protection against infections,mechanical, chemical and biological.

  While ascribing a certain importance to all these,in our opinion, the investigators have not yet touched one of the most importantprotective factor" the antimicrobial antibiotics, formed in the soil by microbesand absorbed there from the plants.

 

Formation and accumulation of antibiotics in soil

  It was recently established that in the soil thereare antibiotic substances which are formed by the microbial antagonists. The formationof antibiotic substances by microbes in the soil was experimentally proved by manyinvestigators. Gottlieb and Siminoff (1952) have shown the presence in soil of chloromycetin,formed by Actinomyces venezuelae. The substance was isolated from the soiland chemically purified. Owing to favorable conditions of growth of the actinomycetes,25.0-27.8 mg/kg of chloromycetin accumulate in the soil. The formation of chloromycetinin the soil by A. venezuelae was also noted by Jefferis (1952). The greatestamount is formed upon the addition of peptone or lucerne hay to the soil. In thepresence of starch or oat meal the antibiotic is not formed.

  Gregory et al., (1952) observed the formation of antibioticsubstances in soil by cultures of bacteria--Bacillus sp. B-6, by actinomycetes--ActinomycesNo 67 and by fungi Pencillium patulum. In the presence of appropriate nutrientsources in soil (soy meal) the following amounts of antibiotics were formed:

 

In sterile soil, units

In nonsterile soil, units

Bacillus sp. B-6

300

30

Actinomyces sp. No 67

5-10

3

Penicillium patulum

100

10

  Hessayon (1951, 1953) showed the formation of the antibiotictrichothecin by the fungus Trichothecium roseum. Large quantities of the antibioticwere detected in loamy soil, and smaller quantities in sandy soils.

  According to Gottlieb (1952) the antibiotic actidioneis formed in the soil in considerable quantities (10 µg/g and more) in the presenceof soy meal. Grossbard (1940) has grown Penicillium patulum in the soil inthe presence of various sources of nutrition. The formation of patulin took placein the presence of beet cake, glucose, fresh wheat straw, timothy grass and certainother plant residues. According to this author, antibiotics are also formed in thesoil by Aspergillus terreus and A. antibioticus in the presence ofwheat straw and certain other sources of nutrition.

  In a later work Grossbard (1952) has shown the formationof antibiotics in the soil by the fungi: Aspergillus terreus--70 units; Penicilliumsp.--80 units; Aspergillus clavatus--110 units and Penicillium clavatus--100units in 1 g of soil.

  On the third day of growth of the fungus Aspergillusclavatus, in the presence of brown sugar, Gottlieb and Siminoff (1952) noticedthe formation of clavacin in the soil in the amount of 16 µg/g. Taking intoaccount the extent of adsorption and inactivation of the substance by soil particles,the authors determined its total production as not less than 50 µ g/g.

  Stevenson and Lochhead (1954) studied the formationof antibiotic substances by the fungus Penicillium sp. and by the actinomycetesK-1 and V-27 in sterile as well as in nonsterile soil. In sterile soil the fungusforms as much antibiotics as on Chapeks medium (about 20--30 µ g/g and in nonsterilesoil 3 times less (7 -l0 µ g/ g). The actinomycetes form up to 16 units/g andmore of the active substance. As sources of nutrition both organisms use variousplant residues--the green bulk of clover, orchard grass, brome grass and wheat.

  Wright (1955) observed the formation of griseofulvinin the soil by the fungus Penicillium nigricans in the presence of other soilfungi. According to his data, a pure culture of Penicillium nigricans formsthe following amounts of the antibiotic (in sterile soils), in podsol at pH 5.3--0.2µ g on the 7th day and 2.4 µ g on the 14th day, in podsol with Ca(OH)2--2.4µ g on the 7th day and 20 µ g on the 14th day; in orchard soil at pH 6.2--0.6µ g on the 7th day and 10 µ g on the 14th day--in l g of soil. Upon infectingthe soil with other fungi the production of griseofulvin either decreases or increases,depending on the species of the fungi. On the 14th day of incubation griseofulvinwas detected in the soil in the following quantities:

 

Amount in µ g

Penicill nigricans at 40 µ g/g (Control)

100

Penicill nigricans + Trichod. viride

3.2

Penicill nigricans + Trichod. viride

3.2

Penicill nigricans + Trichod. viride

0.4

Penicill nigricans + Mucor ramannianus

12

Penicill nigricans + Cladosp. herbrum

100

Penicill nigricans + Penic. expansum

1.5

Penicill nigricans + Penic. frequentans

0

Penicill nigricans + Penic. albidum

200

Penicill nigricans + Penic. stolonifer

200

  In sterile soil which is contaminated with nonsterilesoil griseofulvin accumulates much less, namely about 6 µ g/g.

  The author notes that in sterile and nonsterile soilsantibiotics are also formed by many other fungi. On the 14th day of incubation insoil infected by fungi he found the following quantities of antibiotics:

 

Antibiotic

In 1 g of soil, µ g

Trichoderma viride

gliotoxin

40

Trichoderma viride

viridin

0

Penicill. expansum

patulin

200

Penicill. frequentans

frequentin

20

Penicill. stolonifer

mycophenolic acid

0.16

Penicill. nigricans

griseofulvin

40

  Our studies show that antibiotic substances are formedin the soil by various antagonistic organisms--bacteria, actinomycetes and fungi,if the conditions are suitable.

  In podsol soil in the presence of nutrient sources(soy meal, lucerne or clover straw, sugar or other substances) bacterial antagonistsform 40-180 units/g in sterile soil and 10-80 units/g in nonsterile soil. (Table113). As can be seen from the table, the formation of antibiotic substances is moreintense in sterile soils. As an artificial nutrient, here too, the presence of specialsubstances is essential to the formation of antibiotics, and often those are notthe same substances as those necessary for the nutrition and growth of the antagonisticbacteria. Of many organic substances tested only some of them were suitable for thesynthesis of antibiotics, although growth of the antagonists occurred with any oneof the nutrient sources used in the experiments.

Table 113
Formation of antibiotics in soil by bacteria
(Units/ g on the 5th through 7th day of growth)
Bacteria Sterile soil, antibiotic formed Nutrient source used Nonsterile soil, antibiotic formed Nutrient source used
Bac. mesentericus 180 Starch, soy meal 80 Soy meal, meat-peptone
Bac. subtilis 120 Meat-peptone broth, soy meal 40 Meat-peptone broth
Ps. flourescens 40 Lucerne 10 Lucerne
Ps. nitrrificans 60 Meat-peptone broth 20 Meat peptone broth, lucerne

   Similar data were also obtained upon testingthe actinomycetes-antagonists. The latter, when growing in soils where appropriateorganic sources of nutrition are available, form antibiotic substances in detectableamounts (Table 114). We detected these antibiotics directly by the use of microbiologicalassay methods in the soil, and after extraction with organic solvents. Extractionas a rule, causes lower quantitative results. Where the microbiological assay methodindicates 100units/g by extraction one succeeds in isolating not more than 10-30units/g, i.e., about 10-30% of the total amount present in soil. Depending on thenature of the soil and the properties of the substance itself, the amounts of theextracted substances may vary (Krasil'nikov, 1954c).

Table 114
Formation of antibiotics by actinomycetes under various soil conditions
(units / g)

Soil

Actino- mycete 290, soy meal

Actino- mycete 290, corn extract

Actino- mycete 287, lucerne straw

Actino- mycete 287, starch

Actino- mycete B, lucerne straw

Actino- mycete B, corn extract

Sterile podsol

80

120

80

10

100

100

Nonsterile podsol

30

40

10

0

20

0

Sterile serozem

0

20

20

0

10

0

Nonsterile serozem

0

0

10

0

0

10

Sterile chernozem

30

60

20

0

50

80

Nonsterile chernozem

0

20

10

0

20

20

  Korenyako, Artamonova and Letunova (1955) studied theformation of antibiotics in soil by actinomycetes belonging to different species.In all more than 100 cultures were studied, 13 of them in great detail. The actinomyceteswere grown in soil with the addition of various organic nutrients. All of them, excepta very few, formed antibiotics in soil in greater or smaller amounts. The most activeones are presented in Table 115.

Table 115
Formation of antibiotic substances by actinomycetes in various soils
(units / g)
Actinomycetes

Podsol

Serozem

Cherno- zem

Krasno- zem

Podsol

Serozem

Cherno- zem

Krasno- zem

  Sterile

 

 

 

Non-

sterile

 

 

A. Aurantiacus, 1149

200

50

100

80

80

10

30

0

A. globisporus 81-B

120

100

60

60

60

60

40

20

A. globisporus 2302

80

30

100

20

30

0

20

0

A. globisporus 2570

150

20

60

50

50

20

20

0

A. griseus 2535

170

100

100

120

80

40

30

40

  The data given in the table show that microbial antagonistsform antibiotic substances with different intensities depending on the type of soil.The largest amount of these substances is formed in podsol soil, less in chernozemand chestnut soils and the smallest amounts in krasnozem.

  The antimicrobial properties of antibiotics expressthemselves differently in the soil than in artificial media. Not all the antibioticsare active in the soil. Martin and Gottlieb (1955) have shown, that circulin, neomycinand biomycin do not suppress the growth of the sporiferous bacterium Bac. subtilisin the soil even at very high concentrations (500 µ g/g), whereas on laboratorynutrient media they inhibit its growth at 0. 1 µ g/g. At the same time subtilinnoticeably inhibits the growth of Bac. cereus in these very same soils. Inthese author's experiments antinomycin was the most active. Five µ g/ g actinomycinin soil was a sufficient amount for the suppression of Bac. subtilis growth.

  The effectiveness of antibiotics in the soil is determinednot only by their properties but also by their concentration. In turn the latterdepends on the rate at which these substances are formed and enter, the soil on theone hand, and by the rate of their inactivation on the other hand. As is known, themajority of antibiotics disappear from the soil. Some are inactivated by the soilsolution or destroyed by microbes, others are washed out by water and a considerableportion is adsorbed by soil particles.

  in our experiments (Krasil'nikov, 1954c) we followedthe degree of activity of antibiotic preparations, introduced artificially into varioussoils under different conditions. The rate of inactivation of antibiotics, the extentof their being washed out by water and their adsorption by the soil were studied.The average indexes chernozem soil are presented in Table 116.

Table 116
Changes in the antibiotic content of soil (chernozem)
two hours after the introduction of the former
(in units/g)

Antibiotics

Inactivated

Washed out with water

Remained in active and absorbed state

Streptomycin

850

30

1,120

Globisporin

800

120

1,080

Terramycin

850

250

800

Pennicillin

300

1,320

380

Preparation No 1609

10,000

0

0

  Up to 2,000 units/g of chemically pure preparationswere introduced into the soil. Preparation 1609 was introduced at a concentrationof 10,000 units/g.

  As is seen from these data, streptomycin, globisporinand terramycin are inactivated in chernozem to more or less the same extent (about25%), penicillin, to a lesser extent and preparation No 1609 is completely inactivated.In other soils the inactivation of these antibiotics presents a different picture(Table 117). The least inactivation of these antibiotics is observed in podsol soilsand in krasnozem, and the greatest in chernozem.

Table 117
Minimal doses of antibiotics, at which the soil still exhibits antibacterial properties
(in units/ g)

Antibiotic

Serozem

Chernozem

Krasnozem

Podsol

Streptomycin

350

850

300

80

Globisporin

100

600

400

80

Terramycin

600

850

200

400

Preparation 1609

10,000

10,000

10,000

50,000

Penicillin

120

300

150

60

  In order to determine what part of antibiotics areadsorbed by the soils, we washed the latter with water. Upon introduction of antibioticsinto podsol soil in the amount of 2,000 units/g the following portions could be washedout with water: penicillin, 1,650 units/g; streptomycin, 40 units/g; globisporin,120 units/g; terramycin, 350 units/g, and the antibiotic No 1609--none.

  Penicillin more than other antibiotics can also bewashed out of other soils; e, g., krasnozem, 1, 800 units/ g and from serozem, 1,500 units/ g.

  By comparing the numerical indexes of inactivationof the antibiotics and their proneness to be washed out by water, the amount of antibioticsin the adsorbed state can be determined and therefore also the adsorbing capacityof the soil. According to our data in podsol soil the adsorption picture was as follows:globisporin--1,800 units/g; streptomycin--1,880 u/g, terramycin--1,350 u/g, penicillin--280u/g per gram. In serozem the figures were 1,670, 1,600, 1,200 and 380 units/g respectively.Streptomycin is adsorbed by chernozem at the rate of 1, 120 units /g and by krasnozem--1,650units/g; globisporin--1,080 and 1,540 units/g and terramycin--900 and 1,620 units/grespectively.

  As seen from this data the amounts of antibiotics detectedare considerably smaller than those introduced. In order to create antibacterialactivity in 1 g of serozem soil for example, it is necessary to use a minimum of350 units streptomycin; 100 units globisporin; 600 units terramycin; 130 units penicillinand more than 10,000 units preparation No 1609. Upon introduction into the soil ofsmaller antibiotic doses than those indicated, the antimicrobial properties of thesoil will not be detected either by a qualitative test or by extraction with varioussolvents (water, alcohol, ether, acetone, etc). All these data show that to the numericalindexes obtained upon quantitative determination of antibiotics formed in the soilby microbial antagonists, one should add the amounts of antibiotics adsorbed andinactivated. If 20 units/g of antibiotic substance were found in the soil, the actualamount produced by the antagonists would be considerably higher.

  The rate of destruction of antibiotics in the soilvaries. Some of them are inactivated within a few hours and others may be preservedfor a few days or even weeks, depending on the nature of the substance and the propertiesof the substrate.

  Antibiotics with basic properties such as streptomycinare very quickly inactivated in the soil, Neutral compounds (chloromycetin) are inactivatedslowly, and substances of the acid type occupy an intermediate position.

  Adsorption and inactivation of antibiotic substancesdepend to a considerable degree on soil acidity. Aureomycin and terramycin saturateneutral loamy soils at a concentration of 30,000 µ g/g, while for the saturationof acidic loam 60,000) µ g/g and more are required, i.e., twice as much.

  At pH 3.2 soil rich in humus adsorbs 4,000 µ g/gof antibiotics and at pH 5.6-7.6 only 400 µ g/g, i.e., ten times less.

  Consequently, the antimicrobial action of antibioticswill differ in these soils. In order to inhibit growth of Bac. polymyxa insoil at pH 5.6 a concentration of 5,00 µ g/g of terramycin is required, whileat pH 6.2, 200 µ g/g suffice (Gottlieb and Siminoff, 1952; Martin and Gottlieb,1952).

  The stability of antibiotics in the soil also varieswith the acidity of the latter.

  According to Gregory et al., (1952), the antibioticactidione is preserved in alkaline soil at pH 7.8 for 8 days and in an acidic soilat pH 5.2--for more than 14 days. Clavacin to completely inactivated in the firstday in alkaline soil and in acidic soil it is preserved for 3-4 days. Ninety percent of fradicin is preserved in alkaline soil for 14 days, in an acid soil it isadsorbed and completely inactivated an the first day.

  Chloromycetin is less strongly inactivated in the soilthan are streptomycin and terramycin. When chloromycetin is introduced into sterilesoil, it remains for more than 14 days without change. In nonsterile soil, with alarge number of various microorganisms, the preparation is gradually inactivated;after 3 days only 70% of it is left, after 7 days--about 30% and after 2 weeks only20% are recovered.

  Pramer and Starkey (1052) have found, that streptomycinintroduced into the soil at a concentration of 1,000 µ g/g is preserved in sterilesoil for over 3 weeks and in nonsterile soil for 2 weeks. About 50% of it is destroyedwithin the first week. In the presence of glucose the antibiotic is preserved fora longer period of time.

  Jefferis (1952) tested many antibiotic substances.He introduced them into various soils and observed the rate of their destruction.The following data have been obtained (Table 118).

Table 118
Preservation time of antibiotics in different soils
(days)

Antibiotic

Introduced µ g/g

Nonsterile podsol

Sterile podsol

Nonsterile orchard soil

Sterile orchard soil

Albidin

30

7

14

2

3

Frequentin

100

10

16

2

7

Gliotoxin

20

40

16

2

7

Griseofulvin

30

20

40

16

17

Patulin

2,000

32

32

2

2

Penicillin

50

3

2

2

2

Streptomycin

400

26

16

6

16

Viridin

100

8

16

1

1

  An may be seen, in orchard soil there is a faster inactivationof antibiotics. According to the author, certain substances are destroyed fasterin sterile soil than in nonsterile soil, which is puzzling and disagrees with theobservations of other investigators.

  According to our data (Krasil'nikov, 1954c), the preservationtime of antibiotics varies greatly and depends first of all, upon the propertiesof the substances, and secondly, on the type of soil and external conditions (temperature,humidity, acidity, etc), The same antibiotic is inactivated and parishes at differentrates in different soils. For instance, globisporin is preserved for 7 days in podsol,but for only 2 days in krasnozem. Aureomycin is active 10 days in podsol but notmore than 3 days in krasnozem (Table 119).

Table 119
Preservation time (days) of antibiotic substances

Soils

 

Globi- sporin

Prepar- ation No 112

Aureo- mycin

Terra- mycin

Prepar- ation No 1609

Penicillin

Chernozem Sterile

25

15

30

30

2

10

  Nonsterile

5

2

3

6

0.2

1

Podsol Sterile

90

80

60

50

5

20

  Nonsterile

7

5

10

8

0.1

3

Serozem Sterile

35

20

40

30

3

15

  Nonsterile

5

4

8

8

0.1

1

Red soil Sterile

22

12

20

15

2

5

  Nonsterile

2

2

3

3

0.1

0.5

  Antibiotics are most rapidly inactivated in krasnozemand podsol soils.

  Preparation No 1609 disappeared immediately in nonsterilesoils; other preparations could still be detected after a few days. In sterile soilsantibiotics are preserved for considerably longer periods of time than in nonsterilesoils. As can be seen from Table 119, the majority of active substances disappearin the first 2-3 days in nonsterile soils, while in sterile soils they are preservedfor 2 -3 months.

  Similar data are obtained upon introduction of crudeantibiotics in the soil. Korenyako, Artamonova and Letunova (1955) introduced activeactinomycetal substances in the form of culture liquids into podsol, chernozem, krasnozemand serozem soils. About 700 crude preparations were examined, 12 of them, in greaterdetail. The results are given in Table 120.

Table 120
Preservation time of crude antibiotics in soils
(days)

Antibiotics

Podsol, sterile

Podsol, non- sterile

Ser- ozem, sterile

Ser- ozem, non- sterile

Cher- nozem, sterile

Cher- nozem, non- sterile

Kras- nozem, sterile

Kras- nozem, non- sterile

A. violaceus strain 1806

20

5

20

5

20

5

10

5

A. aurauntiacus strain 1149

180

8

180

8

180

8

100

8

A. globisporus strain 81-B

180

8

180

8

180

8

5

2

A. globisporus strain 76

180

25

180

25

180

25

180

25

A griseus strain 2535

180

2

180

2

180

2

20

2

A. griseus strain 2392

120

20

120

20

120

20

120

20

Control soils

0

0

0

0

0

0

0

0

  The soil solution exerts a slightly inactivating effect.We tested a solution obtained by using a strong press from incubated samples of podsol,serozem and chernozem soils, with 4 antibiotics: penicillin, globisporin, preparationNo 15 (grisein) and preparation No 1609. The soil solution was added in various amountsto the antibiotic solution of a known titer and after a certain period (1-5 hoursor more) the activity of the preparations were examined. The results were not alwaysclear-cut, however, reliable data were obtained in experiments with grisein and,preparation 1609 and in some cases also with penicillin. The solution from incubatedpodsol soil inactivated antibiotics to a lesser degree than solutions obtained fromserozem and chernozem. One ml of solution extracted from podsol neutralized 20-30units of preparation No 1609, 5-7 units of grisein and 2-5 of penicillin. Soil solutionfrom incubated serozem inactivated 50-60 units of preparation No 1609, 7-10 unitsof grisein and 10-12 units of penicillin. Solution of incubated chernozem inactivated80, 15 and 25 units, respectively. In addition it also inactivated approximately5-7 units of globisporin.

  The inactivating force of soil solution is closelyrelated to the species makeup and metabolism of the microorganisms. If one incubatessoil in the presence of' small amounts of organic substance and in addition inoculatesthe soil with certain bacterial species, the solution thus obtained would inactivateconsiderably more antibiotics. We incubated podsol soil with various bacterial species(sporeformers and nonsporeformers) with an admixture of various sources of organicnutrition (soy meal, clover, hay, corn extract, peptone), The greatest effect wasobtained in an experiment with a nonsporeforming bacillus (strain No 6) which wasincubated in soil with soy meal. The solution obtained thereof (1 ml) inactivated100 units of preparation No 1609 and up to 50 units of penicillin, but did not inactivategrisein or globisporin. The latter was inactivated by solution from soil incubatedwith bacterium No 15 in the presence of clover hay.

  The inactivating action of the soil solution was observedby Waksman and Woodruff (1942), They examined the effect of actinomycin in pure solutionand with an admixture of a humus extract. A culture of Bac. mycoides was killedin the former case by 1 µ g and in the latter case, by 10 µ g or more ofthe substance. A similar effect was observed by Skinner (1956) while studying theantibiotic obtained by him from Actinomyces albido-flavus.

  Inactivation of antibiotics in soil is probably mainlyby products of microbial metabolism. It has been shown, that with increase of thelatter, destruction of antibiotics, is enhanced. In the above-mentioned experiments,in soil incubated together with soy meal, the growth of bacteria was very intenseand their composition differed from that in soil with clover or peptone.

  Winter and Willeke (1951 a, b) introduced penicillininto composted soil rich in humus and into loamy soil poor in organic substances.In humus soil, where there was abundant growth of microbes, the antibiotic disappearedafter 2-3 hours. In soil poor in humus the same antibiotic was preserved for 12 hours.If the soil microflora is removed by sterilization, penicillin in such soil is preservedfor more than 3 days, while in nonsterile soil it disappears on the second day.

  We have demonstrated the inactivating role of the soilmicroflora in experiments with pure cultures of bacteria and actinomycetes. It wasfound, that the antibiotics, penicillin, mycetin and streptomycin are inactivatedin different degrees by metabolic products of various species of bacteria and actinomycetes.Some species or strains of bacteria inactivate streptomycin more strongly, whileothers inactivate penicillin and mycetin more strongly. There are cultures the metabolicproducts of which enhance the activity of antibiotics (Krasil'nikov and Nikitina,1951).

  The data given here change our concepts on the preservationof antibiotics in soil. Antibiotics are not always fully inactivated in the soil.Depending on the soil-climatic conditions and also an the chemical properties ofthe antibiotics themselves, they may be preserved and accumulate in the soil.




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