Interaction of Microorganisms with Soil Colloids
Observed by X-Ray Microscopy
Galina Machulla1, Jürgen Thieme2, Jürgen Niemeyer3
1
2
Institut für Bodenkunde und Pflanzenernährung, Martin-Luther-Universität,
Weidenplan 14, D-06108 Halle, Germany
Forschungseinrichtung Röntgenphysik, Georg-August-Universität, Geiststrasse 11,
D-37073 Göttingen, Germany
3
Fachbereich VI - Geowissenschaften, Abteilung Bodenkunde, Universität Trier,
D-54286 Trier, Germany
Abstract. In an X-ray laboratory study the interaction of bacteria with a sterile
montmorillonite suspension was studied. It was found that the inoculation of
the sterile montmorillonite suspension with a culture of soil bacteria resulted
in adhesion and aggregation of montmorillonite platelets on the surface of
bacterium cells. The platelets appear to be stuck in bacterial slime in a typical
edge-to-face association, parallel to each other. The adhesion is due to the
extracellular polysaccharides produced by the soil bacteria.
1 Introduction
Soils build complex environments that generally contain large amounts of
microorganisms. The viability, activity and mobility of bacteria and other microbes
in soil depend strongly on the extent to which they are attached to the surfaces of
organic and inorganic soil particles. Adhesion of microorganisms to mineral soil
colloids may lead to aggregation of soil mineral components, which improves soil
structure and its stability [1]. This, in turn, may increase again the biological activity
and soil productivity.
In view of the fact that microorganisms excrete extracellular polymer substances, the production of microbial substances is assumed to be the major
mechanism by which bacteria and fungi contribute to aggregation processes. This is
the material that first makes contact between a cell and a surface and which can
cause an irreversible adhesion [2].
It is hypothesized that there are four main types of adhesion between microorganisms and soil solids such as mineral particles. These particles can either be larger than microorganisms, of equal size, or smaller, as in the case of clay particles
[3]. The mechanisms involved in this adhesion are of a physicochemical nature.
They include van der Waals, electrostatic, hydrogen-bonding as well as hydrophobic
interactions [4]. Many polysaccharides can adsorb several particles simultaneously,
and thus bind and flocculate them.
Phenomena of building mineral colloidal flocculates, as well as the microbial
biodegradation of some xenobiotics, can be considered as basic features in the pro-
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G. Machulla et al.
cess of soil and water decontamination [5]. Therefore, studies about the interaction of
microorganisms with mineral colloids are fundamental for the science of microbial
ecology and of great practical importance for the disciplines of agronomy and plant
pathology.
The development of a new technique – X-ray microscopy – has made it possible
to study visually the role of bacteria in building colloidal flocculates as well as the
mode of their association. Due to a much shorter wavelength, X-ray microscopy
provides higher resolution than optical microscopy. Most importantly, X-ray
microscopy has the potential for imaging hydrated specimen with high resolution.
Moreover, the preparation of samples is simple and the biotic-abiotical system can be
studied without distortion.
2 Methods and Materials
The method most generally used in laboratory experiments to observe the interaction
of microorganisms with mineral colloids applies liquid systems. This method was
also used for the present study. The microbes observed in our investigations
represent a species of bacterium Bacillus megatherium on one hand and a mixed
culture of several soil bacteria which are common in most native German soils on the
other. The cell length of Bacillus megatherium is about 4 µm and in pure cultures
bacterial chains could be observed. The culture of soil bacteria includes cells of
different size and shape.
To obtain the bacterial suspension and to study the aggregating ability of soil
bacteria, we cultured Bac. megatherium in a standard nutrient medium consisting of
2 g glycerine, 0.25 g peptone, 0.1 g K2HPO4, 0.004 g CaCO3, 0.3 g NaCl and 0.25 g
MgSO4 per liter of distilled water. The culture was grown for 72 h at a temperature
of 28 °C, and after the incubation a 0.1% Na-montmorillonite (Wyoming montmorillonite) suspension was inoculated with this microbial culture at a 1 : 1 - ratio.
Subsequently this suspension was studied by X-ray microscopy.
The mixed culture of soil bacteria was maintained in a diluted (0.1% or 1.0% of
the original medium) and in the original (100%) nutrient medium after the addition
of the Na-montmorillonite (1g per liter). Test tubes containing 2 ml of this bacteriamontmorillonite mixture were incubated at 28 °C for 48 h. At the end of the incubation period the mode of interaction between microorganisms and clay particles was
determined with X-ray microscopy.
3 Results
In Figures 1 and 2 the mode of cell-montmorillonite association and the influence of
the medium concentration on the delimitation of Wyoming montmorillonite in a
liquid system is shown. It can be seen (Fig. 1, top) that the swollen montmorillonite
aggregates disperse into tactoids (or quasicrystals), which consist of several packs of
crystallites. In general, the crystallites are associated in a subparallel manner (faceto-face) and have to be considered as interactions of clay particles with micro-
Interaction of Microorganisms with Soil Colloids
II - 23
organisms. After the culture of Bac. Megatherium was added to the clay suspension,
a close look was taken at the clay platelets (or crystallites, Fig. 1, bottom) and at the
very small clay particles (Fig. 2, top). It was observed (see Fig. 1, bottom) that the
crystallites are in an edge-to-face association with the bacterial cells. The very small
particles, however, are located on the surface of the microbial cells and in between
(Fig. 2, top). In these location the concentration of microbial slime is higher then in
the surroundings. Large quantities of polysaccharide secretion are seen as several
microns thick “shadows“ around the microbial bodies. The extracellular
polysaccharide slime and its ability to bind clay particles has also been observed by
means of ultra-thin section and low-temperature scanning electron microscopy [1, 2].
Clay particles in soil are a main source of nutrients for microbes. The microbes
obtain these nutrients through biological weathering processes. These processes are
of a biochemical nature and result from the secretion of acid metabolites. For pHvalues smaller than 5, it has been reported that mineral destruction by complexation
or dissolution takes place [6]. This biochemical weathering leads to a destruction of
clay quasicrystals, which fall apart into thinner subparticles (Fig. 2, bottom). In the
X-ray micrograph, fully dispersed montmorillonite tactoids in the 1.0% nutrient
solution can be observed. This 1.0% solution appears to be the optimal microhabitat
for an active mixed culture, since it destroys clay mineral tactoids completely. Soil
bacteria cultured in the 0.1% solution, however, appear to be inactive (Fig. 3, top),
because of the extremely low nutrient concentration, whereas in the case of the 100%
solution (Fig. 3, bottom) they find sufficient nutrients in the solution. In both
nutrient solutions (0.1% and 100%) the montmorillonite suspension keeps therefore
its original appearance. It thus seems possible that mineral destruction by soil
microbes occurs in particular, whenever the microbe population begins to starve after
there is a rapid drop of nutrient concentration in the soil solution.
4 Conclusions
1. X-ray microscopy allows the direct visualization of bacteria in soil, their extracellular polymer substances in microsystems, as well as the affected mineral
aggregates and particles.
2. The polysaccharide secretion is noticed as a white „shadow“ that surrounds the
cells.
3. The inoculation of the sterile montmorillonite suspension with bacteria resulted
in the adhesion and aggregation of montmorillonite platelets on the surface of the
cells. The platelets appear to be attached in the bacterial slime in a typical edgeto-face association with a parallel orientation to each other. This result seems to
be due to the extra-cellular polysaccharides produced by the soil bacteria.
4. By secretion of organic polymers and by physicochemical action, microorganisms
can change the organization and physical characteristics of the media in which
they live.
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G. Machulla et al.
Fig. 1. The interaction of soil microbes with colloidal particles: (top) montmorillonite
aggregate dispersion; (bottom) polysaccharide secretion and crystallite adhesion caused by
Bacillus megatherium.
T - tactoid, C - crystallite, P - small clay particles, B - bacterial cell associated with clay
crystallites in an edge- to-face manner.
Interaction of Microorganisms with Soil Colloids
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Fig. 2. The interaction of soil microbes with colloidal particles: (top) small clay particles
adsorption in polysaccharide slime; (bottom) tactoids destruction in a 1.0% nutrient solution.
T - tactoid, C - crystallite, P - small clay particles, B - bacterial cell associated with clay
crystallites in an edge- to-face manner.
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G. Machulla et al.
Fig. 3. Microbe - clay mineral system in the 0.1% (top) and 100% (bottom) nutrient solutions.
Interaction of Microorganisms with Soil Colloids
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