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Chapter 6. Soil Microorganisms and Plant Roots
FRANCIS E. CLARK
VI. Modification of the Root Surface Microflora . . . . . . . . . . .
1. By Seed or Soil Inoculations . . . . . . . . . . . . . . . .
2. By Soil Treatment . . . . . . . . . . . . . . . . . . . .
3. By Plant Treatments . . . . . . . . . . . . . . . . . . .
4. By Application of Herbicides and Insecticides . . . . . . . . .
VII. Influences of the Rhieosphere Flora on Succeeding or Aswciated Plants .
1 . Persistence of the Rhieophilic Flora in Field Soils . . . . . . . .
2. Persistence of Changes in the Soil Environment Brought About by
the Rhizosphere Flora . . . . . . . . . . . . . . . . . .
References. . . . . . . . . . . . . . . . . . . . . . . .
For many years the plant root has been regarded as an absorbing and
anchoring organ, between which and the surrounding soil there exist
numerous relationships. Concepts concerning these relationships must be
subjected periodically to scrutiny. Certain clarifications have already
been made. I n the uptake of nutrients by plant roots, it was formerly
considered that mineral salts entered the cell sap of epidermal cells and
root hairs by processes of simple diffusion from the soil solution. Such
processes have been found inadequate to account for the transfer of
nutrient materials into the root, particularly since solutes may be encountered in the cell solution in higher concentration than in the soil
solution; accordingly, other physical as well as metabolic explanations
are now offered. Further, with the discovery of the phenomenon of base
exchange, it has become recognized that cations come largely from absorption sites in the clay lattice and not simply from the soil solution.
A concept relatively little emphasized is that plant roots are in contact
not merely with a physicochemical environment, but with a microbiological environment as well.
Root hairs, as well as root surfaces, are covered with a mantle of
microorganisms, which affect the growth and welfare of the plant in many
ways, and the activities of which in turn are subject to influences of
higher plants. The concentration of soil microorganisms upon plant root
surfaces is intense, and a t times may reach such proportions that the
roots themselves possess no direct contact with the soil solution. Such is
the case with tree roots that become encompassed by mycorrhizal-forming
fungi. For other roots, the ensheathment is less complete, but, nevertheless, the region of contact between plant roots and the surrounding
soil remains a region of microbiological interest. It is here that the soil
microorganisms may exert their most direct influences upon plants.
Here any beneficial products of microbiological decompositions or syntheses are in immediate contact with the root absorbing surfaces; here
also, any toxic or injurious substance of microbial origin, or any com-
SOIL MICROORGANLSMS AND PLANT ROOTS
petition by microorganisms for nutrients, directly challenges t,he growing
It is beyond the purpose of this review to consider either the manifold
activities involved in microbiological transformations of organic and
inorganic subst.ances in soil, or the physiological responses of plant root$.
Its purpose primarily is to consider the extent t o which the root surface
microflora, by reason either of its composition or its proximity, influence
plant growth and welfare. There is also offered a brief characterization
of the microflora encouraged by growing roots, and a discussion of the
persistence of this microflora under field conditions.
1. Historical Summary
Interest in the microbiology of plant roots dates rather sharply from
Hellreigel and Wilfarth’s report in 1888 on the symbiosis between root
nodule bacteria and leguminous plants. Successful explanation of the
fixation of atmospheric nitrogen by symbiotic bacteria served not only t o
stimulate research in soil microbiology as a branch of science, but it also
led microbiologists to expect that similar relationships, equally clear-cut
and dramatic, might be established between soil bacteria and other plants.
I n the decade following Hellreigel and Wilfarth’s contribution, claims
were made that nonlegumes could be inoculated profitably with diverse
bacteria. Although such claims failed to win general acceptance, they
have not failed to attract enthusiastic supporters even to this day. A
further debate, more academic than that concerning the bacterial inoculation of nonlegumes, developed at the opening of the twentieth century as
to whether there was a specialized microflora associated with plant roots
or whether there was merely a stimulation of the general soil microflora.
This controversy was nonproductive excepting for its emphasis upon the
association of microorganisms and plant roots, and the resulting designation of this ecologic region by Hiltner in 1904 as the rhizosphere. The
rhizosphere is defined as that soil region inside which the soil is subject
to the specific influence of plant roots.
Knowledge concerning the microflora of the rhizosphere developed
slowly during the first quarter of the current century. There were scattered observations that the bacteria associated with roots might affect
plant development. The possibility was suggested that the root microflora, by its contribution to the carbonic acid production in soil, might
affect the solution and availability of mineral salts. It was also admitted
that the rhizosphere population might compete directly with plants in the
assimilation of nutrient elements from soil, and there was observed a
depression of mineralization of nitrogen in soil under the influence of
FRANCIS E. CLARK
A series of papers by Starkey (1929a-l929c, 1931a, b) in the period
1929-1931 called attention to the many microbiological problems afforded
by the rhizosphere. Stmarkeyboth reviewed the earlier literature, and
presented his own extensive data concerning microbial numbers and
activity in the root zone and the unequal stimulation by roots of different
microbial types. He observed that type of plant, age and condition of
any given plant, and proximity to roots influenced microbial activity
in soil; he also emphasized the possible importance of the rhizosphere
flora to the growing plant.
More recent investigations on the rhizosphere have been fostered by
a diversity of interests. Russian microbiologists have developed an extensive program of seed treatment wit,h nonsymbiotic bacteria, claiming
that such inoculation increases yields; they have also been interested in
the relation of the rhizosphere microflora to the formation of a stable soil
structure. Some workers in t,he U S . Department of Agriculture have
studied the saprophytic microflora of the rhizosphere and its relationship
to differences in resistance of plant roots to soil-borne plant pathogens.
Canadian investigators have been similarly intereeted, and t.hey have
also considered the rhizosphere in relation to the nutritive requirements
of soil bacteria. Workers in several countries have considered the role
of the rhizosphere flora in the uptake or availability of nutrient. materials
Krassilnikov (1940) has emphasized the extent to which bacteria
overgrow root surfaces and has reviewed the literature concerning influences of microorganisms on the growth of plants. Kabznelson et aZ.
(1948) have reviewed the more recent literature concerning the microflora of plant roots, particularly that dealing with t,he preferential stimulation by higher plants of certain types of soil organisms.
2. Characterization of the Rhizosphere
The diversity of interests in rhizosphere microbiology has led to a
profusion of experimental techniques and to differences in expression and
interpretation of results. It becomes desirable, therefore, to attempt
some orientation of the biology of the rhizosphere before undertaking to
discuss, firstly, t,hose activities of microorganisms that may affect plant
welfare, and secondly, those alterations within the soil microflora that
are occasioned by plant growth and development.
Hiltner’s definition of the rhizosphere, stated above, refers primarily
to soil adjacent to plant) roots. Nevertheless, nearly all microbiological
studies of the rhizosphere by cultural procedures have included both roots
and soil as material for study. Referring indirectly to the rhizosphere,
Thorn and Smith (1939) speak of “that ball of earth, filled by the roots
SOIL MICROORGANISMS AND PLANT ROOTS
of a particular plant, with the microorganisms that accompany them."
Photomicrographs of Cholodny slides recovered after burial within the
root zone show large numbers of microorganisms to be associated intimately with the root surfaces, very much as an enveloping sheath, with
the surrounding soil relatively sparsely colonized. Both Starkey (1938)
and Linford (1942) have published excellent photomicrographs of associations of microorganisms and roots as shown upon glass slides exposed
within the root zones of growing plants.
Cultural data also show the importance of proximity to roots upon
the magnitude of microbial populations encountered. Representative
data for the cotton plant rhizosphere are shown in Table I. Starkey
(f931a) has published microbiological data for other plants which serve
equally well to emphasize the immediate localization of microorganisms
on root surfaces.
Perot,ti (1926) , in attempting to establish boundaries for the rhizosphere, has considered it to be bounded on one side by the general soil
region, or edaphosphere, and on the other, by the root tissues, or histosphere. Although a sharp line cannot be drawn between the edaphosphere
and the rhizosphere, practically, the boundary may be considered to be
reached wherever plant roots can no longer be shown to have an observable influence on the soil flora. I n Table I, with sample (a) considered
as representative of t,he soil flora apart from and uninfluenced by roots,
Occurrence of Certain Types of Bacteria in Relation to Proximity to Cotton Roots"
Description of sample
(a) Soil, 10 to 15 cm. distant
from roots . . . . . . . . . . . . . . . . . . . .
(b) Soil, 5 to 10 cm. distant
from roots . . . . . . . . . . . . . . . . . . .
(c) Soil, 2.5 to 5 cm. distant
from roots . . . . . . . . . . . . . . . . . .
(d) Soil, 0.5 to 2.5 cm. distant
from roots . . . . . . . . . . . . . . . . . .
(e) Soil, 0 t o 0.5 cm. distant
from roots . . . . . . . . . . . . . . . . . . .
( f ) Root surface scrapings . . . . .
bacillus b Ratio
After Clark (1940).
Hundred thousands per g. of air-dry soil or per g. of root surface scrapings.
FRANCIS E. CLARK
samples (b) and (c) would not be included in the rhizosphere on the
basis of microbiological data presented for them. If appreciable cultural
or direct microscopic differences are not required as criteria in delimit,ing
the rhizosphere, then it must be considered to be the entire field soil at
least to the depth penetrated by plant roots, and perhaps even further,
inasmuch as such soil is, in the main, under the influence of roots. Past
experience and usage is justifiably against such an extensive definition.
The histosphere, which in Perotti’s scheme serves as t.he inner boundary of the rhizosphere, has in the past been considered normally to be
free of microbial cells. Exceptions, to be discussed shortly, occur in
parasitism, in mycorrhizae, in bacteriorrhizae and the nodulation of
legumes, and, of course, during the terminal disintegration and decay of
the root system. Some recent work makes it appear probable that the
interior tissues of plant roots are more commonly colonized by microorganisms than has heretofore been recognized.
Perotti (1926), and almost wit,hout exception other writers publishing
on the rhizosphere, include under that term the det.ritus on the root
surface, sloughed or dying cells, and the microorganisms closely attached
to the root surfaces. Inasmuch as nutrient material coming from the
roots is primarily responsible for this immediate microbial concentration,
this mantle of microorganisms attached to the root surface is undoubtedly
the most important part of what has been called the rhizosphere microflora. Accepting the root surfaces as responsible for the bulk of the
microorganisms found in those combined soil and root samples taken
collectively as the rhizosphere, it would be more logical to speak of this
microflora in terms of the root surfaces than in terms of the soil adjacent
to the root# of plants. This goal can be obtained by the use of the term,
rhizoplune. The rhizoplane is defined as the external surfaces of plant
roots together with any closely adhering particles of soil or debris.
The introduction of the term, rhizoplane, by shifting the microbiological emphasis from the soil around the root to the root surfaces themselves, provides those working either with roots or root surface scrapings
with a means of considering their data on other than a soil basis. The
frequent employment in the literature of the phrase “the microorganisms
associated with root surfaces” indicates an unwillingness to ascribe t o
the surrounding soil the microbes present on root surfaces. Certain workers (Graf, 1930; Poschenreider, 1940) have used the terms “outer rhizosphere” and “closer rhizosphere” in describing sites of microbial
concentration. Berezova (1941) considered there were two distinct zones
in the rhizosphere supporting appreciably different microbial populations
- o n e zone was the adjacent soil, the second, the root surface itself. The
writer is of the opinion that by using the term rhizosphere to describe
SOIL MICROORGANISMS AND PLANT ROOTS
the Soil region adjacent to plant roots, and the term rhizoplane to denote
the plant root surfaces, much of the uncertainty and confusion in the
literature can be avoided.
11. TYPESOF RELATIONSHIPS
Microorganisms associated with plant growth cannot be classified
rigidly as to whether they occur within, on the surface of, or only at a
distance from plant roots. The site a t which any given microorganism
occurs may vary with changes in environmental or physiological conditions. Nor can soil microorganisms be classified rigidly as to the type
of relationship exist,ing between them and plant roots. These relationships may range all the way from chance association to symbiotism or to
1 . Symbiotism
Symbiotism between microorganisms and plant roots, in the sense of
mutualism or benefit to both partners of the associat,ion, is best known
in mycorrhizal formations and in the nodulation of legumes by
Rhizobium. A mycorrhiza is defined as the symbiotic association of a
fungus with the roots of a seed plant, with the symbiosis probably of
mutual benefit, though not unquestionably proved to be so. Where the
hyphae of the fungus form an interwoven mass investing the root tips,
they constitute an ectotrophic mycorrhiza, where they penetrate the
parenchyma of the roots, t.hey constitute an endotrophic mycorrhiza. A
mycorrhiza is the morphologic combination of a fungus and a root. The
term mycotrophy denotes the processes of plant nutrition by means of
mycorrhizae, or the tendency of fungi and roots to form associations of
functional benefit. The literature on mycorrhizae and mycotrophy is
voluminous. A number of review articles have been published; it will
suffice here to mention those of Rayner (1927), Hat.ch (1937), and
Bacterial associations with roots, analogous to those formed by fungi
in mycotrophy, are termed bacteriorrhizae. Very little is known concerning the actual existence or extent of bacteriorrhizae, apart from the
root hair invasions and the resulting localized nodule formations by
rhizobia. Nodules on legumes caused by rhizobia may be considered to
represent a localized bacteriorrhizal condition. Because of their practical
importance, the rhizobia have been studied more extensively than any
other group of soil microorganisms. They are commonly referred to as
the legume bacteria, and their symbiotic relationships with their host
plants are generally acknowledged. Monographs on the nitrogen-fixing
FRANCIS E. CLARK
bacteria have been prepared by Fred et al. (1932), and by Wilson (1940).
Whether actinomycetes form symbiotic associations with roots similar
to those formed by fungi and bacteria is unknown. There remains the
possibility of the eventual discovery of an “actinorrhiza.” Lutman and
Wheeler (1948) observed invasions of cells of potato tubers by microbial
filaments, which they first thought to be actinomycelial in nature, but
which yielded bacilli upon culture. Sanford (1948) has recently reported
that the interior tissues of the stems and tap roots of potato, lucerne,
and sweet clover contain a mixed bacterial flora. T e y e t and Hollis
(1948) also have encountered bacteria in healthy storage organs of
potato and other plants.
The occurrence of fungi, actinomyces, bacteria, and viruses injurious
to plant roots has long been recognized. References and discussions concerning the phytopathogenic microflora of plant roots are available in
the numerous textbooks and monographs on plant diseases.
Because of the excretion of inorganic and organic substances, and the
sloughing of root caps, root hairs, cortical and epidermal cells, all of
which supply available energy material for microbial utilization, a great
number of microorganisms are encouraged to develop immediately upon
the root surfaces, and the majority of these microorganisms are undoubtedly commensalistic in their relationship to the higher plant, living a t
the expense of secreted or excreted material, but not invading the root
tissues with either injurious or beneficial results. Certain workers have
emphasized the scavenger role of microorganisms on root surfaces ; others,
the existence of a successive symbiotic effect, rather than a true or
Undoubtedly the relationships between microorganisms and plant
roots are variable, depending upon environmental and physiological conditions. Microorganisms normally commensalistic may become parasitic
with decreased vigor of plant growth. Similarly, mycorrhizal invasions
vary from absolute parasitism to complete symbiosis, depending upon
t.he vitality of the fungus and the health of the higher plant (McArdle,
1932). Thornton (1935) has suggested that eventually it will be possible
to trace an evolutionary gradient from the purely ecological relationships
that exist between free-living soil bacteria and higher plants through
that too little studied population of organisms existing outside but in
close contact with the roots of higher plants, to those microorganisms
such as mycorrhizal fungi and rhizobia that live symbiotically within