Microbial Associations of Soil Types E. N. MISHUSTIN Department of Soil Microbiology, Institute of Microbiology, Academy of Science of the USSR, Moscow, USSR
Abstract Microorganisms are characterized by wide ranges of distribution. Some groups, however, are known to have zones of active proliferation, and the development o f specific populations in discrete zones results in rather specific microbial associations in some soil types. The soils formed in warm climates are richer in microorganisms and contain more bacilli and actinomycetes. The spectrum of dominant microbial groups varies in different soil types.
The geography and ecology of soil microorganisms have not been well studied until recently. The paucity of research is evident in a fundamental monograph by Alexander [1 ]. The author could find very limited information on the subject. In his classical investigations, Dokuchayev [ 10] developed the doctrine of natural zones, a doctrine that offered scientific substantiation of the zonal distribution of plants, animals, and soils. However, the attempts of a number of scientists to ascertain patterns of distribution of soil microorganism ended in failure. It is believed that microorganisms are cosmopolitan and that their occurrence in soils is influenced only by factors such as climate, vegetation, etc. [ 16, 18, 45]. At the same time, it was shown that the transformation of organic and mineral substances is accompanied by microbial succession [ 15, 19, 20]. Hence, it can be concluded that the geographic environment, because it affects the processes of organic matter transformation in the soil, must promote the development of different microbial associations. Apart from the geographic factors, each zone is certainly defined by its environmental conditions. According to Kluyver [13] the cosmopolitan distribution of microorganisms does not mean that they occur everywhere in large numbers. Their active reproduction is promoted only by certain conditions. It seems that in different soil types, which are geographically dependent formations, different groups of microorganisms must predominate. 97 MICROBIAL ECOLOGY, Vol. 2, 97-118 9 1975 by Springer Verlag New York Inc.
98
E.N. Mishustin
For a long period of time, we have studied the soil microflora of different climatic zones of the USSR. Most observations were performed in warm periods of the year comparing the soil microflora beginning with the polar desert of the extreme north to the deserts of Middle Asia. We tried to find the specific features in their microfloras by ordinary research methods, which admittedly are not free from a number of generally recognized shortcomings. Seasonal fluctuations of numbers of soil microorganisms was found in all soil types examined. These findings have been reported by us [24, 30, 36]. For obvious reasons, the number of non-spore-forming bacteria show the greatest variation in the course of a season. The population fluctuations among the bacilli are less marked. The factors listed above appreciably affect the number of actinomycetes and microscopic fungi. The periodic changes in the number of soil microorganisms do not obscure the specific features characteristic of the microscopic population of certain soil types. It is obvious that the total number of soil microorganisms increases from north to south. Table 1 gives the approximately average values for the numerical composition of the microflora of a number of virgin soils in the European part of the USSR (A horizon). The data are based on a large number of tests. The bacterial abundance was estimated on beef-extract agar, actinomycetes on starch.ammonia agar and the spores of bacilli on mixed beef-extract and wort agar at a 1:1 ratio (pH 7.0). It was observed that the last medium is good for the differentiation of some Bacillus species. A great humber of bacilli are found in soil in the spore state. Thus, to facilitate the estimation of this group, the culture was obtained from pasteurized soil suspensions (10 min at 70~176 The fungi were estimated on wort agar. It is clear from Table 1 that southern soils are the richest in microorganisms. The somewhat high count in the virgin soils of the extreme north can be linked to accumulation on the surfaces of these soils of partially decomposed plant residues, which in the warm periods of the year form a good substrate for microorganisms, mostly fungi and non-spore-forming bacteria. Further to the south, the bacilli and actinomycetes play a more important role in the microbial community [23, 27, 29, 30]. As shown by our model tests, these microorganisms participate in the later stages of plant residue decomposition. This particular role in decomposition is possibly associated both with their powerful eiazymatic apparatus (particularly in actinomycetes) and their requirement for several growth factors (in bacilli). Since in warm climates the decomposition of plant residues is more intensive, the conditions are more favorable forBacillus and actinomycete reproduction. In a still more southern zone, the number of fungi decreases. Due to the increasing competition of bacteria and actinomycetes with
Table 1
c++ moderate. d+++ intensive.
Brown and sierozems
Desert steppe and desert
a+ extremely. b+ slow.
Chestnut
Dry steppe
Podzols and soddypodzolic
Forest meadow
Chernozems
Tundra-gley and gleypodzolic
Tundra and taiga
Meadow steppe and steppe
Soils
Zone
4490
3482
3630
1080
2140
Total number of microorganisms (X 103]g)
45.7%
45.4%
42.4%
77.2%
94.9%
Non-spore forming bacteria
17.7%
19.4%
21.4%
12.0%
0.7%
Bacilli
36.1%
34.6%
35.4%
8.1%
1.5%
Actinomyeetes
0.5%
0.6%
0.8%
2.7%
2.9%
Fungi
+++a
++c
++c
+b
_+a
Nitrification rate
The Average Number of Ma/or Groups of Microorganisms in Different Soil Types of the USSR
Q ~d
~~
o o u ".
100
E . N . Mishustin
the fungi, the number of the latter decreases in these soils, especially those characterized by neutral reactions. It seems appropriate to count the number of microorganisms, not per gram of soil, but per gram of humus since the metabolism of saprophytic microorganisms is associated not with the total mass of soil but with its organic matter. Using such estimations, the high "biogeny" of southern soils is made more evident (Table 2). Data obtained by the conventional technique of microorganism estimation by the use of solid media cannot be considered as absolute, but the general trends noted here can be recognized as fully objective. The higher density of microorganisms in southern soils is supported also by the data obtained by other methods, i.e., direct counting according to the technique of Winogradsky [9] and inoculation of a solid medium by fine earth according to the procedure of Novogrudsky [39]. The resulting absolute indices are, of course, different but the general trends remain the same. Soil types of some mountain ranges in the Caucasus and Middle Asia also have been studied. The effect of altitudinal zonality on the composition of soil microflora is the same as that of latitudinal zonality. Higher in the mountains, where the average annual soil temperature is lower, both the number of bacteria and the percentage of bacilli and actinomycetes decreases [21, 33]. Some soil types have different profiles and distributions of organic matter. These differences affect the microbiological profile of different soils. For example, in chernozems the microbiological profile is much deeper than in podzolic soils. Usually the humus content decreases with soil depth in a more gradual way than the number of microorganisms. Passing on to the characteristic of certain groups of soil microorganisms, I shall comment on the non-spore-forming bacteria. From data obtained in a study employing the commonly used media, the non-spore-forming soil bacteria are usually found to be represented by the genera Pseudomonas, Bacterium, and Mycobacterium. Up to now, we have failed to determine the differences in the composition of non-spore-forming bacteria in certain soil types. It is not unlikely that there are no essential differences whatsoever. The non-spore-forming bacteria are pioneers developing on the decomposing organic residues. Since the presence of fresh organic matter (mainly plant residues) is typical of all soil types and the differences in the chemical composition of this organic matter are not very great, it may well be that identical non-spore-forming bacteria develop in different soil types. Recently it has been found, however, that mycobacteria predominate in the primary phases of the soil-forming process [47]. It should be noted that the composition of the non-spore-forming soil bacteria require additional study. It has been already mentioned that more extensive decomposition of organic residues favors the reproduction of bacilli and actinomycetes. Thus, it might be expected that the specific or group composition of these microorganisms would be
Tundra-gley and gleypodzolic
Podzols and soddy-podzolic
Chernozems
Chestnut
Brown and chernozems
Forestmeadow
Meadow steppe and steppe
Dry Steppe
Desert steppe and desert
Soils
Tundra and taiga
Zone
Table 2
218500
99460
57450
32480
42800
Total number of microorganisms
101500
421 O0
26250
25040
40540
Non-spore forming bacteria
38500
20700
11250
3960
260
Bacilli
77500
36000
19500
2700
600
Actinomyeetes
The Average Number of Microorganisms Calculated per Gram of Humus (IX 103/g)
1000
660
450
780
1400
Fungi
7=
O
o
>
c)
102
E.N. Mishustin
dissimilar in soils of different climatic zones. As is clear from the above, the number of bacilli and actinomycetes increases appreciably in southern soils. Our expectations were realized. Table 3 shows average numerical values based on a large number of studies of groups of spore-forming bacteria characteristic of different soil types. Only virgin soils are considered here. As will be discussed below, cultivation brings about significant changes in the microbial community of soil. All bacilli are cosmopolitan, but for each species there are zones of optimum reproduction. Thus the group B. agglomeratus - B. cereus is rather characteristic of the soils of the extreme north. B. mycoides and B. cereus occur in large numbers of podzols of the central part of the USSR. In chernozem soils, intensive reproduction of the group containing B. idosus and B. megaterium is observed. The group B. mesentericus - B. subtilis is found in chestnut and grey soils. In addition, these soils contain a great number ofB. megaterium cells. Apart from these bacilli, in each soil type are found a certain number of cells of other species. The data obtained are quite convincing. Nevertheless, the groups can be further divided. For example, B. mesentericus is actually a combined species and can be divided into several subspecies. Recently, the distribution of different B. mesentericus strains in the soils of the European part of the US SR was analyzed in our laboratory. The results showed that their distribution is also associated with certain soil types. Thus, B. mesentericus niger is typical only of southern soils. Other strains of this species have a wider distribution range. The specific distribution of certain microbial groups is related to their physiological peculiarities. The bacilli able to use mineral nitrogen (B. megaterium, B. subtilis, B. mesentericus, etc.) obviously predominate in soils with intensive mineralization. The spore-forming bacteria which require organic nitrogen (B. agglomeratus, B. cereus, B. mycoides, etc.) obviously predominate in soils with slow rates of mineralization. In soils of altitudinal zonality, a similar shift of bacterial forms is observed, but since these soils are not quite identical with the zonal ones, certain deviations are observed. Thus, the mountain-meadow soils, like the tundra soils, are poor in B. mycoides but rich in the B. cereus - B. idosus group. B. mycoides occurs in mountain chernozem soils and is practically absent in common chernozems. Recently, the distribution in USSR soils of anaerobic spore-forming bacteria of the genus Clostridium was studied [37]. In the ecology of these microorganisms, certain regularities were also found. Northern soils proved to be much richer in these bacteria than southern soils. In the northern zone, the soils contain chiefly C. pasterianum and a large number ofC. butyricum cells. In southern soils the dominant form is C. acetobutylicum. Table 4 gives average data illustrating the general ecology of bacteria of the genus Clostridium.
Table 3
Gley-podzolic
Podzols Soddy-podzolic
Chernozems
Chestnut
Brown soil and sierozems
Solonetz
Taiga
Forest-steppe
Meadow steepe and steeppe
Dry steppe
Desert steppe
Semidesert
4
1
10
4
[26 [ 25
150[
~
0
0
0
0
10 17
4
0
~
12]
0
~
4
0
~
4
0
6
4
0
0
0
0
30
40
33
63
20 37] 0 21 I 0 - ~
[25
0
~
aThe species making up the typical soil community are outlined. b 0 Means that the species occur in small numbers. CBacilli are differentiated according to Krasilnikov's guide (I 949).
Tundra-gley
Soils
Tundra
Zone
34
26
16
17
0 5
0
0
~
7
8 I
8 [
0
0 0
0
0
4
0
0
0
0 0
0
0
~ ~" ~
--
4
0
0
0 0
0
0
~
~
I
10
8
4
3 4
2
2
~
3 I 10
10
10
4 5
3
4
~'
5
15
-
15
-
42
Species
Unidentified
Composition o f the Spore-Forming Bacteria in A Horizon of Different Soils {~ o f Total Number of Bacilli)a, b,c
%o
o.
o
w,
>
o
104
E.N. Mishustin Table 4
Bacteria of the Genus Clostridiurn in Virgin Soils of Different Horizontal Zonalities (X 1036g) a Butyric acid bacteria
Soil types
Soddy-podzolic
7000
700
20
1
100
30
30
1
10
5
5
10
2.5
5
15
Chernozems Chestnut Sierozems
0.5
C. butyricum
Acetone butyl
C. pasteurianum
Pectinolytic clostridia
(C. acetobutylicum )
a The values shown are the maximum values observed.
As is clear from model tests, C.pasterianum develops intensively during the first period of plant residues decomposition, and C. acetobutylicum during the final period. The latter prefers a protein-containing medium and requires a complex of vitamins, and thus it can reproduce only on organic residues adequately enriched with microbial protein and vitamins. Thus, it is natural that southern soils (richer in microorganisms) are more suitable for C. acetobutylicum and northern soils for C. pasterianum. Since the clostridia studied are nitrogen-fixing bacteria, I shall dwell briefly on the ecology of other free-living nitrogen fixers studied by us, namely, Beijerinckia and Azotobacter.
Beijerinckia was found only in the subtropical red earth of Georgia, soils having a low pH value. This genus occurs there in both virgin and cultivated soils. In humid and hot climates, large numbers of Beijerinckia are readily found in the soil at any season of year. There are many publications dealing withAzotobacter and its distribution in different soil types of the USSR and of many other countries [6, 12, 22, 35, 44, 46]. Azotobacter is rather fastidious in its particular nutrition. It develops in soils rich in available phosphorus compounds, having a pH close to neutrality, containing organic substances, provided with water, etc. A complex of such conditions is not common, even in the zone of rich chernozem soils. Thus, it is not surprising thatAzotobacter is not always found in podzolic soils and that it occurs mostly in neutral soils of flood-plains and highly cultivated lands. In the zone of chernozems
Microbial Associations of Soil Types
105
and more southerly soils, it occurs regularly only in irrigated and sufficiently mostened soils. In the nonirrigated chernozems, chestnut soils, and sierozems, Azotobacter reproduces only in the spring. Apparently, its appearance mainly depends on the increased availability of water. Because of this relationship and a number of others, the organism is with full reason brought together with the algae. A similar picture is observed in chestnut and sierozem soils [22, 48]. From the discussion above, it is clear that a true picture of the ecology of Azotobacter can be obtained only through regular observations. An analysis of randomly collected soil samples without consideration of soil condition results in meaningless conclusions on the ecology of Azotobacter. In general, the distribution of Azotobacter is not related to any particular soil-climatic zone. It may be found both in the north and in the south but only in highly fertile soils. The presence of this organism is a good index of agronomically valuable soil properties. The evidence presented supports the conclusion that some microorganisms, such as Azotobacter, do not reflect the soil type but rather certain ecological conditions. The effect of geography on the distribution of microscopic fungi in soil has been considered in a number of studies. Some points relating to the distribution of microscopic fungi in soil have become generally recognized. For example, it is generally accepted that the variety of soil fungi increases in southern soils, although their total number in these soils markedly decreased [ 17]. It has been found that fungi of the genusAspergillus are largely absent from northern soils and are rather common in southern soils [43, 54]. During the studies of soils of different zones, a considerable amount of information on fungal ecology was collected in our laboratory. A part of it has already been published [25]. The data are summarized in Table 5. An analysis of the results allows for the establishment of a number of generalizations. Thus, in soils with nonintensive mineralization, the relative abundance of fungi of the genera Penicillium and Mucor increases. Further to the south, fungi of the genus Penicillium are replaced by fungi of the genus Aspergillus. The probable reason for this change is that representatives of the genusAspergillus are able to assimilate more complex organic compounds. It should be noted that the relative percentage of Aspergillus not infrequently increases in alkali soil located in the chernozem zone.
Mucor strains that prefer organic nitrogen compounds are common in the upper layers of virgin soil containing many nondecomposed plant residues. Mucor ramanianus is found only in forest soils. Representatives of the genus Choanephora occur only in sierozems.
106
E. N. Mishustin Table 5 Composition o f Microscopic Fungal Flora in A Horizons of Different Soils f~ o f the Total Number of FungiJa, b
Zone
Soils
Tundra
Tundra-gley
72
0
3
Taiga
Gley-podzolic, soddy-podzolic
71
0
6
Steppe
Chernozems
71
1
Dry steppe
Chestnut
63
Desert steppe
Sierozems
47
Semidesert
Solonetz and solonchak
48
1
7
2
7
0
11
10
5
0
7
1
7
1
0
6
4
8
4
0
5
~
aOutlined are fungi characteristic of a particular soil type. b0 Means that the genus occus in small numbers.
Of interest is not only the changing number of Penicillium in soils of different soil-climatic zones but the changing ratio of representatives of sections of this genus. In soils with less intensive biochemical changes, the biochemically less active representatives of the genus Penicillium predominate. In soils with less favorable conditions, say, in solonetz (alkaline), solonchak (saline), and takyr (desert soils), fungi of the Monoverticullata section dominate. The section B iverticullata is chiefly largely represented in forest soils. The section Asymmetricullata is widely distributed in most parts of cultivated soils. Representatives of the genus Fusarium are concentrated in soils covered by grass vegetation. Forest soils are practically free of these fungi. Chernozem, chestnut, and sierozem soils are rather rich in representatives of this genus. Thus, it can be concluded that the composition of the estimated fungi undergoes essential regrouping in different soil types. It would be more than desirable to estimate not only the generic composition of microscopic fungi in different soils but the specific composition as well. It can be supposed that the specific features of the fungal community in different soil types could be described still more clearly.
Microbial Associations of Soil Types
107
It is appropriate to make some remarks on soil yeasts. On the basis of our data as well as the results of Pumpyanskava [42] it is possible to conclude that large numbers of yeasts occur in soils rich in partially decomposed organic materials. That is quite natural since yeasts utilize only simple sugars and organic acids. Thus yeasts are rather widespread in northern soils. The study of Babyeva and Golovleva [5] dealing with yeast ecology should be mentioned. The authors have thoroughly studied a diversity of soils beginning with the tundra zone and finishing with the dry steppe, and they confirmed that the largest number of yeasts are found in soils rich in slightly decomposed plant residues and characterized by an acid reaction and rather high moisture content. In other soils, yeasts occur in small numbers. It is of interest that different yeast types are characteristic of certain soils. Representatives of the genus Candida having a mycelial stage in its developmental cycle predominate in acidic podzolic and soddy-podzolic soils. In tundra-gley and boggy soils, capsulated yeast of the genus Cryptococcus are mainly present. Yeasts of the genus Lipomyces are constantly found in grey forest and chernozem soils. In addition, pink yeasts (Rhodotorula) are also often found in chernozems. As regards the diversity of yeast species, the peaty soils are the richest. Yeasts of the genus Torulopsis are constantly foundthere together with other species. This yeast type is not found in other soils. Literature on yeast ecology is far from abundant, but it shows the differences in the prevalence of this group of certain soil types. The general regularities in actinomycete distribution are quite clearly seen from the data collected, and these distributions are confirmed by a number of other studies [40, 49]. The data allow the conclusion to be drawn that northern soils are not only poor in actinomycetes but the composition of the actinomycete microflora is very limited in them. Further to the south, both the number and the diversity of actinomycetes increase greatly. Table 6 gives average data obtained by us in our studies of the actinomycetes of a number of soils. There is no doubt that the data need further definition, but still some conclusions can be drawn from them. Thus, it can be stated that in southern soils there is a great number of actinomycete groups. A higher diversity of species can be also postulated. Representatives of some species of Streptomyces, such as albus, griseus, globisporus, and chromogenes, occur in appreciable numbers in almost all the soils studied. Nevertheless, further to the south, the numbers of S. griseus and S. globisporus decrease. Some actinomycetes are represented in larger numbers either in northern or in southern soils. Thus, the representatives of the S. albidus and S. viridis generally occur in considerable numbers in northern soils whereas S. fradiae, S. flavus, and S. rubroaurantiacus are more often found in the soils of the northern zone. Southern soils of the USSR are undoubtedly richer in pigmented actinomycetes.
14.7
10.5
Solonetz
Solonchak
0
0
0
6.8
4.2
3.5
6.8 0
8.2
1.8 3.
~
0
0
3.5
0
8.8 i
5.9
0
0
0
0
0
0
13.5 10.1 I 0
6.2
o~ O
1.9
0
34.5 0
16.7
28.13 3.5
16.13 0
3.7
0
0
0
0
0
0
Asporogenic
7.7 15.4[ 7.7 0
~"
3.7 20.7 I 0
0
0
I~
3.5 24.1
.51110.41 14.7
0
4.5
0
0
0 2.3
0
0
0
0
aOutlined are the species listed making up the typical soil community. ~0 Means that the species occurs in small numbers.
28.0
0111.3
30.7 115.4 7.7 15.4 0 I 17.8111.7 35.3 11.7 0 r 17.2 17.0 1.9 i 20.0 0 4.5110.1 7.
Chestnut
Typical chernozem
Greyforest
Soddy-podzolic
Podzolic
Soils
~"
Streptomyces Species
Table 6 Composition of Actinomycetes Microflora of Different Soils (% of Total Number of Actinomycetes]
3.5
0
0
0
0
0
z K ~r
O0
109
Microbial Associations of Soil Types
Table 7 Distribution of Pigmented Actinomycetes in the soils of Kazakhstan (% of the Total Number of Actinomycetes) Soil M o t m t a i n - m e a d o w , alpine
Mountain chernozem, virgin Mountain chcrnozen~, cultivated Sierozem, virgin Sierozem, cultivated
Pigmented 21.0 43.4 46.0 46.0 51.2
Nonpigmented 79.0 56.6 54.0 54.0 48.8
According to Tyeplyakova [49], who studied the vertical zonation of the soil microflora of in Ala-Tau mountains of Kazakhstan, the number of pigmented actinomycetes (Table 7) decreases as the absolute altitude of soils above the sea level increases. The specific composition of actinomycetes of different soils has received little study. Nevertheless, from the available data, certain interesting regularities appear. Thus, as reported by Tyeplyakova, Actinomyces (Streptomyces)fumosus, A. candidus, and A. actinoides often occur in mountain-meadow soils, while they are largely absent from sierozems.* A. verticullatus, A. virgatus, and A. longisporous are, however, not found in mountain soils, while they are common largely represented in sierozems. Some species of actinomycetes, such as A. globisporus and A. griseus, occur in all soil types. In my l a b o r a t o r y , m i c r o o r g a n i s m s of the genus Nocardia (proactinomycetes), which are related to the actinomycetes, were studied by Tepper [ 5 0 - 5 2 ] and Aristarkhova [3]. Proactinomycetes are not easily differentiated in ordinary laboratory media. Thus, for the estimation of Nocardia, a nutrientdeficient agar medium was chosen, one similar in composition to the medium used by Winogradsky for the cultivation of nitrite-forming bacteria. The small colonies of Nocardia which develop in this medium are easily differentiated by use of the low power on the microscope. Physiological studies of a rather large collection of Nocardia have led to the conclusion that a considerable number of them can mineralize humus constituents. In agreement with this conclusion, it has been established that in soils having high rate of humus mineralization the percentage of proactinomycetes in soils increases. The data given below show the percentage of microorganisms of the genus Nocardia in the total number of colonies estimated on the media mentioned above. *Usage in the United States differs,
Streptomyces.
andActinomyces as used in this report is commonly designated
110
E.N. Mishustin
As is evident from these data, the soils become richer in proactinomycetes as one progresses from north to south. Cultivation also results in soil enrichment with these microorganisms: % of Proactinomyces 1.9 9.4 6.8 16.0 25.0
Northern peat-boggy soils Soddy-podzolic soils Chernozems Light chestnut soils (virgin soils) Meadow (cultivated soils)
Certain species ofproactinomycetes are abundant in different soils. Thus the soddy-podzolic soils are rich in N. corallina, chernozem in N. symbioticum, and chestnut soils in N. citrica. The increasing number of microorganisms in southern soils and the higher percentage of bacilli, actinomycetes, and proactinomycetes are indicative of more intensive mineralization of organic compounds of soil. A direct index of this greater activity is the more rapid nitrification. It was established by Kostychev [ 14] that nutrification increases from north to south, and his observations are fully supported by our results. The titer of nitrifying bacteria is very low in podzolic and soddy-podzolic soils. The numbers of nitrifying bacteria increase greatly in chernozems and more southern soils. Nitrification processes correspondingly increase in southern soils. A similar phenomenon is observed in mountain soils. While studying the microbiological features of the A horizon in soils of Zailiysky Alatau vertical zonality, Tyeplyakova [49] came to the conclusion that the nitrifying ability of the soils of warmer climates is much higher than that of soils formed under more severe conditions. The estimation of nitrification activity of soils has shown that the quantity of nitrate accumulating per kg of humus in the mountain-meadow soil in a three-week period is 1.2 rag, in chernozem 6.9 rag, and in zierozem 62.2 mg (Table 8). Thus, the soils that are richer in microbial biomass are more actively forming inorganic compounds of nitrogen. Table 8
Nitrification in Different Virgin Soils of Zailiysky Alatau Soil
Mountain-meadow Mountain-forest Chernozem Dark-chestnut Sierozem
Depth (cm)
Amount of NO~ mg per kg of humus
0-10 0-12 0-13 0- 9 0- 6
1.2 2.9 6.9 26.5 62.9
Microbial Associations of Soil Types
111
It should be noted that the greater effect of equal quantities of mineral fertilizers in the north than in the south is a very important practical result of the differences in mineralization processes in different soil types [2, 53] This greater influence in the north can be ascribed to the slower rate of formation :of plant nutrients in northern soils. The abundance of cellulose decomposing microorganisms is also indicative of mineralization rates [5] in soil. It is known that cellulose can be decomposed both by bacteria and fungi. Cellulose decomposing microorganisms differ greatly in their demands for available nitrogen, however. Among the cellulollytic microflora, fungi of the generaDematium andPhoma are least specific in their demands, and that is why they predominate in soils with slow mineralization rates. Cellulose decomposing fungi such as Chaetomium and Fusarium require a higher level of nitrate for growth. Bacteria require a still larger quantity of available nitrogen than fungi. Thus, in soils with high nitrate levels, a considerable number of myxobacteria (Polyangium and Myxococcus) appear, and at still higher concentrations, vibrios (Cellvibrio) and myxobacteria of the genus Cytophaga assume prominence. These generalizations apply to soils of the same zone. However, since the rate of mineralization in soil increases from north to south, the relative content of cellulose decomposing bacteria in warmer soil-climatic zones increases greatly compared to that of the fungi. More detailed information on this aspect has been published [30, 36]. In these biological characteristics of certain soil types, we have used data referring only to the A1 horizon of virgin soils. It should be stated however, that the composition of the microflora changes with depth. The deeper soil layers are relatively more rich in bacilli and, not infrequently, in actinomycetes. This is particularly evident in the chernozem and sierozem, the soil types in which these microbial groups are found in large numbers (Table 9). The specific composition of microorganisms is also affected by depth, as can be illustrated from data obtained in studies of by chernozem soil. Thus, some spore-forming bacteria, for example, B. idosus, penetrate rather deep into the soil
Table 9 Number of Microorganisms in the Profile of a Chernozem Soil in the Kharkov region, • 103/g of soil Depth (cm)
0- 5 5 - 10 20- 30 40- 50 70 - 80
Humus (%) Bacteria
9.2 9.1 7.7 4.5 2.7
8950 6650 835 200 147
Bacilli
Actinomycetes
Fungi
815 945 825 169 127
835 1015 126 24 13
37.0 36.5 19.5 17.2 0.3
112
E.N. Mishustin
layer. Other bacteria such as B. megaterium and B. mycoides are concentrated in the upper layer of soil. It is the same with fungi. Fungi of the genus Penicillium are the major representatives of the microflora of deeper soil layers. The genera Mucor, Fusarium, and Trichoderma inhabit chiefly the top layers of soil. Table 10 illustrates the changes in the soil micro flora with depth. It shows the distribution of a number of microorganisms in the profile of a chernozem soil. The data illustrate the effect of geographic factors on the composition of the microflora. Thus, it is necessary to consider the effect of ecological factors and the influence of cultivation on the microbial community. Such factors as mechanical composition of soil, type of vegetation, and increased percentage of salt produce a rather noticeable effect on the composition of the soil microflora. The ecological factors cannot be neglected, but its effect is influenced by a stronger effect of geographical zonality. The type of changes in the microbial community under the influence of vegetation and cultivation can be seen from Table 11, in which some chernozem soils of nearby areas of the Voronezh region are considered. The application of organic fertilizers, etc., produces a strong effect on the soil microflora, and this influence has already been discussed [28, 30, 36] so that only some general remarks need to be made. Cultivation improves the temperature and water-air status of soil, resulting in more rapid mineralization of organic materials, and the soil acquires a number of characteristics typical of more southerly soils. An increase is observed in the number of microorganisms, the percentage of bacilli (particularly the number ofB. megaterium cells), and the percentage of bacteria among the cellulose-decomposing microorganisms. The abundance of fungi is reduced, whereas the nitrifiers become more numerous. The application of mineral fertilizers, especially those containing nitrogen, results in significant changes in the composition of some groups of microorganisms. Thus, for example, the application of ammonium sulfate to the soddy-podzolic soil results in rapid reproduction of bacteria of the genus Cellvibrio and a sharp reduction in the abundance of fungi. Soil fertilization gives rise to a large number of Cytophaga cells, etc. Barnyard manure contains many thermophiles, and manured soils thereby become enriched with this group of microorganisms. Barnyard manure rich in organic substances also stimulates reproduction of a number of microorganisms, including Azotobacter. The data collected allow for the establishment of a complex of microbiological indices characteristic of the soil type and its cultivation status. This calls for a special discussion. Recently the study of the soil community composition has utilized the aid of capillary studies [4, 41] and electron microscopy. In our laboratory, we have used electron microscopy in studies of soil [7, 8, 38], and it has been shown that the number of microorganisms in soil is much higher than is usually revealed by ordinary research methods. A number of " n e w " forms of microorganisms were found, and these have
Microbial Associations of Soil Types
1 13
7~ichoderma
oO
r
r
~
0
0
0
o r
eq. o
o
o
"4
0
oO
t~
Cq
r
r--
--
06
t'-
tz
~o
~
~
c5
t~
tt~
t1%
q~
0"~
tO)
tt3
r
t"q
tr
tl~
c5
c5
o
o
o
o
~
d~
d-
6
6
"o"
t"-
O~
Fusarium
Mucor
Penieillium
tt~
B. i d o s u s
6
tae)
B. m e g a t e r i u m
B. m e s e n t e r i c u s
B. m y c o i d e s
oo
6
114
E . N . Mishustin
Mucor
Chaetomium r~
Fusariu m
r~
Aspergillus
tq
Penicillium
B. mesentericus
B. m y c o i d e s
~.N ~.~
c5o
oc5
~5c5
oo
o
r
r'qo
0
B. megaterium
B. agglomeratis
0 r~
0
Microbial Associations of Soil Types
115
been rather thoroughly studied. What is the relationship between the common, well-studied microorganisms and the " n e w " ones, and is it necessary to radically revise our concept of the soil community? It seems that these questions can now be answered. Winogradsky divided the soil microflora according to its functions into two groups: (a) zymogenous and (b) autochthonous. The first decomposes organic residues, the second, soil humus. The data collected allow one to conclude that the common saprophytic microorganisms (bacteria, fungi, actinomycetes, etc.) must be referred to the zymogenous group that are chiefly responsible for metabolizing the organic matter entering the soil. Furthermore, a specific microflora decomposing only humus does not exist; this function is fulfilled by some saprophytic microorganisms, including representatives ofPseudomonas, Nocardia, and other microorganisms [26, 32, 34, 52]. Thus, the so-called "autochthonous" microflora should also include certain representatives of the zymogenous microflora which are able to decompose humic substances as well as simple compounds. The studies of a number of " n e w " microorganisms carried out in our laboratory suggest that they are mostly oligocarbophiles, developing in media with a reduced concentration of carbohydrates. Obviously, the group mineralizes the residues of organic materials decaying in the soil and can be considered as a satellite. According to the terminology suggested by Zavarzin [ 11 ], these microorganisms can be called the "dispersion microflora." The composition of this group in different soils requires additional study. The author expresses thanks to M. Alexander for the correction of the translated text of this article.
References 1.
Alexander, M. 1971. "Microbial Ecology." Wiley, New York.
2.
Anikst, D. M. 1969. O geografii deistviya mineralnykh udobrenii na urozhai yarovoi pshenitsy. J. Agrokhimiya 10: 37-42.
3.
Aristarkhova, V. I. 1968. Uchastiye pochvenykh proaktinomitsetov v razlozhenii organicheskikh veshchestv. Izvestia AN SSSR, Ser. Biol. 2:301-305.
4.
Aristovskaya, T. V. 1945. "Mikrobiologiya Podzolistykh Pochv." Publ. h. Nauka.
5.
Babyeva, I. P. and Golovleva, L. A. 1963. Drozhevaya flora osnovnykh tipov pochv Yevropeiskoi chasti SSSR. Sbomik "Mikro-Organizmy v Selskom Khozyaistve." Publ. h. Moskovskogo Gos. Universiteta, p. 231-251.
6.
Blinkov, G. N. 1959. "Azotobakter i Yego Znacheniye Dlya Rastenii." Publ. h. Tomskogo Universiteta.
116
E. N. Mishustin
7.
Vasilyeva, L. V. 1972. Osobenosti ultrastruktury i tsikl razvitiya Stella humosa, lzvestiya AN SSSR, Ser. Biol. 5: 782-788.
8.
Vasilyeva, L. V. 1972. O tsikle razvitiya i tsitologicherskikh osobennostyakh novogo pochvenogo mikroorganizma oblada-yushchego porstekami, lzvestiya AN SSSR, Ser. Biol. 6: 860-864.
9.
Winogradsky, S. N. 1952. Pryamoi metod v mikrobiologicheskom issledovanii pochvy. "Mikrobiologiya Pchvy." AN SSS.
10.
Dokuchayev, V. V. 1899. "Kucheniyu o Zonakh Prirody. "SPB.
11.
Zavarzin, G. A. 1970. K ponyatiyu mikroflory rasseyaniya v krugovorote ugleroda. J. Obshchaya Biologiya 31(4): 386-393.
12.
Zinovyeva, Kh. G. 1962. "Azotobakter i Selskokhozyaistvennyye Rasteniya. "Publ. h. AN Ukr. SSR.
13.
Kluyver, A., and van Niel, C. B. 1956. "The Microbes Contribution to Biology. "Harvard University Press, Cambridge.
14.
Kostychev, S. P. 1930. Byvody agrotekhnicheskogo kharaktera iz rabot otdela po biodinamike pochvy. Trudy Instituta Selkhoz. Mikrobiologii 4: 29-36.
15.
Krasilnikov, N. A., and Nikitina, N. I. 1945. Vliyaniye razlagayush-chikhsya kornei na sostav mikroflory v pochve. Pochvovedeniye 2: 132-145.
16.
Krasilnikov, N. A. 1958. "Mikroorganizmy Pochvy i Vysshiye Rasteniya. "Publ. h. AN SSSR.
17.
Kursanov, L. I. 1940. "Mikologiya." Uchpedgiz.
18.
Kriss, A. E. 1947. Mikroorganizmy tundrovyh i polyamopustynnykh pocbv Arktiki. Mikrobiologiya 16: 5,437.
19.
Mai, H. 1970. Bacterial succession in degradation of Alfalfa, rye-straw and maize root. Zentr. Bakteriol. H 124, 508-524.
20.
Mishustin, E. N., and Timofeyeva, A. G. 1944. Smena mikroflory pri protsesse razlozheniya organicheskikh ostatkov. Mikrobiologia 13: 272-283.
21.
Mishustin, E. N., and Mirzoyeva, V. A. 1950. Rastitelnyye poyasa gor i ikh otrazheniye v sostave bacteeialnogo naseleniya pochvy. Mikrobiologiya 19 (4): 299-307.
22.
Mishustin, E. N. 1954. O rasprostranenii azotobaktera v pochvakh. Mikrobiologiya 22: vypusk 4, 408-424.
23.
Mishustin, E. N. 1958. Geografichesky faktor i rasprostraneniye pochvenykh mikroorganizmov. lzvestiya AN SSSR, Set. Biol. 6: 661-676.
24.
Mishustin, E. N., and Teplyakova, Z. F. Sezonnaya dinamika mikrobiologicheskikh protesessoy i eye agronomicheskoye znacheniye, lzvestiya AN Kaz. SSR, Ser. Botanika i Pchvovedeniye 3 (6): 15.
25.
Mishustin, E. N., Pushkinskaya, O. I., and Teplyakova, Z. F. 1960. Ekologogeograficheskiye zakonomernosti v rasprostranenii pochvenykh gribov. Izvestiya AN SSR, Ser. Biol. 5: 641-665.
26.
Mishustin, E. N., and Nikitin, D. 1. 1961. Atakuyemost guminovykh kislot pochvenymi bakteriyami. Mikrobiologiya 33 (5): 841-847.
27.
Mishustin, E. N. 1962. Regularities of incidence of Actinomyces in the soils of the USSR. Abstr. 8. Inter. Cong. Microbiology, p. 61.
28.
Misbustin, E. N., and Tepper E. Z. 1963. Vliyaniye dlitelnogo sevooborota, monokultur i udobrenii na sostav pochvenoi mikroflory, lzvestiya TSKhA 6: 85-94.
Microbial Associations of Soil Types
1 17
29.
Mishustin, E. N. 1964. Les differents types de sols et la specificity de leur micropopulation. Ann. Inst. Pastrur 107: 3, 63-78.
30.
Mishustin, E. N. 1966. Geografichesky faktor, pochvenyye tipy i ikh mikrobnoye naseleniye. Sbomik "Mikroflora Pochv Severnoi i Srednei Chasti SSR". Nauka, p. 3-23.
31.
Mishustin, E. N. 1966. Microorganismes cellulolytique des sols de L'URSS. Ann. Inst. Pasteur 110: 596-603.
32.
Mishustin, E. N., and Mrysha, G. N. 1967. Fraktsiya peregnoinykh soyedinenii i ikh razrusheniye mikroorganismami. Symposium Humus et Planta IV. Prague, p. 55-61.
33.
Mishustin E. N., Mirzoyeva V. A. 1968. Sporeforming bacteria in the soils of the USSR. The ecology of Soil Bacteria. Liverpool U. P., p. 458-473.
34.
Mishustin, E. N., Nikitin, D. I., and Tepper, E. Z. 1968. "Autochtonous Soil Microflora." and Festkrift til H. L. Jenson, p. 77-87.
35.
Mishustin, E. N., and Shilnikova, V. K., 1968. "Biologicheskaya Fiksatsiya Atmosfernogo Azota." Publ. h. Nauka.
36.
Mishustin, E. N. 1972 "Mikrrorganizmy i Produktivnost Zemledliya." Publ. h. Nauka.
37.
Mishustin, E. N., and Yemtsev, V. I. 1973. Anaerobic nitrogen fixing bacteria in USSR soils. Soil. Biol. Biochem. 5: 97-107.
38.
Nikitin, D. I., Vasilyeva, L. V., and Lokhmacheva, R. A. 1966. "Novyye i Redkiye Fomay Pochvenykh Mikroorganizmov." Publ. h. Nauka.
39.
Novogrudsky, D. M. 1948. Novy metod issledovaniya pochvenoi mikroflory. Publ. h. AN SSSR, Ser. Biol. 6:211-236.
40.
Panasyan, A. K., and Tumanyan, V. G. 1958. O nekotorykh biologicheskikh osobenostyakh aktinomitsetov v pochvakh Armyanskoi SSR. Sbomik "'Voprosy S and Kh i Promyshlenoi Mikrobiologii, " 7-75-87.
41.
Perfilyev, B. V., and Gabe, D. R. 1961. "Kapillyarnyye Metody lzucheniya Mikroorganismov." Publ. h. AN SSSR.
42.
Pumpyanskaya, L. V. 1938. Rasprostraneniye drozhei v raznykh pochvakh SSSR v svyazi s ikh fiziko-khimicheskimi osobennostyami. Doklady VASKHNIL No. 1-2: 41-47.
43.
Rippe], A. 1940. Uber die Verbreitung yon Aspergillus niger insbesondere in Deutschland. Arch. Mikrobiol. II, Abt. G. B. I1, p. 1-32.
44.
Rubenchik, L. I. 1940. "Azotobakter i Yego Primeneniye v Selskom Khozyaistve." Publ. h. AN USSR.
45.
Severin, S. A., 1909. Bakterialnoye naseleniye neskolkikh obraztsov pochv dalekogo Severa. Vestnik Bakteriologo Agronomicheskoi Stantsii 15: 166-178.
46.
Sushkina, N. N. 1949. "Ekologo-geograficheskoye Rasprostraneniye Azotobaktera v Pochvakh SSSR" Publ. h. AN SSR.
47.
Sushkina, N. N., and Gordeikina, N. I. 1972. O sostave mikrotlory primitivnykh vysokogornykh pochv Vostochnogo Pamira. Vesmik Mosk. Gos. Univer~'iteta, No 1, 76-85.
48.
Teplyakova, Z. F., Sitnikova, A. S., and Karagushiyeva, D. K. 1953. Rasprostraneniye azotobaktera v nekotorykh pochvakh Kazakhstana. Mikrobiologiya 22 (3): 164-170.
49.
Teplyakova, Z. F. 1961. Aktinomitsety gomykh i podgomykh pochv Zailiyskogo Ala-Tau. Sbornik "Fiziologiya i Ekolologiya Mikroorganizmov", Publ. h. AN KazSSR, P. 129-138.
50.
Tepper, E. Z. 1963. O bakteriyakh avtokhtonnoi mikroflory pochvy, razlagayushchikh gumusovyye veshchestva. Mikrobiologiya 32 (4): 555-564.
1 18
E . N . Mishustin
51.
Tepper, E. Z., and Koryagina, L. A. 1965. Rasprostraneniye proaktinomitsetov v dernovopodzolistykh i chernozemnykh pochvakh, lzv. AN SSSR, Ser. Biol. 5: 772-787.
52.
Tepper, E. Z. 1969. "Proaktinomitsety, kak Predstaviteli Avtoktonnoi Mikroflory." Dokt. Disser. Moskva.
53.
Tyurin, I. V., and Sokolov, A. V. 1968. Tipy pochv i effektivnost udogrenii, lzv. AN SSSR, Ser. Biol. 6: 644-661.
54.
Waksman, S. A. 1927. "Principles of Soil Microbiology." Williams and Wilkins, Baltimore.
Abstract Microorganisms are characterized by wide ranges of distribution. Some groups, however, are known to have zones of active proliferation, and the development o f specific populations in discrete zones results in rather specific microbial associations in some soil types. The soils formed in warm climates are richer in microorganisms and contain more bacilli and actinomycetes. The spectrum of dominant microbial groups varies in different soil types.
The geography and ecology of soil microorganisms have not been well studied until recently. The paucity of research is evident in a fundamental monograph by Alexander [1 ]. The author could find very limited information on the subject. In his classical investigations, Dokuchayev [ 10] developed the doctrine of natural zones, a doctrine that offered scientific substantiation of the zonal distribution of plants, animals, and soils. However, the attempts of a number of scientists to ascertain patterns of distribution of soil microorganism ended in failure. It is believed that microorganisms are cosmopolitan and that their occurrence in soils is influenced only by factors such as climate, vegetation, etc. [ 16, 18, 45]. At the same time, it was shown that the transformation of organic and mineral substances is accompanied by microbial succession [ 15, 19, 20]. Hence, it can be concluded that the geographic environment, because it affects the processes of organic matter transformation in the soil, must promote the development of different microbial associations. Apart from the geographic factors, each zone is certainly defined by its environmental conditions. According to Kluyver [13] the cosmopolitan distribution of microorganisms does not mean that they occur everywhere in large numbers. Their active reproduction is promoted only by certain conditions. It seems that in different soil types, which are geographically dependent formations, different groups of microorganisms must predominate. 97 MICROBIAL ECOLOGY, Vol. 2, 97-118 9 1975 by Springer Verlag New York Inc.
98
E.N. Mishustin
For a long period of time, we have studied the soil microflora of different climatic zones of the USSR. Most observations were performed in warm periods of the year comparing the soil microflora beginning with the polar desert of the extreme north to the deserts of Middle Asia. We tried to find the specific features in their microfloras by ordinary research methods, which admittedly are not free from a number of generally recognized shortcomings. Seasonal fluctuations of numbers of soil microorganisms was found in all soil types examined. These findings have been reported by us [24, 30, 36]. For obvious reasons, the number of non-spore-forming bacteria show the greatest variation in the course of a season. The population fluctuations among the bacilli are less marked. The factors listed above appreciably affect the number of actinomycetes and microscopic fungi. The periodic changes in the number of soil microorganisms do not obscure the specific features characteristic of the microscopic population of certain soil types. It is obvious that the total number of soil microorganisms increases from north to south. Table 1 gives the approximately average values for the numerical composition of the microflora of a number of virgin soils in the European part of the USSR (A horizon). The data are based on a large number of tests. The bacterial abundance was estimated on beef-extract agar, actinomycetes on starch.ammonia agar and the spores of bacilli on mixed beef-extract and wort agar at a 1:1 ratio (pH 7.0). It was observed that the last medium is good for the differentiation of some Bacillus species. A great humber of bacilli are found in soil in the spore state. Thus, to facilitate the estimation of this group, the culture was obtained from pasteurized soil suspensions (10 min at 70~176 The fungi were estimated on wort agar. It is clear from Table 1 that southern soils are the richest in microorganisms. The somewhat high count in the virgin soils of the extreme north can be linked to accumulation on the surfaces of these soils of partially decomposed plant residues, which in the warm periods of the year form a good substrate for microorganisms, mostly fungi and non-spore-forming bacteria. Further to the south, the bacilli and actinomycetes play a more important role in the microbial community [23, 27, 29, 30]. As shown by our model tests, these microorganisms participate in the later stages of plant residue decomposition. This particular role in decomposition is possibly associated both with their powerful eiazymatic apparatus (particularly in actinomycetes) and their requirement for several growth factors (in bacilli). Since in warm climates the decomposition of plant residues is more intensive, the conditions are more favorable forBacillus and actinomycete reproduction. In a still more southern zone, the number of fungi decreases. Due to the increasing competition of bacteria and actinomycetes with
Table 1
c++ moderate. d+++ intensive.
Brown and sierozems
Desert steppe and desert
a+ extremely. b+ slow.
Chestnut
Dry steppe
Podzols and soddypodzolic
Forest meadow
Chernozems
Tundra-gley and gleypodzolic
Tundra and taiga
Meadow steppe and steppe
Soils
Zone
4490
3482
3630
1080
2140
Total number of microorganisms (X 103]g)
45.7%
45.4%
42.4%
77.2%
94.9%
Non-spore forming bacteria
17.7%
19.4%
21.4%
12.0%
0.7%
Bacilli
36.1%
34.6%
35.4%
8.1%
1.5%
Actinomyeetes
0.5%
0.6%
0.8%
2.7%
2.9%
Fungi
+++a
++c
++c
+b
_+a
Nitrification rate
The Average Number of Ma/or Groups of Microorganisms in Different Soil Types of the USSR
Q ~d
~~
o o u ".
100
E . N . Mishustin
the fungi, the number of the latter decreases in these soils, especially those characterized by neutral reactions. It seems appropriate to count the number of microorganisms, not per gram of soil, but per gram of humus since the metabolism of saprophytic microorganisms is associated not with the total mass of soil but with its organic matter. Using such estimations, the high "biogeny" of southern soils is made more evident (Table 2). Data obtained by the conventional technique of microorganism estimation by the use of solid media cannot be considered as absolute, but the general trends noted here can be recognized as fully objective. The higher density of microorganisms in southern soils is supported also by the data obtained by other methods, i.e., direct counting according to the technique of Winogradsky [9] and inoculation of a solid medium by fine earth according to the procedure of Novogrudsky [39]. The resulting absolute indices are, of course, different but the general trends remain the same. Soil types of some mountain ranges in the Caucasus and Middle Asia also have been studied. The effect of altitudinal zonality on the composition of soil microflora is the same as that of latitudinal zonality. Higher in the mountains, where the average annual soil temperature is lower, both the number of bacteria and the percentage of bacilli and actinomycetes decreases [21, 33]. Some soil types have different profiles and distributions of organic matter. These differences affect the microbiological profile of different soils. For example, in chernozems the microbiological profile is much deeper than in podzolic soils. Usually the humus content decreases with soil depth in a more gradual way than the number of microorganisms. Passing on to the characteristic of certain groups of soil microorganisms, I shall comment on the non-spore-forming bacteria. From data obtained in a study employing the commonly used media, the non-spore-forming soil bacteria are usually found to be represented by the genera Pseudomonas, Bacterium, and Mycobacterium. Up to now, we have failed to determine the differences in the composition of non-spore-forming bacteria in certain soil types. It is not unlikely that there are no essential differences whatsoever. The non-spore-forming bacteria are pioneers developing on the decomposing organic residues. Since the presence of fresh organic matter (mainly plant residues) is typical of all soil types and the differences in the chemical composition of this organic matter are not very great, it may well be that identical non-spore-forming bacteria develop in different soil types. Recently it has been found, however, that mycobacteria predominate in the primary phases of the soil-forming process [47]. It should be noted that the composition of the non-spore-forming soil bacteria require additional study. It has been already mentioned that more extensive decomposition of organic residues favors the reproduction of bacilli and actinomycetes. Thus, it might be expected that the specific or group composition of these microorganisms would be
Tundra-gley and gleypodzolic
Podzols and soddy-podzolic
Chernozems
Chestnut
Brown and chernozems
Forestmeadow
Meadow steppe and steppe
Dry Steppe
Desert steppe and desert
Soils
Tundra and taiga
Zone
Table 2
218500
99460
57450
32480
42800
Total number of microorganisms
101500
421 O0
26250
25040
40540
Non-spore forming bacteria
38500
20700
11250
3960
260
Bacilli
77500
36000
19500
2700
600
Actinomyeetes
The Average Number of Microorganisms Calculated per Gram of Humus (IX 103/g)
1000
660
450
780
1400
Fungi
7=
O
o
>
c)
102
E.N. Mishustin
dissimilar in soils of different climatic zones. As is clear from the above, the number of bacilli and actinomycetes increases appreciably in southern soils. Our expectations were realized. Table 3 shows average numerical values based on a large number of studies of groups of spore-forming bacteria characteristic of different soil types. Only virgin soils are considered here. As will be discussed below, cultivation brings about significant changes in the microbial community of soil. All bacilli are cosmopolitan, but for each species there are zones of optimum reproduction. Thus the group B. agglomeratus - B. cereus is rather characteristic of the soils of the extreme north. B. mycoides and B. cereus occur in large numbers of podzols of the central part of the USSR. In chernozem soils, intensive reproduction of the group containing B. idosus and B. megaterium is observed. The group B. mesentericus - B. subtilis is found in chestnut and grey soils. In addition, these soils contain a great number ofB. megaterium cells. Apart from these bacilli, in each soil type are found a certain number of cells of other species. The data obtained are quite convincing. Nevertheless, the groups can be further divided. For example, B. mesentericus is actually a combined species and can be divided into several subspecies. Recently, the distribution of different B. mesentericus strains in the soils of the European part of the US SR was analyzed in our laboratory. The results showed that their distribution is also associated with certain soil types. Thus, B. mesentericus niger is typical only of southern soils. Other strains of this species have a wider distribution range. The specific distribution of certain microbial groups is related to their physiological peculiarities. The bacilli able to use mineral nitrogen (B. megaterium, B. subtilis, B. mesentericus, etc.) obviously predominate in soils with intensive mineralization. The spore-forming bacteria which require organic nitrogen (B. agglomeratus, B. cereus, B. mycoides, etc.) obviously predominate in soils with slow rates of mineralization. In soils of altitudinal zonality, a similar shift of bacterial forms is observed, but since these soils are not quite identical with the zonal ones, certain deviations are observed. Thus, the mountain-meadow soils, like the tundra soils, are poor in B. mycoides but rich in the B. cereus - B. idosus group. B. mycoides occurs in mountain chernozem soils and is practically absent in common chernozems. Recently, the distribution in USSR soils of anaerobic spore-forming bacteria of the genus Clostridium was studied [37]. In the ecology of these microorganisms, certain regularities were also found. Northern soils proved to be much richer in these bacteria than southern soils. In the northern zone, the soils contain chiefly C. pasterianum and a large number ofC. butyricum cells. In southern soils the dominant form is C. acetobutylicum. Table 4 gives average data illustrating the general ecology of bacteria of the genus Clostridium.
Table 3
Gley-podzolic
Podzols Soddy-podzolic
Chernozems
Chestnut
Brown soil and sierozems
Solonetz
Taiga
Forest-steppe
Meadow steepe and steeppe
Dry steppe
Desert steppe
Semidesert
4
1
10
4
[26 [ 25
150[
~
0
0
0
0
10 17
4
0
~
12]
0
~
4
0
~
4
0
6
4
0
0
0
0
30
40
33
63
20 37] 0 21 I 0 - ~
[25
0
~
aThe species making up the typical soil community are outlined. b 0 Means that the species occur in small numbers. CBacilli are differentiated according to Krasilnikov's guide (I 949).
Tundra-gley
Soils
Tundra
Zone
34
26
16
17
0 5
0
0
~
7
8 I
8 [
0
0 0
0
0
4
0
0
0
0 0
0
0
~ ~" ~
--
4
0
0
0 0
0
0
~
~
I
10
8
4
3 4
2
2
~
3 I 10
10
10
4 5
3
4
~'
5
15
-
15
-
42
Species
Unidentified
Composition o f the Spore-Forming Bacteria in A Horizon of Different Soils {~ o f Total Number of Bacilli)a, b,c
%o
o.
o
w,
>
o
104
E.N. Mishustin Table 4
Bacteria of the Genus Clostridiurn in Virgin Soils of Different Horizontal Zonalities (X 1036g) a Butyric acid bacteria
Soil types
Soddy-podzolic
7000
700
20
1
100
30
30
1
10
5
5
10
2.5
5
15
Chernozems Chestnut Sierozems
0.5
C. butyricum
Acetone butyl
C. pasteurianum
Pectinolytic clostridia
(C. acetobutylicum )
a The values shown are the maximum values observed.
As is clear from model tests, C.pasterianum develops intensively during the first period of plant residues decomposition, and C. acetobutylicum during the final period. The latter prefers a protein-containing medium and requires a complex of vitamins, and thus it can reproduce only on organic residues adequately enriched with microbial protein and vitamins. Thus, it is natural that southern soils (richer in microorganisms) are more suitable for C. acetobutylicum and northern soils for C. pasterianum. Since the clostridia studied are nitrogen-fixing bacteria, I shall dwell briefly on the ecology of other free-living nitrogen fixers studied by us, namely, Beijerinckia and Azotobacter.
Beijerinckia was found only in the subtropical red earth of Georgia, soils having a low pH value. This genus occurs there in both virgin and cultivated soils. In humid and hot climates, large numbers of Beijerinckia are readily found in the soil at any season of year. There are many publications dealing withAzotobacter and its distribution in different soil types of the USSR and of many other countries [6, 12, 22, 35, 44, 46]. Azotobacter is rather fastidious in its particular nutrition. It develops in soils rich in available phosphorus compounds, having a pH close to neutrality, containing organic substances, provided with water, etc. A complex of such conditions is not common, even in the zone of rich chernozem soils. Thus, it is not surprising thatAzotobacter is not always found in podzolic soils and that it occurs mostly in neutral soils of flood-plains and highly cultivated lands. In the zone of chernozems
Microbial Associations of Soil Types
105
and more southerly soils, it occurs regularly only in irrigated and sufficiently mostened soils. In the nonirrigated chernozems, chestnut soils, and sierozems, Azotobacter reproduces only in the spring. Apparently, its appearance mainly depends on the increased availability of water. Because of this relationship and a number of others, the organism is with full reason brought together with the algae. A similar picture is observed in chestnut and sierozem soils [22, 48]. From the discussion above, it is clear that a true picture of the ecology of Azotobacter can be obtained only through regular observations. An analysis of randomly collected soil samples without consideration of soil condition results in meaningless conclusions on the ecology of Azotobacter. In general, the distribution of Azotobacter is not related to any particular soil-climatic zone. It may be found both in the north and in the south but only in highly fertile soils. The presence of this organism is a good index of agronomically valuable soil properties. The evidence presented supports the conclusion that some microorganisms, such as Azotobacter, do not reflect the soil type but rather certain ecological conditions. The effect of geography on the distribution of microscopic fungi in soil has been considered in a number of studies. Some points relating to the distribution of microscopic fungi in soil have become generally recognized. For example, it is generally accepted that the variety of soil fungi increases in southern soils, although their total number in these soils markedly decreased [ 17]. It has been found that fungi of the genusAspergillus are largely absent from northern soils and are rather common in southern soils [43, 54]. During the studies of soils of different zones, a considerable amount of information on fungal ecology was collected in our laboratory. A part of it has already been published [25]. The data are summarized in Table 5. An analysis of the results allows for the establishment of a number of generalizations. Thus, in soils with nonintensive mineralization, the relative abundance of fungi of the genera Penicillium and Mucor increases. Further to the south, fungi of the genus Penicillium are replaced by fungi of the genus Aspergillus. The probable reason for this change is that representatives of the genusAspergillus are able to assimilate more complex organic compounds. It should be noted that the relative percentage of Aspergillus not infrequently increases in alkali soil located in the chernozem zone.
Mucor strains that prefer organic nitrogen compounds are common in the upper layers of virgin soil containing many nondecomposed plant residues. Mucor ramanianus is found only in forest soils. Representatives of the genus Choanephora occur only in sierozems.
106
E. N. Mishustin Table 5 Composition o f Microscopic Fungal Flora in A Horizons of Different Soils f~ o f the Total Number of FungiJa, b
Zone
Soils
Tundra
Tundra-gley
72
0
3
Taiga
Gley-podzolic, soddy-podzolic
71
0
6
Steppe
Chernozems
71
1
Dry steppe
Chestnut
63
Desert steppe
Sierozems
47
Semidesert
Solonetz and solonchak
48
1
7
2
7
0
11
10
5
0
7
1
7
1
0
6
4
8
4
0
5
~
aOutlined are fungi characteristic of a particular soil type. b0 Means that the genus occus in small numbers.
Of interest is not only the changing number of Penicillium in soils of different soil-climatic zones but the changing ratio of representatives of sections of this genus. In soils with less intensive biochemical changes, the biochemically less active representatives of the genus Penicillium predominate. In soils with less favorable conditions, say, in solonetz (alkaline), solonchak (saline), and takyr (desert soils), fungi of the Monoverticullata section dominate. The section B iverticullata is chiefly largely represented in forest soils. The section Asymmetricullata is widely distributed in most parts of cultivated soils. Representatives of the genus Fusarium are concentrated in soils covered by grass vegetation. Forest soils are practically free of these fungi. Chernozem, chestnut, and sierozem soils are rather rich in representatives of this genus. Thus, it can be concluded that the composition of the estimated fungi undergoes essential regrouping in different soil types. It would be more than desirable to estimate not only the generic composition of microscopic fungi in different soils but the specific composition as well. It can be supposed that the specific features of the fungal community in different soil types could be described still more clearly.
Microbial Associations of Soil Types
107
It is appropriate to make some remarks on soil yeasts. On the basis of our data as well as the results of Pumpyanskava [42] it is possible to conclude that large numbers of yeasts occur in soils rich in partially decomposed organic materials. That is quite natural since yeasts utilize only simple sugars and organic acids. Thus yeasts are rather widespread in northern soils. The study of Babyeva and Golovleva [5] dealing with yeast ecology should be mentioned. The authors have thoroughly studied a diversity of soils beginning with the tundra zone and finishing with the dry steppe, and they confirmed that the largest number of yeasts are found in soils rich in slightly decomposed plant residues and characterized by an acid reaction and rather high moisture content. In other soils, yeasts occur in small numbers. It is of interest that different yeast types are characteristic of certain soils. Representatives of the genus Candida having a mycelial stage in its developmental cycle predominate in acidic podzolic and soddy-podzolic soils. In tundra-gley and boggy soils, capsulated yeast of the genus Cryptococcus are mainly present. Yeasts of the genus Lipomyces are constantly found in grey forest and chernozem soils. In addition, pink yeasts (Rhodotorula) are also often found in chernozems. As regards the diversity of yeast species, the peaty soils are the richest. Yeasts of the genus Torulopsis are constantly foundthere together with other species. This yeast type is not found in other soils. Literature on yeast ecology is far from abundant, but it shows the differences in the prevalence of this group of certain soil types. The general regularities in actinomycete distribution are quite clearly seen from the data collected, and these distributions are confirmed by a number of other studies [40, 49]. The data allow the conclusion to be drawn that northern soils are not only poor in actinomycetes but the composition of the actinomycete microflora is very limited in them. Further to the south, both the number and the diversity of actinomycetes increase greatly. Table 6 gives average data obtained by us in our studies of the actinomycetes of a number of soils. There is no doubt that the data need further definition, but still some conclusions can be drawn from them. Thus, it can be stated that in southern soils there is a great number of actinomycete groups. A higher diversity of species can be also postulated. Representatives of some species of Streptomyces, such as albus, griseus, globisporus, and chromogenes, occur in appreciable numbers in almost all the soils studied. Nevertheless, further to the south, the numbers of S. griseus and S. globisporus decrease. Some actinomycetes are represented in larger numbers either in northern or in southern soils. Thus, the representatives of the S. albidus and S. viridis generally occur in considerable numbers in northern soils whereas S. fradiae, S. flavus, and S. rubroaurantiacus are more often found in the soils of the northern zone. Southern soils of the USSR are undoubtedly richer in pigmented actinomycetes.
14.7
10.5
Solonetz
Solonchak
0
0
0
6.8
4.2
3.5
6.8 0
8.2
1.8 3.
~
0
0
3.5
0
8.8 i
5.9
0
0
0
0
0
0
13.5 10.1 I 0
6.2
o~ O
1.9
0
34.5 0
16.7
28.13 3.5
16.13 0
3.7
0
0
0
0
0
0
Asporogenic
7.7 15.4[ 7.7 0
~"
3.7 20.7 I 0
0
0
I~
3.5 24.1
.51110.41 14.7
0
4.5
0
0
0 2.3
0
0
0
0
aOutlined are the species listed making up the typical soil community. ~0 Means that the species occurs in small numbers.
28.0
0111.3
30.7 115.4 7.7 15.4 0 I 17.8111.7 35.3 11.7 0 r 17.2 17.0 1.9 i 20.0 0 4.5110.1 7.
Chestnut
Typical chernozem
Greyforest
Soddy-podzolic
Podzolic
Soils
~"
Streptomyces Species
Table 6 Composition of Actinomycetes Microflora of Different Soils (% of Total Number of Actinomycetes]
3.5
0
0
0
0
0
z K ~r
O0
109
Microbial Associations of Soil Types
Table 7 Distribution of Pigmented Actinomycetes in the soils of Kazakhstan (% of the Total Number of Actinomycetes) Soil M o t m t a i n - m e a d o w , alpine
Mountain chernozem, virgin Mountain chcrnozen~, cultivated Sierozem, virgin Sierozem, cultivated
Pigmented 21.0 43.4 46.0 46.0 51.2
Nonpigmented 79.0 56.6 54.0 54.0 48.8
According to Tyeplyakova [49], who studied the vertical zonation of the soil microflora of in Ala-Tau mountains of Kazakhstan, the number of pigmented actinomycetes (Table 7) decreases as the absolute altitude of soils above the sea level increases. The specific composition of actinomycetes of different soils has received little study. Nevertheless, from the available data, certain interesting regularities appear. Thus, as reported by Tyeplyakova, Actinomyces (Streptomyces)fumosus, A. candidus, and A. actinoides often occur in mountain-meadow soils, while they are largely absent from sierozems.* A. verticullatus, A. virgatus, and A. longisporous are, however, not found in mountain soils, while they are common largely represented in sierozems. Some species of actinomycetes, such as A. globisporus and A. griseus, occur in all soil types. In my l a b o r a t o r y , m i c r o o r g a n i s m s of the genus Nocardia (proactinomycetes), which are related to the actinomycetes, were studied by Tepper [ 5 0 - 5 2 ] and Aristarkhova [3]. Proactinomycetes are not easily differentiated in ordinary laboratory media. Thus, for the estimation of Nocardia, a nutrientdeficient agar medium was chosen, one similar in composition to the medium used by Winogradsky for the cultivation of nitrite-forming bacteria. The small colonies of Nocardia which develop in this medium are easily differentiated by use of the low power on the microscope. Physiological studies of a rather large collection of Nocardia have led to the conclusion that a considerable number of them can mineralize humus constituents. In agreement with this conclusion, it has been established that in soils having high rate of humus mineralization the percentage of proactinomycetes in soils increases. The data given below show the percentage of microorganisms of the genus Nocardia in the total number of colonies estimated on the media mentioned above. *Usage in the United States differs,
Streptomyces.
andActinomyces as used in this report is commonly designated
110
E.N. Mishustin
As is evident from these data, the soils become richer in proactinomycetes as one progresses from north to south. Cultivation also results in soil enrichment with these microorganisms: % of Proactinomyces 1.9 9.4 6.8 16.0 25.0
Northern peat-boggy soils Soddy-podzolic soils Chernozems Light chestnut soils (virgin soils) Meadow (cultivated soils)
Certain species ofproactinomycetes are abundant in different soils. Thus the soddy-podzolic soils are rich in N. corallina, chernozem in N. symbioticum, and chestnut soils in N. citrica. The increasing number of microorganisms in southern soils and the higher percentage of bacilli, actinomycetes, and proactinomycetes are indicative of more intensive mineralization of organic compounds of soil. A direct index of this greater activity is the more rapid nitrification. It was established by Kostychev [ 14] that nutrification increases from north to south, and his observations are fully supported by our results. The titer of nitrifying bacteria is very low in podzolic and soddy-podzolic soils. The numbers of nitrifying bacteria increase greatly in chernozems and more southern soils. Nitrification processes correspondingly increase in southern soils. A similar phenomenon is observed in mountain soils. While studying the microbiological features of the A horizon in soils of Zailiysky Alatau vertical zonality, Tyeplyakova [49] came to the conclusion that the nitrifying ability of the soils of warmer climates is much higher than that of soils formed under more severe conditions. The estimation of nitrification activity of soils has shown that the quantity of nitrate accumulating per kg of humus in the mountain-meadow soil in a three-week period is 1.2 rag, in chernozem 6.9 rag, and in zierozem 62.2 mg (Table 8). Thus, the soils that are richer in microbial biomass are more actively forming inorganic compounds of nitrogen. Table 8
Nitrification in Different Virgin Soils of Zailiysky Alatau Soil
Mountain-meadow Mountain-forest Chernozem Dark-chestnut Sierozem
Depth (cm)
Amount of NO~ mg per kg of humus
0-10 0-12 0-13 0- 9 0- 6
1.2 2.9 6.9 26.5 62.9
Microbial Associations of Soil Types
111
It should be noted that the greater effect of equal quantities of mineral fertilizers in the north than in the south is a very important practical result of the differences in mineralization processes in different soil types [2, 53] This greater influence in the north can be ascribed to the slower rate of formation :of plant nutrients in northern soils. The abundance of cellulose decomposing microorganisms is also indicative of mineralization rates [5] in soil. It is known that cellulose can be decomposed both by bacteria and fungi. Cellulose decomposing microorganisms differ greatly in their demands for available nitrogen, however. Among the cellulollytic microflora, fungi of the generaDematium andPhoma are least specific in their demands, and that is why they predominate in soils with slow mineralization rates. Cellulose decomposing fungi such as Chaetomium and Fusarium require a higher level of nitrate for growth. Bacteria require a still larger quantity of available nitrogen than fungi. Thus, in soils with high nitrate levels, a considerable number of myxobacteria (Polyangium and Myxococcus) appear, and at still higher concentrations, vibrios (Cellvibrio) and myxobacteria of the genus Cytophaga assume prominence. These generalizations apply to soils of the same zone. However, since the rate of mineralization in soil increases from north to south, the relative content of cellulose decomposing bacteria in warmer soil-climatic zones increases greatly compared to that of the fungi. More detailed information on this aspect has been published [30, 36]. In these biological characteristics of certain soil types, we have used data referring only to the A1 horizon of virgin soils. It should be stated however, that the composition of the microflora changes with depth. The deeper soil layers are relatively more rich in bacilli and, not infrequently, in actinomycetes. This is particularly evident in the chernozem and sierozem, the soil types in which these microbial groups are found in large numbers (Table 9). The specific composition of microorganisms is also affected by depth, as can be illustrated from data obtained in studies of by chernozem soil. Thus, some spore-forming bacteria, for example, B. idosus, penetrate rather deep into the soil
Table 9 Number of Microorganisms in the Profile of a Chernozem Soil in the Kharkov region, • 103/g of soil Depth (cm)
0- 5 5 - 10 20- 30 40- 50 70 - 80
Humus (%) Bacteria
9.2 9.1 7.7 4.5 2.7
8950 6650 835 200 147
Bacilli
Actinomycetes
Fungi
815 945 825 169 127
835 1015 126 24 13
37.0 36.5 19.5 17.2 0.3
112
E.N. Mishustin
layer. Other bacteria such as B. megaterium and B. mycoides are concentrated in the upper layer of soil. It is the same with fungi. Fungi of the genus Penicillium are the major representatives of the microflora of deeper soil layers. The genera Mucor, Fusarium, and Trichoderma inhabit chiefly the top layers of soil. Table 10 illustrates the changes in the soil micro flora with depth. It shows the distribution of a number of microorganisms in the profile of a chernozem soil. The data illustrate the effect of geographic factors on the composition of the microflora. Thus, it is necessary to consider the effect of ecological factors and the influence of cultivation on the microbial community. Such factors as mechanical composition of soil, type of vegetation, and increased percentage of salt produce a rather noticeable effect on the composition of the soil microflora. The ecological factors cannot be neglected, but its effect is influenced by a stronger effect of geographical zonality. The type of changes in the microbial community under the influence of vegetation and cultivation can be seen from Table 11, in which some chernozem soils of nearby areas of the Voronezh region are considered. The application of organic fertilizers, etc., produces a strong effect on the soil microflora, and this influence has already been discussed [28, 30, 36] so that only some general remarks need to be made. Cultivation improves the temperature and water-air status of soil, resulting in more rapid mineralization of organic materials, and the soil acquires a number of characteristics typical of more southerly soils. An increase is observed in the number of microorganisms, the percentage of bacilli (particularly the number ofB. megaterium cells), and the percentage of bacteria among the cellulose-decomposing microorganisms. The abundance of fungi is reduced, whereas the nitrifiers become more numerous. The application of mineral fertilizers, especially those containing nitrogen, results in significant changes in the composition of some groups of microorganisms. Thus, for example, the application of ammonium sulfate to the soddy-podzolic soil results in rapid reproduction of bacteria of the genus Cellvibrio and a sharp reduction in the abundance of fungi. Soil fertilization gives rise to a large number of Cytophaga cells, etc. Barnyard manure contains many thermophiles, and manured soils thereby become enriched with this group of microorganisms. Barnyard manure rich in organic substances also stimulates reproduction of a number of microorganisms, including Azotobacter. The data collected allow for the establishment of a complex of microbiological indices characteristic of the soil type and its cultivation status. This calls for a special discussion. Recently the study of the soil community composition has utilized the aid of capillary studies [4, 41] and electron microscopy. In our laboratory, we have used electron microscopy in studies of soil [7, 8, 38], and it has been shown that the number of microorganisms in soil is much higher than is usually revealed by ordinary research methods. A number of " n e w " forms of microorganisms were found, and these have
Microbial Associations of Soil Types
1 13
7~ichoderma
oO
r
r
~
0
0
0
o r
eq. o
o
o
"4
0
oO
t~
Cq
r
r--
--
06
t'-
tz
~o
~
~
c5
t~
tt~
t1%
q~
0"~
tO)
tt3
r
t"q
tr
tl~
c5
c5
o
o
o
o
~
d~
d-
6
6
"o"
t"-
O~
Fusarium
Mucor
Penieillium
tt~
B. i d o s u s
6
tae)
B. m e g a t e r i u m
B. m e s e n t e r i c u s
B. m y c o i d e s
oo
6
114
E . N . Mishustin
Mucor
Chaetomium r~
Fusariu m
r~
Aspergillus
tq
Penicillium
B. mesentericus
B. m y c o i d e s
~.N ~.~
c5o
oc5
~5c5
oo
o
r
r'qo
0
B. megaterium
B. agglomeratis
0 r~
0
Microbial Associations of Soil Types
115
been rather thoroughly studied. What is the relationship between the common, well-studied microorganisms and the " n e w " ones, and is it necessary to radically revise our concept of the soil community? It seems that these questions can now be answered. Winogradsky divided the soil microflora according to its functions into two groups: (a) zymogenous and (b) autochthonous. The first decomposes organic residues, the second, soil humus. The data collected allow one to conclude that the common saprophytic microorganisms (bacteria, fungi, actinomycetes, etc.) must be referred to the zymogenous group that are chiefly responsible for metabolizing the organic matter entering the soil. Furthermore, a specific microflora decomposing only humus does not exist; this function is fulfilled by some saprophytic microorganisms, including representatives ofPseudomonas, Nocardia, and other microorganisms [26, 32, 34, 52]. Thus, the so-called "autochthonous" microflora should also include certain representatives of the zymogenous microflora which are able to decompose humic substances as well as simple compounds. The studies of a number of " n e w " microorganisms carried out in our laboratory suggest that they are mostly oligocarbophiles, developing in media with a reduced concentration of carbohydrates. Obviously, the group mineralizes the residues of organic materials decaying in the soil and can be considered as a satellite. According to the terminology suggested by Zavarzin [ 11 ], these microorganisms can be called the "dispersion microflora." The composition of this group in different soils requires additional study. The author expresses thanks to M. Alexander for the correction of the translated text of this article.
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2.
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116
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Microbial Associations of Soil Types
1 17
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