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Part IV, continued: 

 

Concerning the Epiphytic Microflora

  In discussing the problems of interaction between microorganismsand higher plants one cannot ignore the epiphytic microflora. Microbial epiphytesare the organisms which concentrate on the surface of the green parts of vegetatingplants and are nourished by the excretions of the latter.

  The epiphytic microflora has been but little studied,especially its quantitative and qualitative composition on various plants.

  On the surface of the aerial parts of plants one findsdifferent microorganisms--bacteria, actinomycetes, fungi, yeasts, algae and protozoa.Their number may by very high. Duggeli (1904) counted many thousands of microorganismson the surface of cereal seeds. From 80,000 to 25 million bacterial cells and from4,000 to 7,200 fungi in one gram of wheat seeds have been detected by Morgentaller(1918). The author points out that on healthy seeds there are almost no fungi.

  In germinating wheat seeds there are 60,000 bacterialcells per gram of grains, and in nongerminating seeds--13 millions. Mack (1936),Kent-Jones and Amos (1930), inspected 21 samples of wheat seeds from different countries,and found from 8,000 to eight million bacterial cells on the surface of one gramof seeds. Gustafson and Parfeitt (1933) in a similar study counted from 46,000 to3,260,000 bacterial cells in one gram of wheat seeds.

  Rautenshtein (1939) studied the microflora of wheatseeds in the various stages of ripening: milky, waxy, and full maturity stage. Theresults of his observations are given in Table 136.

Table 136
Quantitative and group composition of microorganisms on ripening seeds of wheat
(number of cells in thousands in one gram of seeds)

Wheat variety

Maturation stage

Total No of microbes

Bacteria

Fungi

Actino- mycetes

Yeasts

Cesium 0111, second class

Milky

8,050

7,250

150

650

0

 

Waxy

5,525

5,275

225

25

0

 

Full

17,650

17,050

500

100

0

Cesium 0111, first class

Milky

33,375

32,500

625

250

0

 

Waxy

72,000

70,900

500

600

0

 

Full

41,500

41,250

250

0

0

  Bacteria are the most numerous among the microorganismgroups which were found. Yeasts are not always encountered.

  With ripening of the seeds the number of microbes ontheir surface increases. James, Wilson and Stark (1946) counted from 280,000 to 164million microorganisms in one gram of wheat seeds. The numbers of microbes foundon the surface of the green parts of plants are not smaller. Many investigators countedfrom 49,000 to 6,300,000 epiphytes in one gram of tissue (Khudiakov, 1953; Thomasand Hendricks, 1950; Stirling, 1951; James, 1955, and others). According to Kroulik,Burkey and Wiseman (1955). the number of epiphytes in one gram of tissue of greenplants of corn, oats, clover, lucerne, garden grass, and other plants varies from1,540,000 to 99,200,000. These numbers vary from species to species, in relationto the age of the plants, and also with the soil-climatic conditions. As a rule,the number of epiphytes on the surface of young plants is larger than the numberon ripening ones. In relation to seeds, the opposite picture was observed. Bacteriaform the greatest part of the epiphytic microflora. The species composition of thebacteria is quite diverse, but the dominating part of it is considerably small. Almostall the investigators noted the predominance of bacteria with a yellow pigment, classifiedas Ps. herbicola on plants. This bacterial species was described by Duggeli(1904), He found that these bacteria were the dominant species. Their total numberreached 380,000 and more in one gram of tissue.

  According to Weller (1929), Ps. herbicola comprises90-100% of the total bacterial flora on seeds of wheat and rye. Upon germinationof seeds in the soil, these bacteria soon disappear, reappearing toward the end ofripening. On growing plants, according to the author, there is abundant growth oflactic-acid bacteria. On seeds of barley and oats Weller found sporeforming bacteria.Rautenshtein (1939, a, b) found 75-98% of Ps. herbicola among the bacteriapopulating the surface of wheat seeds. Cocci and Sarcina are encountered in a fewcases. He also found a great number of lactic-acid bacteria. Among the fungal flora,Rautenshtein found the fungi Cladosporium herbarum, Trichoderma koningliiand less often Dematium, Asperigillus, Penicillium, Oospora and also the speciesA. globisporus and A. griseus and the yeast species of the genus Torulopsis.Many heat-resistant and thermophilic bacterial forms were found. The majority ofthese forms belonged to the sporeforming species of the type Bac. mesentericus.They also grow at a temperature of 17-20° C.

  James, Wilson and Stark (1946) distinguish betweentwo types of epiphytic bacteria--type A and type B. Type-A bacteria form yellow coloniesand are all considered to be cultures of the same species--Ps. herbicola.The other type belongs to the colorless Pseudomonas species. Of the fungalflora these authors found the following on plants: Acrostalagmus, Alternaria,Penicillium, Aspergillus, Botrytis, Cephalosporium, Fusarium, Torula, Moniliaand other fungi. There were also phytopathogenic species among them Helminthosporiumsativum, Hormodendron pallidum, H. viride, Alternaria tennis, Fusarium culmorum,Cladosporium herbarum, Septoria nodorum, etc.

  James (1955) gives the following data on the extentof distribution and accumulation of Ps. herbicola. Out of 200 plants of oats,barley, and flax, growing in different regions of Canada which were studied, morethan half contained this microbe in amounts of 100,000 and more, about 15 % of theplants contained from 10,000 to 100,000 bacteria and some samples were free of thisbacterial species altogether.

  Clark (1947) and others observed the yellow bacteriain great numbers on the green parts of the cotton plant.

  The predominance of these bacteria on other plantswas noticed by many investigators (Burri, 1903; Mack, 1936; Thomas and Hendriks,1950; and others).

  Wallace and Lochhead (1951) point out the connectionbetween the epiphytic and rhizosphere microflora. The latter, according to theseauthors, is intermediate between the microflora of the soil and the epiphytic microflora.

  Khudiakov studied a great number of plants (1953a).He investigated the migration of bacterial epiphytes from the surface of the seedsduring the germination of the latter, to the seedlings and later to all the plantorgans including fully formed and mature seeds. More than 20 microbial species wereisolated from different wheat varieties by this author. Among these microbes therewere three species of yeasts, two species of fungi and the remaining were bacteria,among which three,. cultures produced a yellow pigment, three others orange or red,one green, and all the others colorless. According to the author's observations,each of these species is the predominant form on some seed variety.

  Upon analysis of preharvest seeds of wheat (Moscowvariety, 2411) grown in the Moscow Oblast', 97% of the bacteria found were Ps.herbicola; no yeasts or colorless bacteria of the Ps. fluorescens groupwere observed. On another wheat variety which was grown alongside the former andunder the same conditions, there were 60% yeasts and no Ps. herbicola bacteriawere observed.

  Reciprocal cross infection of the wheat varieties bythe isolated cultures has shown that the latter were not specific. Epiphytes fromone wheat variety, when transmitted to another variety, grew as well as they didon seedlings of their own host plant. Khudiakov has shown that epiphytic microfloracan be changed at will by treating the seeds before sowing with the correspondingmicroflora. He treated sterilized oat seeds with cultures of epiphytic yeasts andbacteria which he isolated, and sowed them in open ground.

  Those microbes that had been artificially introduced(Table 137) were found on these plants. On the control plants the bacteria whichusually concentrate on this species predominated.

Table 137
Effect of bacterial inoculation of seeds on the composition of the epiphytic microflora of oats

Organisms introduced with the seeds

Number of colonies on plate

% yeasts, No 1 red

% yeasts, No 2 mycelian

% yeasts, White

% Bacteria, Ps. herbicola

% bacteria, yellow green

% bacteria, others

Red yeasts No 1

80

80.8

0

6.2

0

0

5.0

Mycelial yeasts No 2

107

1.8

90.6

2.8

0

0

4.6

Bacterium sp. yellow-green

132

5.3

4.5

14.4

7.5

65.1

3.0

Control seeds (not inoculated with bacteria)

84

2.3

2.3

9.5

80.9

0

4.6

  Kvasnikov and Sumnevich (1953) investigated woody plantsin Central Asia--poplar, apple, pear, cherry, maple, Greek nut, etc and grassy cultures:corn, lucerne, cotton, Sorghum cernium, milo, potato, sugar beet, cabbage,etc. In all cases, lactic-acid bacteria were found on all the plants in great quantities.According to the authors, on wild plants the number of these bacteria is considerablysmaller. The nearer the plants are to places of human habitation, the more lactic-acidbacteria one finds on the surface of plants.

  Kroulik, Burkey and Viseman (1955) divide the bacteriainto chromogenic and colorless groups. Among the former, the Ps. herbicolatype predominates and among the latter--lactic-acid bacteria Lactobacterium plantarum.

  In our studies we investigated various species of grassyand woody plants growing in the central belt of the USSR. The number of bacteriaand fungi on the surface of leaves and branches has been determined. Similarly toother investigators, we also detected hundreds of thousands and millions of bacterialcells per gram of tissue. In the different plants the predominant epiphytic microfloravaries in its species composition. In some plants Ps. herbicola predominatesand in others--other species of the genus Pseudomonas and Bacterium,and sometimes--lactobacilli. Quite often one finds large numbers of yeasts of thegenus Torula (T. rosea, or T. alba) and of the genera Sporobolomycesand Mycotorula.

  Large numbers of microorganisms are found on the surfaceof fruits. Studies show that on berries and fruits there are bacteria, fungi andyeasts, actinomycetes and even protozoa. Epiphytes are encountered on wild as wellas on cultivated fruits.

  On berries as on other parts of the plants the mostnumerous group of microbes are the bacteria with fungi and yeasts following. Oftenthere are as many as hundreds of thousands or even millions of them on 1 g of berries.The number of microbes varies with the variety and species of the berries, with thedegree of naturity, and with climatic and other external conditions. As a rule, theirnumber increases with the ripening of the berries.

  The quantitative ratio between bacteria, yeasts andfungi also change.

  The microflora of the vine grapes has been the mostthoroughly studied. According to the data of Akhinyan (1952) the total number ofmicroorganisms on the surface of vine grapes of the "Kakhet" variety rangesbetween 3,000 and 4,000,000 per 1 g, depending on the region and where the vine wasgrown (Table 138).

Table 138
Distribution of microflora on the surface of vine grapes
(according to Akhinyan, 1952) (number of cells in 1 g of berry)

Region (Armenian SSR)

Yeasts

Fungi

Bacteria

Ashtarak region

 

 

 

Oshakan village

20,850

4,100

650,000

Voskevaz village

75,000

--

296,000

Artashat region

 

 

 

Aizestan village

4,008,600

123,000

7,500

Yuva village

3,500

--

60,000

  The presence of such a large microflora on the surfaceof plants cannot be explained by their being carried over mechanically from the air.The accumulation of certain specific species speaks against it. The latter evidentlygrow and multiply on the plant surface. Consequently, they must find there sufficientquantities of food substances necessary for mass reproduction.

  Plants, as has been pointed out above, excrete variousvolatile and nonvolatile substances--with the aid of special glands or by guttation.In the drops formed by the guttation of rye grass glutamine was found (Chibnall,1939), Genkell (1946) observed the excretion of mineral salts together with the fluidexcreted by plants of salty marshes. Vigorov (1954) found in the guttation dropsof 7-to 9-day-old wheat seedlings 1.8 mg/ml of dry substance, containing 5-10 mg/mlammonium-nitrogen and 40-45 mg/ml phosphorous compounds. The author observed, thatthe intensity of guttation depends on illumination, soil humidity, on the presenceof nitrogen and other nutrient elements. Introduction of ammonium salts into thesoil elevates the excretion of nitrogen compounds in the guttation drop. One of thetests for the presence of organic substances in the fluids is the growth of microorganismsin them. According to the author, fungi grow abundantly in guttation drops.

  According to the data of Kholodny (1944 a,b,c) allor many of the organic substances excreted by plants are used by microbes as sourcesof nutrition.

  The significance of the epiphytic microflora in thelife of the plant is many-faceted. Among the epiphytic microflora there are manyactivators (Ps. herbicola, yeasts, etc), which form biotic substances--vitamins,auxins, folic acid, thiamine, riboflavin and other compounds, and also organismsforming antibiotic substances with strong antimicrobial properties.

  On the surface of leaves and stems of plants thereare microorganisms forming toxic substances. In the epiphytic group there are alsoparasitic and phytopathogenic forms.

  One should assume that the metabolic products of theepiphytic microflora behave in a certain manner in the plant tissue, having a definiteeffect on them. The ability of leaves to absorb various substances was known fora long time. On this basis methods of extrarhizal feeding have been elaborated andalso methods of introduction of substances with the aim of changing certain physiologicalfunctions shedding of leaves, arresting of flowering, etc.

  It was also established that plants can absorb variousmicrobial metabolites, vitamins, antibiotics and other compounds through the leafsurface. As was indicated above, these substances not only enter the plant throughthe leaves but they can be introduced by this route in large quantities for the purposeof feeding as well as for fighting bacterial and fungal infections. Among the epiphyticmicroflora there are numerous antagonists which produce antibiotic substances whichsuppress their competitors, and among them also phytopathogenic microbes. Growingabundantly on plants, such organisms may fulfill a protective role removing or suppressinginfectious agents originating from without. If we were to change the compositionof the epiphytic microflora on the surface of the green parts of plants at will,and form certain coenoses of antagonists there, this would prove to be of great valueto plant and fruit growing.




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