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Accurate prediction of the response of soil biological communities to environmental perturbation is predicated upon the premise that an accurate or at least a reasonably representative understanding of the ecosystem properties controlling soil processes has been developed. This prerequisite presents a meaningful conceptual problem to those seeking to understanding basic soil microbial processes. Microbiological principles and metabolic reactions are understood at the level of the individual microbial cell, colony or the molecular level (i.e. the microlevel), whereas soil properties delimiting the expression of microbial potential are assessed at a macrolevel (e.g. ranging from several grams of soil to a total ecosystem characterization). Thus, development of a true picture of the effect of soil microbes on large‐scale ecosystem function requires extrapolation of the microsite and macrosite processes. These three spheres of concern (microsite, ecosystem‐wide, worldwide) are not always easily merged, but such reconciliation of data is essential. The task would be simpler were microbial function controlled by one or a few physical, chemical or biological soil properties. In reality, microbes in soil are responding to a combination of soil properties acting in concert. For example, it is relatively easy to dismiss a proposed 1–3 °C rise in soil temperature due to climate change as being insignificant. Yet, soil temperature variation due to climate change has a large effect on the hydrological cycle (occurrence of droughts or floods as well as the seasonal variation of soil water content) at a local level. Additionally, the anticipated declines in soil water result in increased soil salinity, osmotic and matric potential impacts on microbial activity (e.g. Asghar et al. 2012; Chowdhury et al. 2011). Clearly, the soil microbe is encountering a complex, stressful environment.
Macro‐ and microsite reality and ecosystem heterogeneity: A primary difficulty associated with the merging of microbial and soil chemical and physical analyses results from the high degree of variability of soil properties both in a vertical and horizontal plane within the soil profile. A false impression of the microsite traits may be gained from study of the relatively large soil samples, both in situ at the soil site as well as in soil samples incubated in the laboratory or greenhouse. Values for soil properties, such as pH or cation exchange capacity, are meaningfully variable at microdistances within a soil. Soil properties measured in a sample of soil as small as 10 cm3, for example, may be quite different from those measured in a large soil sample.
This telescoping nature of the application of soil microbiological research reveals a characteristic and, to some degree, an intellectual trap of our discipline. Data are necessarily a summation of a variety of reactions occurring in highly variable soil microsites. Thus, a composite representation of soil processes must result in the emphasis of dominant reactions, but other processes of major importance to ecosystem integrity and function may be overlooked or the importance of their contribution to system complexity minimized. For example, the bulk of terrestrial soils are aerobic. Molecular oxygen diffuses freely into the pores of nonwaterlogged soils as well as being transferred deep into the soil matrix through mass transfer driven by water infiltration. For example, mass transfer of atmospheric gases into soil can occur when groundwater is drained from the soil or as a result of the percolation of oxygen‐bearing irrigation or rainwater through soil pores.
Thus, it is easy to emphasize aerobic processes in conceptual models of ecosystem function to the detriment of anoxic reactions. Yet, we know that anaerobic soil bacteria occur and are active in essentially all soils and that the products of anaerobic microbial activity are essential for ecosystem survival (e.g. denitrification). Our understanding of soil microbial processes is constructed on a seemingly contradictory concept of the system. The occurrence of both aerobic and anoxic microbial processes in close proximity in most surface soils provides an excellent example of this quandary. Obligate aerobic processes, such as aerobic nitrification (oxidation of ammonium to nitrate), occur in close physical proximity in soil to obligatorily anoxic processes, such as denitrification (reduction of the nitrate produced by nitrification to nitrous oxide and dinitrogen). The distribution of aerobic and anaerobic microsites in close proximity in surface soils and the diffusion of nitrate into the anoxic sites following its synthesis by the nitrifiers allows this seemingly contradictory interlocking of these incongruent reactions to occur.
Thus, a foundational concept for this analysis of process control in soil is that all microbial processes are first the summation of the activity of a variety of individuals, whether a cell, multicellular complex (that is, a biofilm), or even a collection of enzymes. Second, the level of activity of the microbe is controlled by the summation of a complex, interacting soil physical and chemical properties. Initial considerations involve an evaluation of the concept of a limiting factor and experimental quantification of the impact of various environmental properties on biological activities. This will be followed by elucidation of the variation of specific soil chemical and physical traits. Soil properties interacting with biological entities to be evaluated herein include nutrient requirements, soil moisture, oxygen tension, redox potential, pH, and temperature.
5.1 Microbial Response to Abiotic Limitations: General Considerations
5.1.1 Definition of Limitations to Biological Activity
The classical definition of an environmental factor that determines the rate of a biological activity can be summarized by Liebig's law of the minimum: “Under steady‐state conditions, the essential material available in amounts most closely approaching the critical minimum needed by a given organisms will tend to be the limiting one” (Dommergues et al. 1978). This principle was originally developed to describe the impact of variation of nutrients on microbial growth and activity, but it has been expanded in recent years to include a number of growth limitors, including temperature and a variety of inorganic and organic metabolic inhibitors.
It must be stressed that the application of Liebig's law to a particular ecosystem is appropriate only if the system is at steady state. This condition is necessitated by the fact that the organisms and their interactions cannot be adequately assessed in relationship to rate‐controlling soil conditions except at steady state; otherwise, populations and/or their activities may be increasing or decreasing to reach the density or activity level allowed by the newly established environmental conditions.
Although it is conceptually attractive, Liebig's law is of limited applicability for describing the dynamics of soil ecosystems. This results from three conditions.
· As exemplified above in regard to the complexity of changes in soil properties due to drying, soil microbes in general must deal with an array of stress factors, only in extreme situations, such as highly acidic or temperature stressed situations would the microbes have the luxury of dealing with a single stress factor.
· This combined stress produces different responses by the microbial community than would be anticipated to result from any soil property acting singly on the microbial cells.
· The structure of the soil ecosystem makes it difficult to determine the exact conditions at the microsite where the organisms are living.
5.1.1.1 Microbial Responses to a Stressful World
Limitations to microbial growth are generally due to a combination of biological, chemical and physical soil properties acting in concert. It is rare for one soil property to preclude life processes. Exceptions to the multiplicity of stresses may be exemplified, to some degree, by consideration of microbial populations in the dry deserts of Antarctica (Benoit and Hall 1970; Cameron 1972) or incidence of extreme anthropogenic intervention (e.g. acidification due to acid mine drainage, nuclear power plant cooling water outflows, or soils receiving massive influxes of heavy metal‐containing mine wastes). Essentially, all soil microbes are subject to compromises in the rates at which they function. More commonly, a combination of several soil physical and chemical conditions stresses soil microbial populations sufficiently to cause them to operate at suboptimal levels. The effect of interacting soil properties on microbial processes frequently cannot be predicted by examining microbial behavior in laboratory culture. It may be predicted from laboratory studies that a microbial population of interest would be unaffected by a particular soil property; yet, in native soils, the growth of the organisms is precluded under seemingly acceptable conditions or its range of occurrence is truncated. For example, fungi grow in culture within the pH range of 3.0–8.0, but in soil the practical range is closer to 3.0–6.5. In this case, the impact of a chemical controller is altered by the microbe's capacity to compete with cohabiting biological populations. That is, the activity of soil fungi is controlled by soil pH plus the organisms' ability to compete with soil bacteria. The latter populations are better competitors at the higher pH than are the fungi. Similarly, some environmental properties may compensate for cellular inadequacies, thereby allowing microbial populations to develop outside the range predicted from laboratory studies for its maximum, minimum, or optimal activity. For example, it is not unusual to isolate soil microbes capable of growth at 30 °C in culture but not at 37 °C in a particular medium. If growth factors (or vitamins) are added to the culture medium, the organisms grow quite well at the higher temperature. This results from the fact that an enzyme essential for the production of the growth factor by the microbes could be temperature sensitive; that is, it is inactive at the higher temperature. Availability of this obligatory growth factor in the environment allows the previously inactive organism to proliferate in the seemingly detrimental habitat. Therefore, if a microbe is growing in the presence of an organism producing the compensating growth factor (e.g. another microbe or evan a plant root), its temperature tolerance will be extended.
A further difficulty with application of Liebig's law of the minimum to conditions in native soil systems is more of a practical concern than a conceptual difficulty. Due to soil spatial and temporal variability, data may imply nonexistent restrictions to microbial growth. Basically, these observations result from the fact that a microorganism must physically encounter an inhibitory or stimulatory soil component before its growth or activity is affected. Microbes generally grow as microcolonies in soil microsites where their needs are met. Thus, microbial growth is limited by the distribution of carbon, nitrogen, and other nutrients; aeration; anoxic conditions; and so forth. Any troublesome soil property occurring within a short distance from the perimeter of the colony is nonexistent from the viewpoint of the microbes. Furthermore, environmental conditions vary on a daily, seasonal, and annual basis. This is easily envisioned by consideration of the rise in temperature in the surface centimeter of soil on an early spring day. In the early morning hours, the temperature may be suboptimal for microbial activity; perhaps the soil is frozen. As the hours pass, soil temperature increases, resulting in a stimulation of biological processes until some optimal or maximal rate is reached. On the rare hot days, temperature combined in association with the resultant increased desiccation rate (soil moisture loss) could present a situation that is inhibitory to the life processes occurring in the very top portion of the soil profile. Similar temporal effects could be described for other variables, such as nutrient concentration, pH, and moisture. Nutrients may be leached through soil during rainfall events that clearly cause dramatic changes in soil moisture levels. Similarly pH variation may occur at the microsite level due to acid production by the microbes themselves or result from a periodic influx of human products, such as acid rain or acid mine drainage waters. Thus, analysis of the soil site at a particular time of day or season of the year may indicate that the growth‐promoting or ‐inhibiting conditions do not exist, whereas if the measurements were taken a few hours earlier or later, contrasting results would be obtained.
These observations indicate the necessity of modification of our concept of a limiting factor to fit the complexities of soil. A more appropriate basis for consideration of control of soil microbial process is the following: “The growth of a soil microbiological population depends on a combination of several limiting factors acting collectively and interdependently at the soil microsite wherein the microbe is functioning.” For example, denitrifying bacteria in soil can be limited by variation in easily metabolizable carbon levels, nitrate concentrations, as well as oxygen concentrations. Autotrophic nitrifiers are frequently controlled by soil pH, ammonium concentrations, and oxygen tension. It is likely reasonable to assume that none of the soil properties at the microsite where the microbes grow are optimal for the organism. Thus, it can be concluded that the specific level of activity of each individual is controlled by the combined effect of nearly all physical, chemical, and biological properties of the site. Again, exceptions would be those times or situations when one soil property occurs at such an extreme value that its effect overrides all others (e.g. soil moisture so low that minimal cellular respiration is possible, or temperatures approaching the maximum tolerated level). In the majority of the situations, though, it is the impact of the combination of limitations acting in concert that determines the extent of microbial population development in a soil site.
5.1.2 Elucidation of Limiting Factors in Soil
Under most circumstances, the complexities of the soil environment as well as the microscopic and submicroscopic size of the soil microbial community preclude clear determination of limiting factors in the field. In some situations, field observations are quite useful (e.g. Antarctic dry deserts or anthropogenically affected sites where extremes in physical or chemical conditions are easily discerned), but for the vast majority of the terrestrial ecosystem, controlled, laboratory‐based study of the system is required to quantify the contribution of each individual soil property to biological reaction rates. Options for determining biological activities in soil include variation of single or combinations of properties in laboratory‐incubated soil samples or assessment of field activities in several diverse systems.
5.1.2.1 Limitations of Laboratory‐Generated Data
Scientific conditioning creates a desire of experimenters to study the effect of variation of each individual soil property on biological processes under controlled conditions in the laboratory. This offers the security of simplicity and reproducibility. In laboratory experiments, soil conditions may be varied singly or in concert, and the resultant impact on microbial activity assessed accurately. Easily manipulated factors can include pH, temperature, moisture, and nutrient levels. These conceptually alluring studies may yield results that describe the capacity of existing microbial populations in the soil system to cope with alteration of their habitat in the short term. The utility of such data for prediction of the resiliency of the native soil population to long‐term perturbation is frequently limited. To achieve the latter objective, soil samples should be incubated for a sufficient time period to allow the microbial community to adjust to the newly imposed conditions, particularly if the chemical or physical properties of the soil are being altered.
The limitations resulting from failure to allow community development in laboratory study are exemplified by evaluating the impact of varying soil pH beyond ranges normally detected in the soil ecosystem studied. It is frequently desirable to determine the pH range over which an enzymatically or biologically catalyzed process occurs. It could appear reasonable that the impact of variation in this parameter could readily be observed by varying the pH of one or a few representative soils. For this experiment, soil could be amended with different amounts of sulfuric acid, yielding soil samples with pH varying at set intervals over the pH range of interest. (Note that in this example, we are not considering any effect of the elevated sulfate concentration on soil biological function.) The biological activity would then be measured. The results of such an experiment are reasonably predictable. An ecological maxim is that the active microbial population in any soil is that which is best capable of coping with the prevailing soil pH. Thus, the most likely outcome of the experiment is that major changes in the soil pH will be detrimental to the soil microbial activity. The experimental data support an a priori conclusion that the biological community in that particular soil sample has a defined pH range in which it is capable of functioning similar to the hypothetical curve plotted in Figure 5.1. Were this be applied to our understanding of the function and resilience (sustainability), we would be lead to a conclusion that soil pH should be rigidly maintained within a rather limited range. The conclusion results more from limitations of the experimental design than actual field situations.
Figure 5.1 Example of variation of an hypothetical biological activity with changes in soil pH. For this representation, an optimum of approximately pH 7 with rapid declines in activity above and below this value is depicted.
In this type of experiment, the potential for development of alternative microbial communities capable of functioning at the newly imposed soil pH is overlooked. Since acidophilic or alkylinophilic microbes exist in low numbers in the original soil sample or be present in a resting state, an alternative microbial community capable of functioning under the newly established conditions can and generally does evolve. An appropriate conclusion for the experiment would be that in the short term, a significant change in soil pH would be detrimental to microbial function. An evaluation of the microbial populations developing over extended timeframes would result in a more realistic conclusion that pH does indeed result in immediate declines in activity but given time, the resilience of the existing population of microbes in the soil allows development of an alternate microbial community optimized for the newly established conditions. That is, the existence of inactive microbial species or strains capable of functioning under the newly established acidic or alkaline conditions in the original soil sample, given appropriate time for growth and induction of the requisite enzymatic activity, increases the adaptability and sustainability of soil systems.
This examination of the effects of pH on microbial populations also provides an excellent example of the complications from the interaction of soil chemical processes on the biological response to alteration of the soil pH. Complications of data interpretation associated with adjustment of soil pH result not only from selection of the existent populations best adapted to the newly established soil pH, but also from the fact that the reaction of community to the alteration of the soil pH may be moderated due to the soil buffer capacity. Sufficient acid can be added to a soil sample to reach an acidic pH selected for a particular experimental design, but if the pH of the soil sample is measured a few hours or days after initial adjustment, it is commonly found that the pH level has returned to a value approaching the preamendment level. The impact of the added acid on the soil pH has been moderated by the soil buffer capacity (see Brady and Weil 2017) for further discussion of this process). Thus, not only is it necessary to incubate the soils for sufficient period to allow selection of acid‐resistant microbial populations, but the natural tendency for the soil to return to native conditions must be considered. Changes of the soil pH during the incubation period may allow continued activity of the native population rather than development of acidophiles or more acid‐tolerant organisms. Thus, to prevent misinterpretation of the laboratory studies, soil pH of the amended soils must be monitored throughout the study.
In conclusion, native soil samples may be adjusted to mimic variation in a particular soil property, but the data produced will reflect capabilities of the native soil community to cope with the change, not the capabilities of an ecologically adapted community. To achieve a measure of the latter situation, the pH‐adjusted soil samples would have to be incubated for months or longer to allow the results of the selective processes to develop fully.
This consideration of microbial community reactions to changes in soil pH also provides an excellent example of the impact of the microbial functional diversity on the composition of the active portion of the microbial community. Changes in a specific soil property, such as pH or even growth of plant roots, can result in emergence of microbial species or strains better capable of functioning under the newly imposed conditions. One could envision the soil microbial populations as consisting of the fully functional individuals best adapted to the existing conditions plus those incapable of competing with their better adapted associates but fully capable of emerging as full participants in community function should the conditions change. This aspect of soil life is essential for the sustainability and resilience of soil life in its changing world. An excellent example of this sustainability/adaptability property of the system is exemplified by the response of soil microbes to short‐ and long‐term variation in soil salinity. Stresses encountered by matric and osmotic potential fluctuations due to water evaporation necessarily induce microbial community activity and composition changes. (See Asghar et al. 2012; Chowdhury et al. 2011; Mavi et al. 2012 and Yan and Marschner 2012 for further information on soil salinity variation and microbial function.)
5.1.2.2 Evaluation of Soil Properties and Microbial Variability in Native Samples
An alternative available to overcome this potential for laboratory‐generated misunderstandings of the complexity of soil community resiliency and adaptability is to study microbial population dynamics in several native soil samples that already exhibit variation of the property of concern. Microbial populations in native soils could be reasonably anticipated to have adapted to the soil site. Thus, if pH interactions with microbial activity were to be studied, it could be assumed that the microbial community existent in each soil is the one best adapted to function therein. A major caveat for such studies is the need to remember that the specific organisms active in a soil sample are those selected by the specific existing combination of soil properties, not necessarily the individual property under study. Selection of appropriate soil samples for these studies is complicated. Except under fortuitous situations, it is rare to find a series of soil sites where only one or a few soil properties vary in an otherwise constant environment. Thus, for study of native soil samples, not only is it necessary to assess the biological activity of interest, but all possible chemical and physical soil properties that could impinge upon that activity must also be quantified.
Problems in data interpretation with these field survey experiments are encountered in that statistically significant relationships may be obtained that are irrelevant. For example, the relationship may be fortuitous in that the variables being measured could have correlated with a third factor that was not considered. A potential solution to this difficulty is that once significant correlations are determined in the field, the data can be verified under more controlled conditions in the laboratory. A further means of confirmation would be to use the field‐collected relationships to construct a mathematical (geostatistically derived) model of the biological activity of the soil ecosystems. Field validation of the model may provide evidence in support of the conclusion that appropriate relationships have been elucidated and quantified.
5.2 Impact of Individual Soil Properties on Microbial Activity
From a somewhat naive point of view, soil appears to present a reasonably accommodating habit for microbial growth, Mineral nutrients, organic matter (including fresh plant material), and growth factors necessary to meet microbial nutrient requirements generally exist within appropriate temperature, moisture, and pH ranges for microbial growth. Also, most soils contain microbial populations with the necessary genotypic capacity to catalyze all requisite reactions for ecosystem development. Unfortunately for the microbe, soils also contain a variety of growth inhibitors – for instance, heavy metals, as well as organic and inorganic acids. Therefore, to fully understand biological activity in soil, the needs of the organisms must be considered in relation to the form and distribution of chemical and physical prerequisites for microbial growth.
5.2.1 Availability of Nutrients
Microbes require an energy source, electron acceptors, trace minerals, and several macronutrients plus, frequently, a variety of growth factors (i.e. vitamins and amino acids that are incorporated into cellular substituents intact) to proliferate. Although these substances are generally present in soil, the extent of microbial community development is determined by their distribution and concentrations within the microsites where the microbes reside. Therefore, the following discussion is presented with the objective of evaluating the biological requirements for growth and reproduction and the availability of these nutrients to soil communities. (Details of several microbial mechanisms for energy recovery and incorporation of micro‐ and macronutrients into biomass are presented in Chapter 4.)
5.2.1.1 Soil Microbial Nutrient Resources
Energy source: The primary energy supply for all living systems is solar energy. Plants and microbes (photoautotrophs) convert this energy into the chemical energy used by chemoautotrophs (heterotrophs) organisms to support cellular growth and replication. Note that due to the diversity of metabolic capacities of soil bacteria, a portion of the soil microbial community is also capable of taking advantage of the nonsolar‐derived energy sources available in soil including reduced minerals (e.g. S0, Fe2+) and products of cellular respiration (e.g. NH4, H2). Typical energy‐yielding transformations of these substances are:
Although there are many opportunities for chemolithotrophic (energy recovery from oxidation of inorganic chemicals) metabolism in soil, the bulk of the soil community is driven by the oxidation of organic substances. Heterotrophic organisms primarily use photosynthetically synthesized organic carbon, most commonly entering the soil ecosystem as root exudates, dead or decaying plant biomass, or soluble organic substances leached into the soil from surface litter. Plant biomass can also become available to soil microbes through anthropogenic intervention such as through soil amendment with manure or composts or with a variety of anthropogenically modified and industrially produced waste materials (e.g. petroleum by‐products and industrial wastes). Because the energy contained in newly synthesized plant biomass is easily recovered by soil microbes, sites receiving these substances could be likened to oases within a desert of biological activity. This conclusion is based on the fact that in the absence of the reasonably constantly supplied source of carbon and energy produced by growing plants, soil microbes generally are left to metabolize the components of soil humus. This latter nutrient source is of lesser quality than the fresh, green plant biomass. Previous microbial activity has reduced the concentrations of easily metabolized organic matter. Thus, the substance remaining, the humus, is enriched in more complex organic compounds and microbial products.
5.2.1.2 Terminal Electron Acceptors
Recall that a primary chemical maxim is that all oxidative reactions must be balanced by reductive reactions (see Chapter 4). The biological reactions catalyzed within soil are not an exception to this rule. The most common electron acceptor in soil is molecular oxygen, which is reduced to water. For facultatively anaerobic organisms, oxygen and organic are terminal electron acceptors whereas the strictly anaerobic bacteria reduce organic carbon acceptors such as organic acids. Some aerobic organisms growing under anoxic conditions reduce nitrate or nitrite (denitrification) or sulfate (dissimilatory sulfate reduction) (see Chapter 4 for further details of these processes).
5.2.1.3 Macronutrient Sources
Following energy sources, the next most abundant requirement for cellular growth and proliferation in soil is a variety of macronutrients (i.e. carbon, nitrogen, phosphorus, and sulfur supplies). These substances are the major building blocks for cellular biomass. Carbon and nitrogen are generally derived from the mineralization of plant biomass, whereas phosphorus and sulfur are not only components of biomass but are also present in high concentrations in soil minerals. Thus, pathways of carbon and nitrogen mineralization are frequently interlinked whereas phosphorus and sulfur supplies and transformation in soil also generally involve both transformations of organic compounds and of soil minerals.
5.2.1.4 Determination of In situ Limiting Nutrients
Evaluation of nutrient limitations in soil communities is usually focused on energy or macronutrient supplies. Typically, documentation of such limitations entails amendment of soil samples with a nutrient that is suspected to control microbial metabolic activity and measurement of microbial respiration and/or growth. If the soil amendment is utilized by the soil microbes, a variety of responses of the microbial community may be observed. An idealized depiction of the type of data that could be recovered from such studies is presented in Figure 5.2. If the microbial population was not controlled by the soil amendment or an alternative factor limited biological activity, curves of type A could be observed. For example, assume that the objective of an experiment is to determine the primary limitation to aerobic microbial respiration in a soil collected from a swamp. Observation that metabolizable fixed carbon compounds occur at extremely low levels in the soil leads to development of the hypothesis that microbial populations are carbon limited.
A typical experiment to test the hypothesis could be to amend the soil with an easily metabolized carbon source, such as glucose, glycerol, or acetate, under the conditions occurring in the field site and observe the effect of the amendment on microbial respiration or growth. When this is done under the anoxic conditions of a swampy soil, no change in microbial growth of culturable aerobic bacteria may be observed. An easy explanation is that the carbon source was not the limiting nutrient controlling microbial growth. In this situation, the absence of the electron acceptor, free oxygen, likely precluded the rapid catabolism of available carbon and resultant multiplication of the aerobic organisms. Thus, prior to soil amendment, the low levels of organic substrate were sufficient to support the amount of microbial respiration occurring therein at a maximal level. Oxygenation of the system or provision of an alternative electron acceptor (e.g. nitrate) could result in development of a carbon‐limited system. The probable primary delimiter of microbial activity (a terminal electron acceptor), in that case, would be relieved by addition of free oxygen or nitrate, (i.e. the available energy source could become the prime candidate or primary obstacle to maximal microbial activity).
Figure 5.2 Hypothetical curves describing response of soil biological activity to amendment with proposed growth limiting nutrients. A: The added substance is not controlling cell replication. B: The additive is nonlimiting and toxic to the biological community. C: The substance was present in the soil sample in growth controlling concentrations. D: The additive is growth limiting, but it is also toxic in elevated concentrations.
An alternative to this neutral response of soil amendment is the situation described by Curve B in Figure 5.2. These data reflect the condition where increases in soil concentrations of a hypothetical controlling nutrient results in inhibition of overall microbial respiration; that is, the amended substance is toxic at higher concentrations. An example of this effect of soil amendment is provided by the frequently observed result of addition of a variety of aromatic compounds to a soil sample. In low concentrations (typically less than 0.1% (wt/wt)), microbial respiration may be stimulated by such compounds, such as catechol or benzoic acid, but as their concentration is increased, microbial respiration is inhibited. Thus, if maximally tolerated levels of these substances already exist in a soil site, further amendment with them inhibits cellular respiration.
More easily interpreted examples of relief of metabolic limitations are exemplified by Curves C and D. In these situations, the test compound controls microbial activity. As its concentration is increased, microbial activity or respiration increases until a plateau is reached. No further increase in activity is detected. At this point, another soil factor has become limiting to the microbial community or population studied. At maximum concentration, the amendment may have no further impact on soil activity (Curve C) or it may become inhibitory (Curve D), as was described above by Curve B. Note that with time, the microbial respiration may again increase. This change in respiration rate would result from induction of new enzyme synthesis by exiting microbes, or given sufficient time, new microbial growth. The prior leveling of the activity could be interpreted to result from the enzyme or microbial populations being the factor preventing the microbes from utilizing the more abundant nutrient and energy supply. Of course, other nutrients or soil properties (such as space for microbial growth) could also have precluded further increases in microbial respiration and/or growth.
Fluctuations in soil microbial populations following clear‐cutting of a forest provide an excellent example of the impact of nutrient availability on biological activity in situ (e.g. Lundgren 1982). Generally, clear‐cutting of the forest initially results in an increase in soil microbial biomass. This microbial response results from the decomposition of excess organic matter, both living and that previously accumulated in soil as plant residues. These substances become available to the microbial community through destruction of the aboveground portions of the plant biomass. Roots die and carbon compounds from the debris produced by the cutting of the trees leach into the soil profile. Furthermore, machinery and logging traffic on the soil surface results in disruption of soil structure, which enhances availability of any organic matter occluded in soil aggregates.
Three microbial communities can be described for this system in transition: (i) the native forest soil community existing prior to anthropogenic intervention, (ii) an expanded community supported by the release of plant debris during logging (a disturbed soil community), and (iii) a community that develops in response to the newly established ecosystem properties (bare soil). In a nutrient‐controlled situation, the disturbed soil community would exhibit maximal population density, whereas the bare soil reflects a nutrient‐ and energy‐limited community structure reflective of the denuded, minimally productive aboveground ecosystem. With time, a new microbial community reflective of the newly established ecosystem emerges. Similar situations could be proposed to exist during the annual cycle of crop growth in a wheat field. In the spring season, the young succulent plants would be providing carbon compounds to the microbial community through root exudates. The nature and quantities of the nutrients leaking from the roots change during the growing season, resulting in alteration of the nutritional pressures imposed on the microbial community. The potential mid‐season reduction of root exudates would be followed by harvesting of the crop when the plant debris and decaying roots would become the primary controller of carbon metabolism.
5.2.1.5 Carbon Limitation of Microbial Activity
Examination of the occurrence of macronutrients in a soil sample (carbon, nitrogen, sulfur, and phosphorus) and their ratios in soil microbial communities reveals that of these substances, carbon resources are generally the primary controller in soil microbial community development. Soil microbial biomass contents of soils usually correlate with soil organic carbon contents and are generally stimulated by fixed‐carbon amendments (e.g. Knapp et al. 1983; Schniirer et al. 1985). The capacity for soil microbes to utilize efficiently and effectively the pools of metabolizable carbon contained in soil and to be easily stimulated by soil amendment with such materials has been adequately and frequently documented. Similarly, it is commonly noted that nitrogen is rarely limiting to soil microbes, but the active incorporation of nitrogen into soil microbial biomass may cause nitrogen to become limiting to the plants growing in the ecosystem. The low quantities of sulfur and phosphorus required by soil microbes may be supplied through mineralization of plant biomass or through solubolization of soil minerals. Again, although these macronutrients are commonly present in growth‐controlling concentrations for higher plants, soil generally contains adequate sulfur and phosphorus pools for microbial development.
Growth factors: Growth factors are organic compounds required by the cell in small quantities – for example, vitamins and amino acids, purines, and pyrimidines. They are incorporated into the cell intact. It must be noted that if sufficient concentrations of these materials exist in soil, they may also be used as carbon, nitrogen, and/or energy sources. For example, a large proportion of the bacteria growing in the rhizosphere are capable of mineralizing amino acids to meet these nutrient and energy requirements.
Species requiring growth factors are more common in soil than are those capable of synthesizing them de novo. This implies that common sources of growth factors must be present in soil. Four reservoirs of microbial growth factors can be readily identified.
· Other organisms growing in soil: Each microbe in soil is capable of synthesizing a portion of the growth factors that are necessary for its biomass production (prototrophs). A microorganism may produce more than is required for internal consumption. Excess growth factors thus may leak from the cell into the suspending milieu. This cross‐feeding of nutrients (utilization of growth factors leaking from a producer cell to cohabitants of the habitat) is frequently observed when viable cells are grown in petri plates containing vitamin‐ or amino acid‐free media that support development of heterotrophic bacterial colonies. Small satellite colonies develop around the prototrophic bacterial colonies. Alternatively, supplies of the growth factor become available to auxotrophs (those requiring the growth factor) upon death and lysis of the neighboring cells.
· Root exudates and sloughed cells: Plants commonly leak a mixture of growth factors (including vitamins and amino acids) from their roots. Furthermore, as the root extends through soil, packets of nutrients as sloughed cells are released from the root surface and consumed by the soil microbial community.
· Anthropogenically supplied organic substances: Organic matter incorporated into the soil (waste disposal, sludges, composts, and accidental spills) also contains a variety of vitamins as well as macronutrients. These substances are found within the residual cells of the composted vegetative materials as well as in the associated microbial community. Lysis of the plant cells and death and lysis of the microbial cells liberates the nutrients for consumption by the soil microbes.
· Whole plant and animal cells: Predators and parasites receive their growth factors directly from the host cells.
The dominant organisms present in an ecosystem are usually those requiring a few specific growth factors, at most. More fastidious metabolic needs are usually associated with the microbial populations associated with living plants or animals (e.g. rumen or intestinal organisms, and root symbionts). In these latter situations, an organism capable of scavenging growth factors from its environment clearly has an advantage over one that must expend energy to synthesize the factor, but once the available supply of the vitamin or amino acid becomes limiting, the roles are reversed. In a soil system barren of growth factor resources, prototrophs are better competitors than are auxototrophs. An exception to this generalization is associated with those microbes that are capable of forming resting structures, such as endospores. For these microbes, once the easily recoverable supply of growth factors is depleted and their competitive advantage is lost, they form a resting structure wherein they persist until growth conditions improve.
5.2.1.6 Trace Metal Supplies in Soil
A variety of metallic ions are also required for microbial growth. Examples of universally required cations are potassium, iron, and magnesium, whereas most living cells need calcium, copper, and zinc. Cobalt is likely required by most organisms for synthesis of vitamin B12. Molybdenum is required for nitrogen fixation and nitrate metabolism. As would be anticipated by the predominance and variety of mineral components of soils, except for molybdenum, these metals are generally not limiting to soil microbes.
5.2.2 Soil Water
Water is an essential contributor to the growth of soil microbes. Water not only provides an essential medium for growth of microbial populations, but it is also a primary participant in a variety of cell processes. The major roles for water in soil can be summarized as follows.
· Water molecules are reactants in essential metabolic processes: Water is a reactant in hydrolysis reactions (for example, cleavage of cellulose into glucose and proteins into amino acids) as well as hydroxylations, such as the hydroxylation of picolinic acid and other pyridine ring‐containing compounds (e.g. see Dagley and Johnson 1963; Hirschberg and Ensign 1972; Hochstein and Dalton 1967; Tate and Ensign 1974).
· Affects gas exchange in soil: Soil pores are filled with air and water. If all pores are water‐filled, the soil is waterlogged or saturated. Since gaseous diffusion is greater in air than in water, the greater the water content of a soil, the higher the probability of its becoming oxygen limited or even anoxic.
· Affects microbial nutrient supply: Water is the transport medium of microbial nutrients to the cell and of waste materials away from the cell. For nonwater‐soluble nutrients, water serves as the transport medium for the extracellular enzymes or solubolizing agent to reach the substrate and convert it into water‐soluble products or forms. Extracellular enzymes are generally involved in the cleavage of complex polymers into water‐soluble subunits, whereas surfactants increase the availability of hydrophobic materials (such as lipoidal substances) to microbial cells.
· Affects soil temperature: The higher the soil water content, the more resistant is that soil ecosystem to temperature fluctuations. For example, soil temperatures rise more slowly during daily cycles in moist or water‐saturated soils during the spring than occurs in the desiccated or air‐dried soils of midsummer.
· Serves as a growth medium for microbial colonies: Microbes function in the water layer on soil particles or within soil pores. Even in a seemingly dry soil sample, a layer of water must exist surrounding microbial microcolonies and soil particles to support a viable soil community. In reality, although soil is predominantly a particulate system, microbial life forms contained therein live in an aquatic world. This water‐saturated system may consist of a microfilm a few microns thick or it may extend throughout the soil micro‐ or macropore system. In either case, for the microorganism to function properly, it must be totally bathed in water.
5.2.2.1 Measurement of Soil Moisture
Standard techniques for quantification and expression of soil moisture levels are a necessity to the development of predictive mathematical models of the impact of soil moisture variation on soil processes and to the provision of a basis for comparison of moisture effects on biological activity in contrasting soil types. The procedure should be easily conducted and reproducible, as well as measure an aspect of soil moisture that directly affects soil microbial activity.
Traditionally, soil moisture has been reported as gravimetric water content [ω, g H20 g−1 dry soil (usually dried to a constant weight, such as at 105 °C)], and volumetric water content (θ, cm3 H2O cm−3 soil). These two means of expressing soil moisture are related to the soil bulk density (pb, g solids cm−3) by the following relationship:
(5.1)
A third term encountered on a perhaps all too frequent basis is g H20 g−1 wet soil. Use of the latter term is particularly distressing. It provides some means for comparison of soils data when a single soil type is considered, but no commonalty for evaluation of results between soil types exists.
Although gravimetric and volumetric water content are frequently used to measure soil moisture by soil microbiologists, their use is somewhat problematic in that neither value describes the actual availability of water to the soil microbial community. Furthermore, these values do not vary independently of soil type. Thus, comparable gravimetric or volumetric moisture values in two different soil types do not represent comparable water conditions impacting the resident microbial communities. Although a relationship between soil moisture and a specific microbial activity can be derived for a single soil type using these values, it is not possible to determine the generalized principles behind the relationship.
A partial solution to this difficulty is to use water potential as a measure of water availability in soil pores. Water potentials – measurements with a long history of use by soil scientists in general – are governed by surface tension forces and the radius of the menisci of the water. Thus, values can be assessed by principles of capillary rise. The mathematical expression of this relationship is as follows:
(5.2)
where r is the equivalent or effective capillary radius, τ is the suction necessary to absorb the water, and γ is the surface tension of water. Note that with this relationship, as the equivalent capillary radius becomes smaller, less water is present and the suction necessary to absorb the water increases. Equivalent expressions of suction are as follows:
Common water potential values found in the soil microbiology literature are the moisture content of soil at −0.3 bar (field capacity). The basic advantage of this means of assessing and expressing soil water content is that it is based on the energy required to removed the water from the soil matrix. Water adjacent to soil particles requires more energy to remove it from the soil than does the water more distant from the soil particles (see Brady and Weil (2017) for a more complete description of water distribution in soil).
Another useful expression of soil moisture is water‐filled pore space (Linn and Doran 1984). Soil microbes exist in a mixed soil air‐water system. In reality, it is the degree of displacement of air with water within the soil pore that combines with the rate of microbial respiration to produce anoxic or anaerobic microsites within the soil profile (see Skopp et al. (1991) for a detailed study of this process). Complete displacement of air with water restricts diffusion and thereby hastens imposition of oxygen‐limited conditions. When moisture approaches or exceeds field capacity, the percent soil pore space filled with water or air becomes a better indicator of aerobic vs anaerobic microbial activity than does water content or water potential. Use of percent water‐filled pore space to express soil moisture levels is practical since it only requires knowledge of the soil bulk density and the gravimetric soil water content. (The particle density of mineral soils is assumed to be 2.65 Mg m−3.) The percent water‐filled porosity is defined by the following relationship:
(5.3)
where θv is equivalent to the product of the gravimetric water content (ω) and the soil bulk density (ρb) and TP is the percent total porosity (1 – ρb/ρp pp) (100), where ρb is the soil bulk density and ρp is the soil particle density. For aerobic processes, maximal activity is detected at a percent water‐filled porosity of about 60%. Below 60%, water limits activity (insufficient moisture exists to provide the contiguous surface film required by the soil microbes), whereas above 60% aerobic microbial activity decreases (apparently due to reduced oxygen availability). Caution is necessary in use of these generalized figures. Neilson and Pepper (1990) found discrepancies in use of percent saturation or water‐filled pore space to characterize soil aeration, especially when soils of differing bulk densities or those receiving particulate organic matter amendments are compared.
The effect of water in a heterogeneous system can also be expressed as water activity. This measurement of moisture is most commonly used in food storage to describe water availability. Water activity is related to the equilibrium relative humidity (ERH) as follows:
(5.4)
The difficulties with using water activity in soil microbiological research are the complications associated with its measurement. Long incubation times in a chamber where the moisture level of the test sample is allowed to equilibrate with the chamber atmosphere are usually required. Data can be extremely variable. Water availability affects species diversity, survival, movement, and activity of microbes. Of all of the expressions that may be used to define the moisture conditions under which the soil microbe is abiding, water activity is probably the most useful. Unfortunately, it is also the most difficult and time‐consuming (i.e. impractical) measure of soil moisture.
Impact of moisture variation on soil microbial activity: Total microbial activity varies from nearly nonexistent levels at low water availability to a maximal level under optimal soil air/moisture mixtures (Figure 5.3). Typically, soil bacteria are capable of functioning under seemingly severe soil moisture tensions. For example, Wilson and Griffin (1975) found that respiration of a mixed bacterial population in soil was maintained at a reasonably high level between −8 and −30 bar, but was negligible at −50 bar. Similar results have been reported by Wildung et al. (1975) and Knight and Skujins (1981). As soil moisture levels are saturated and free oxygen supplies are depleted, overall activity again declines. A generalized curve (Figure 5.3) is applicable to the summation of microbial respiratory activities and to a certain degree individual processes. In the latter situation, the points of minimal or optimal activity are characteristics of the microbial process being studied. Obligatorily anaerobic processes clearly are optimized under saturation conditions.
Tolerance to restrictive soil moisture levels varies with major microbial groups as well as with individual microbial species. Generally, bacteria are more exacting than are actinomycetes and fungi. Thus, the latter organisms tend to predominate in drier soils. This observation results at least in part from the capacity of the latter groups of organisms to form resting structures. Robertson and Firestone (1992) postulated that bacteria may use extracellular polysaccharide to enhance survival under moisture‐limiting conditions. A purified extracellular polysaccharide from a Pseudomonas species was shown to contain severalfold its weight in water at low moisture tension. Within the realm of the bacteria, a gradient of tolerance is noted to occur. For example, ammonium tends to accumulate in dry soils due to the ammonifying microorganisms being less sensitive to the low water activities than are nitrifiers.
Figure 5.3 Variation in aerobic biological activity in soil. In this hypothetical example, optimal moisture is approximately 0.3 mPa.
Selection of microbial populations with increased resistance to moisture limitations in chronically arid soils was demonstrated by Knight and Skujins (1981). These workers used adenosine triphosphate and soil respiration measurements as indicators of microbial biomass and activity. The impact of variation of soil moisture between −2 and −100 bar tension on microbial activity in two arid soils and a subalpine forest soils was evaluated. The microbial activity increased in arid soils between −2 and −20 bar soil moisture, whereas the optimal moisture in the chronically wetter forest soils was −2 bar and activity continually decreased below that level. Thus, it was concluded that a characteristic microbial populations capable of managing the prevailing environmental conditions had developed in each of the extreme environmentally different soils studied.
It is important for the student of microbiology to understand clearly what these observations reveal about the function of the soil microbial community and at least as importantly, what is not stated. The positive side of this observation is that it follows that as long as the soils are not so dry as to preclude biological function, an appropriate microbial population does develop. The negative aspect is that it is not implied that this microbial population selected for growth under restricted soil moisture is as active as a comparable population growing under less stringent conditions. That is, were microbial respiration to be measured under limited soil moisture and at optimal soil moisture, even if the organisms in each situation were absolutely optimized to their environment, less respiration would be found in the low‐moisture soil. Practically, this means that one should not anticipate as much organic matter to be decomposed in desert soils without irrigation as could occur in an irrigated field of the same soil.
The impact of excessive soil moisture (flooded or chronically waterlogged conditions) on soil microbial activities is more predictable than situations observed in habitats experiencing a shortage of free water. Interactions of soil moisture levels with availability of molecular oxygen become evident in the flooded or chronically waterlogged soil sites. A traditional division of the microbial realm has been separation by oxygen tolerance (anaerobes, facultative anaerobes, and aerobes). Flooded soils are not necessarily oxygen free, but should available free oxygen supplies be depleted, anaerobic and facultative microbial populations are stimulated while obligate aerobes die or enter resting stages. The most dramatic effect of institution of anaerobic conditions in soil is the induction of facultative (e.g. fermentations), anoxic aerobic metabolism (e.g. denitrification), or strictly anaerobic processes (e.g. methanogenesis). Organic acids may accumulate, nitrogen oxides could be evolved, and ultimately, methane generation or sulfate reduction may occur. Practically, anoxic soils may be recognized by precipitation of metal sulfides, or evolution of an amine or sulfide odor. Characteristics of the microbial community are dependent upon the duration of the flood period and the nature of electron acceptors present. Methanogenic bacteria require development of highly reducing conditions, whereas fermentative organisms are less stringent (see Chapter 4).
As stated above, a flooded soil is not necessarily either anoxic or highly reducing. Water movement through the environment as well as diffusion of atmospheric gases into the system may prevent exhaustion of free oxygen. This phenomenon was shown in a study of the muck soils of south Florida (Tate 1979). With flooding of the soils, an increase in aerobic bacterial populations was initially observed (Figure 5.4). The soils were flooded to maximum depths of approximately 31 cm. As the free oxygen and easily metabolized carbon source levels declined, a parallel reduction of aerobic bacterial populations occurred. Thus, were the experimenter assuming that flooded and anoxic conditions were equivalent, during the initial portion of the study, a behavior of the soil microbes contrary to what occurred in situ would have been expected.
Figure 5.4 Effect of flooding a dry histosol on aerobic bacterial populations. Soils were flooded to a maximum depth of approximately 31 cm commencing on day 0 (arrow A). Arrow B indicates the time of drainage of the flood waters.
Source: Adapted from Tate (1979).
5.2.2.2 Soil Systems with Fluctuating Soil Moisture
Further underlying assumptions or perhaps attitudes that must be developed in relationship to understanding the effect of soil moisture on the microbial community are (i) the moisture level of a particular site tends to be continually changing and (ii) moisture effects on biological activity are interactive with other system properties.
Moisture fluctuation may be on a short‐term, localized basis as would be found (i) with daily impact of temperature variation and water evaporation, (ii) following a heavy rainfall, or (iii) due to seasonal flooding, as occurs in a wetland rice soil. For example, microbial biomass fluctuations can reveal the short‐term effect of soil moisture levels induced by precipitation (Clarholm and Rosswall 1980). Precipitation caused increases in microbial biomass lasting one to two days, even in nonmoisture‐limited soil. This observation suggests that although data descriptive of macrosite soil moisture tensions support a conclusion that the microbial populations are not limited by this soil property, it is microsite discontinuity in moisture films covering soil particles that determines the rate of change in and ultimate level of microbial population densities. In flooded rice field soils, anoxic conditions induce anaerobic microbial populations during the somewhat long term of flooding. Draining of the site following harvest or for establishment of seedlings induces aerobic and facultative microbial populations. Strict anaerobes may be further limited by the flow of oxygenated water through the system. In that situation, a mosaic develops in soil, with pockets of anoxic soil interspersed with oxygenated soil.
Note that soil systems are described as containing a mixture of aerobic and anaerobic micro‐ or macrosites, not “a partially aerobic system.” By definition, each individual soil microsite is either anaerobic (anoxic) or aerobic. “Partially aerobic” is an inappropriate description in that “aerobic” indicates the presence of oxygen in the system at the location of the soil microbe, which is an all‐or‐none state. Even if only a few molecules of oxygen are present per cubic centimeter of soil, the site is still described as being aerobic.
The interactive nature of soil moisture with other soil properties is documented by examining wet/dry cycling on soil organic matter availability as well as salinity impacts on microbial activity (see Rousk et al. 2011; Setia et al. 2011). Variation of soil moisture with time not only affects the nature of the microbial population present in the soil, but it may also alter the quantity of nutrients or the physical structure of the microbes' environment. Cycling of soil moisture liberates occluded organic matter through such processes as disruption of soil aggregate structure (for example, see Asghar et al. 2012) or by death of the more stringent microbial populations. Therefore, microbial oxidation of soil organic matter is greater in soils with fluctuating moisture than in constantly wet or dry conditions (e.g. see Lund and Goksoyr 1980; Orchard and Cook 1983; Reddy and Patrick 1975; Sorensen 1974; Thiemann and Billings 2012). Jager and Bruins (1975) noted that the loss of carbon due to microbial respiration in a soil experiencing approximately 60 wet/dry cycles was also proportional to drying temperature, with greatest loss (31.2%) in soil dried at 85 °C and least (18%) in soil dried at 35 °C. Drying resulted in an ultimate reduction in microbial population densities, but the highest population densities were found in soil dried at 85 °C, suggesting that more organic matter was made available to support microbial growth by the more drastic treatment. A practical implication of this observation relates to maintenance of soil organic matter levels in mineral soils and the minimization of subsidence of histosols (see Tate 1980). In both situations, maintenance of the native organic matter levels is desirable for preservation of the ecosystem and/or encouragement of optimal soil structure (see Tate 1987).
An indirect effect of declining soil moisture is the impact of osmotic pressure on biological activity. Generally, the osmotic potential of water is less important than water availability, but in some localized or transient situations, variations in osmotic potential emerge as significant controllers of microbial growth. Potentially lethal or inhibitory salt concentrations may be the product of microbial metabolism (e.g. mineral dissolution through acidification of microsites) and chemical equilibria (e.g. ionization of water‐soluble salts), as well as physical phenomena (evaporation). Localized problems are associated with areas of salt encrustation due to water evaporation; saline waters such as are found in some irrigation systems, around saltwater bodies; or in association with saline soils. Transient difficulties may be associated with severe desiccation of soil.
Major impacts of osmotic pressure are likely averted in soil because of the fact that reduction of soil moisture results in induction of cellular survival adaptations (for example, sporulation). Also, in regions with chronically saline soils, selection of organisms with increased resistance to pressure changes may occur. For a discussion of microbial adaptations to osmotic changes, see Csonka and Hanson (1991).
5.2.3 Aeration
A major role of molecular oxygen in soil is as a terminal electron acceptor. As was indicated in Chapter 4, energy is produced during electron transport to oxygen. In fact, oxygen is the highest energy‐producing electron acceptor. Thus, the greatest cell yields are produced by the total oxidation of an organic carbon‐containing substance to carbon dioxide and water. This important function of molecular oxygen in aerobic metabolism is augmented by its direct incorporation into molecular structures of such compounds as aromatic ring‐containing substances and alkanes by oxygenases. The significance of aerobic metabolism to the soil microbial community is underscored by the observation that most microbes in surface soils are aerobes. Exceptions are soils that are consistently flooded, such as bog soils.
Soil oxygen tension is controlled by gaseous diffusion rates as well as by the respiratory activity of the soil biota. Since diffusion is a relatively slow process, it is not unusual for the rate of consumption of this essential gas to be greater than its rate of replacement. Thus, free oxygen tensions are rarely too high for soil aerobes, but clearly soil oxygen levels may be too low for obligate aerobes.
Indirect limitors of soil oxygen are soil moisture and the concentrations of metabolizable organic matter. As was indicated above, moisture affects soil oxygenation in that the greater the soil moisture, the fewer air‐filled pores exist, and the slower the diffusion rate. Decomposable organic carbon availability controls the rate of oxygen consumption indirectly in that as this fixed carbon is oxidized, free oxygen may be depleted. With an influx of easily metabolized organic matter, it is not unusual for anoxic conditions to develop. Examples of practical consequences of oxygen depletion due to soil microbial respiration are as follows.
· Oxygen depletion in cropped soils after a heavy rainfall event can result in crop damage.
· After a heavy rainfall, with the presence of nitrate and suitable carbon sources, significant losses of fixed nitrogen from soil can result from induction of denitrifiers.
· Spills of easily decomposable organic matter in soil or incorporation of readily metabolizable organic matter can result in depletion of free oxygen supplies thereby resulting in ecosystem disruption through oxygen starvation of plant roots, death of small animals, earthworms, nematodes, mites, and so forth.
As the free oxygen in soil is depleted, a number of predictable changes in the microbial activity occur. When the soil oxygen tension has been reduced to less than 1% (v/v), the microbial population appears to shift from being predominantly aerobic to anaerobic. With the development of a reducing atmosphere, growth yields decline since the energy yielded per mole of fixed carbon oxidized anaerobically is far less than that produced from aerobic respiration (see Chapter 4). (Note that dissimilatory respiration of aerobic bacteria [denitrification and sulfate reduction] cell yields approximate those with oxygen as the terminal electron acceptor since a cytochrome based electron transport processes occurs and the carbon substrates are oxidized completely to carbon dioxide, water, and, ammonia.) Fermentation results in accumulation of incompletely oxidized carbonaceous substrates (organic acids, amines, etc.). When highly reducing conditions develop so that methanogenesis and sulfate reduction occur, complete mineralization of fixed carbon substrates again occurs.
5.2.4 Redox Potential
It is difficult to separate the impact of redox potential from oxygen tension effects experimentally. To artificially raise the redox potential in the absence of free oxygen, it is necessary to add oxidizing agents, which are generally toxic to microorganisms. Thus, it must be noted that changes in soil redox potential are related to changes in oxygen levels. If organic matter is added to soil, oxygen is depleted and the potential drops – at times quite precipitously. This is a microbial reaction since inhibitors of microbial activity prevent both oxygen depletion and development of reducing conditions – which is somewhat logical since in an actively respiring community, oxygen is used as an electron acceptor.
As has been alluded to above, the occurrence of a variety of microbial processes is related to specific redox potentials. Some of these are as follows.
· Aerobic carbon oxidation >0.2 V
· Denitrification 0.15 to 0.2 V
· Methanogenesis −0.2 to −0.1 V
· Sulfur reduction −0.2 to −0.1 V
Although these data suggest a somewhat mutually exclusive nature of microbial activity in soil, it is not unusual to observe these processes occurring concurrently across a landscape of limited size. Again, this results from the heterogeneity of the soil system. Two examples of this situation (variation of redox potential in a flooded soil and within the profile of a drained histosol) are provided in Figure 5.5. In both situations, an oxygen‐rich, high redox potential region overlays an anaerobic portion of the soil profile. A region of variable conditions generally occurs between these two extreme environments. Similarly, a mosaic of activities can occur in a well‐aggregated soil where aerobic processes predominate on the surfaces of the aggregates and anoxic processes occur in the interior.
Figure 5.5 Redox potential and oxygen tension patterns in an hypothetical flooded soil (a) and a soil with an elevated water table (b).
5.2.5 pH
The microbiologist's bias tends to favor conducting experiments with soils with approximately neutral pH and the study of the microbes capable of functioning therein. Our laboratory media developed for the culture of microorganisms are so constituted, and the majority of the soil systems studied tend to fall in a pH range of approximately 5.5–7.5. Yet acidic soils constitute a major portion of the world's soil resources, especially as exemplified by soils of tropical regions and temperate forests, and acidification is a major reclamation problem in the normally near‐neutral soils of the temperate region. For example, with many mining processes, metal‐sulfide‐bearing slags are produced. Biological activity in these waste materials results in oxidation of the sulfide to sulfate. Subsequent drainage of these acid‐bearing waters presents major problems for downstream soils and receiving waters. Thus, it is imperative to understand the effect of soil pH on soil microbial community development and function.
Acidic conditions present a particularly stressful situation to the microbial cell. Organisms existing in acidic habitats could be described as being in a somewhat schizophrenic state. The internal cell pH must be highly buffered at or near neutrality, the pH optimum for internal enzymes. Thus, the effect of variation of the external pH necessitates development of cell surface and membrane‐associated mechanisms to control the internal pH. Furthermore, metabolic processes occurring at the cell surface must be adapted to cope with the acidic conditions.
A variety of pH‐associated metabolic problems must be surmounted by the cell. For example, cellular surface charge varies with environmental pH. This controls the interaction with soil particles as well as with charged nutrients. Furthermore, surface‐bound enzymes as well as extracellular enzymes (see Chapter 6) must also be capable of functioning at the elevated hydrogen ion concentration. Maximal activity must either be expressed at the predominant pH, or enzyme synthesis rate must be modified to compensate for the reduced enzymatic activity occurring under the acidic conditions. Also, adjustment of enzyme synthesis rates may be necessary to compensate for loss of those enzymes that are denatured under acidic conditions. If the enzyme product is essential for cell function, then the organism must adjust the enzyme synthesis rate so that it is always present at effective levels.
5.2.5.1 Indirect Mechanisms for pH Limitation of Biological Activity in Soil
Nutrient concentrations and toxicity of environmental substituents are both affected indirectly by soil acidity. Trace mineral availability provides an excellent example of pH interactions with nutrient availability. Both iron and manganese are more water soluble at low pH, whereas molybdenum is precipitated at low pH.
Similarly, capacity to transport charged carbonaceous substances such as fatty acids and amino acids into the cell is dependent upon the charge of the molecules. Reduction of the soil pH below the pKa of acids results in conversion of the negatively charged entity to a neutral compound. Zwitter ions may have an overall negative charge or positive charge, or be uncharged depending upon the pH of the suspending medium. For example, glycine (+H3NCH2COO−) has both positive and negative charges at neutral pH. As the pH is varied, the positive and negative charges are neutralized independently. Each of these states would result in alteration of the capacity of the cell or its catabolic enzymes to catalyze transformation of these materials.
Under acidic conditions, organic acids can be toxic to microbial growth. This is not a pH effect, but is an impact of the presence of the organic acid itself in an uncharged or neutral state. Thus, accumulation of organic acids under anoxic conditions in acidic environments could be a mechanism for limiting microbial activity. This interaction of pH and anaerobic conditions may contribute to organic matter accumulation in acid swamps.
A final indirect effect of acidic soil pH conditions on biological activity relates to the potential for solubolization of toxic compounds or elements. Aluminum is more available under acidic conditions. It is not uncommon for toxic levels of this cation to be detected in acidic tropical soils.
Direct modifications of biological activity by pH variation: The direct effect of pH relates to the fact that each microbial strain has an intrinsic range of pH within which it is capable of functioning. The diversity of soil life is exemplified by the observation that microorganisms grow from about pH 1–11. Similarly, since it is to a large degree the property of an organism's enzymes that determine the diversity of processes catalyzed, a comparable reaction to environmental pH is found for these cell components. Although microbial life occurs at essentially all naturally occurring soil pH levels, each individual species (and to some extent, strain) has a characteristic pH range and optimum pH for growth. For example, Thiobacillus thiooxidans, a key organism in oxidation of reduced sulfur‐containing compounds, grows only at extremes of acidity (pH 2–4), whereas Nitrosomonas sp., a key player in biological nitrification, has an optimum pH in the alkaline range and is not active below about pH 5.5. Similarly, another sulfur‐oxidizing bacterium, Thiobacillus thioparus, grows only at near‐neutral pH.
Some selection of overall contributors to biological activity results from variation of soil pH. In acid soils, the dominant activity is generally the result of the fungal community. This results from the poor growth of most heterotrophic bacteria at acid pH. An exception is with waterlogged acid soils. Here, the aerobic fungi are inhibited by oxygen limitations, so the acid‐tolerant bacteria are better able to compete.
Some microbially catalyzed processes occur in soils with a specific pH range. For example, bacterial catalyzed nitrification (oxidation of ammonium through nitrite to nitrate) is suggested to occur primarily in soils with a pH greater than 5.5. This observation results from the fact that the bacteria strains involved in nitrification belong to a small group of autotrophic bacterial species that are pH sensitive (Dommergues et al. 1978).
It can be generalized that if a process occurs over the entire pH range conducive to development of living cells, the transformations result from the activity of a variety of microbial species or strains with varying pH optima. For example, carbon and nitrogen mineralization occur in essentially any soil ecosystem where life is possible. Thus, a measure of the diversity of the microbial community is revealed by the breadth of conditions under which the organisms are able to function. This maxim relates not only to soil pH but also to other properties, such as moisture and aeration.
5.2.5.2 Conflicts Between Laboratory Observations and Actual Field Activities
Some anomalous situations have been encountered. These are associated with conditions where laboratory experiments with axenic microbial cultures yield data suggesting that the only known microbes capable of catalyzing a particular process cannot function at the prevailing soil pH of the site under study, yet the products of the reaction are detected. For example, as indicated above, acidophilic autotrophic nitrifiers have not been detected in acidic forest soils, yet nitrate is commonly found in these soils. Several explanations have been proposed to explain this seemingly anomalous observation (for example, see Dommergues et al. 1978). Alternative explanations include the following: the microbes may be growing in soil microsites with a more favorable pH. That is, the acid‐limited microbes may be constrained to islands of more moderate conditions contained within a sea of restrictive soil. This situation would be more likely to occur in soils where the overall pH is near the extremity of function of the microorganism. That is, an overall soil pH of approximately 5.0 could be measured, yet microsites 0.5 pH units above or below that value may exist.
An alternative explanation to the anomalous occurrence of a pH‐limited process may involve our capability of culturing the responsible microbes or even discovering the existence of microbes with appropriate capabilities. This limitation is exemplified by the increasing number of anaerobic ammonium‐oxidizing (anammox) bacterial strains being isolated that are capable of catalyzing processes previously considered to be pH and oxygen limited. (See the following publications for further discussion of this topic: Gijs Kuenen 2008; Kartal et al. 2008; Schmidt et al. 2002, 2005.) Only a small portion of the microbial community in any soil can be cultured and studied in the laboratory. Techniques for the study of the vast majority of this diverse community remain to be developed. Thus, occurrence of processes in the field may be a better indicator of microbial capabilities than is the library of information collected from axenic cultures.
Lastly, a spontaneous chemical conversion of the reactants may occur outside the pH range optimal for biological activity. This is exemplified by denitrification under acidic conditions. In this situation, bacteria can be cultured that catalyze reduction of nitrate to nitrous oxide and dinitrogen under acidic conditions, but their function in the native acidic soil is questioned. The kinetics of the spontaneous chemical reaction favor that process over the biologically catalyzed nitrogen oxide reduction.
In conclusion, a variety of interactions of soil hydrogen ion concentration and soil biological process have been elucidated. Life does occur in extremely acidic or alkaline soils. The participants may vary from those under less stringent conditions, and the range of processes occurring therein may be somewhat truncated. Stewardship of acid soils and reclamation of acid‐affected soils requires consideration of the reactions occurring therein and the limitations of the system. Due to the intrinsic buffer capacity of soil, management of an acidophilic community may be necessitated by the inability to adjust the pH of many soils to a more neutral status (e.g. Magdoff and Bartlett 1985).
5.2.6 Temperature
Greatest resistance to temperature extremes among living entities is found in the microbial community. The typical growth range for soil inhabitants is more restrictive than the extremes associated with other ecosystems. The typical breadth of temperatures supportive of microbial growth in soil is from approximately 0 °C to 70 °C. This contrasts on the upper side of the range to growth of bacteria in hot springs, which are capable of growth at 100 °C (Heinen and Lauwers 1981). Minimal growth temperatures have been shown to be slightly below 0 °C in Antarctic soils. With either maximum or minimal growth temperatures, the limiting factor appears to be free water. As long as the conditions allow existence of free water (either through boiling temperature elevation by augmented pressure or freezing point reduction by salt concentration), microorganisms appear to be capable of growth, albeit slow growth at the reduced temperatures.
Reduction of soil temperatures below freezing results in significant declines in microbial populations (Biederbeck and Campbell 1971). The cause of cell death is not the freezing of the cell, in that freezing itself causes only a small reduction in microbial numbers. The lethal factor appears to be associated with slow thawing of the soil. As soils thaw, ice crystals form in the microbial cells and disrupt cellular integrity. Sensitivity also relates to the growth status of the microbial cells in that recently cropped soils exhibited greater population declines due to freezing than was detected in inactive soils (Biederbeck and Campbell 1971). Furthermore a differential sensitivity of the microbial populations was noted in that bacterial population reduction was greater (92%) than that of fungi (55%), which was greater than that of actinomycetes (33%). Similar differential sensitivity of soil bacteria to freezing and slow thawing was noted by Nelson and Parkinson (1978) in a study of a Pseudomonas sp., Bacillus sp., and Arthrobacter sp. Survival was dependent on soil moisture, storage time, and the thaw rate.
More typically for soil microbes, growth can be depicted as an activity range (Figure 5.6). The growth rate increases until a maximal temperature, characteristic of the organism, is reached. The maximum temperature that supports growth is determined by the sensitivity of the most fastidious enzyme essential for cell replication. This temperature sensitivity curve is a good example of the law of tolerance which states that a microorganism will have an ecological minimum and maximum with a range in between that represents the limits of tolerance.
Tolerance to temperature is commonly used to divide microorganisms into three classes. Psychrophiles are defined to grow optimally at temperatures less than 20 °C. Mesophiles grow best between 15 and 45 °C, whereas thermophiles are active at temperatures greater than 45 °C. Few if any psychrophiles are found in most soils. Exceptions are associated with Arctic and Antarctic sites. Mesophiles constitute the bulk of the soil microbial community, whereas most soils contain some thermophiles, even if the soils never achieve the high temperatures.
Figure 5.6 Typical response of a mesophilic bacterial population to temperature variation. An optimum temperature of 25–30 °C is depicted.
The presence of these classes of microbes assures occurrence of metabolic activity over a wide temperature range. Typically, soil biological activity increases from a minimum at or near 0 °C to a maximum and terminus point near 70 °C. The rate increase can be represented by several mathematical relationships, among which are the Arhenius and Q10 relationships. Historically, the latter is likely the most commonly used parameter for soil microbiological studies. The Q10 is equal to the reaction rate at temperature one divided by the rate at temperature one minus 10 °C, that is:
(5.5)
Q10 values for biological processes generally range from 1.5 to 3.0, with a mode of approximately 2.0. Generally, if the Q10 is much greater than 3.0, an environmental parameter besides temperature is controlling the reaction rate. For example, Q10 values for petroleum decomposition at low temperatures (10–20 °C) are commonly greater than 5. At the low temperatures, catabolism of the oil is controlled by its water solubility and viscosity. These parameters dramatically change as the temperature is raised, so a disproportional impact of temperature elevation of the microbial oxidation is observed.
5.3 Microbial Adaptation to Abiotic Stress
From an analysis of the diversity of soil properties affecting microbial activity, it almost appears that life in soil would be limited at best. Yet, nearly every site on Earth supports active microbial populations. Even in situations where a priori considerations would predict that two factors, such as moisture and temperature, exist at limits that preclude microbial growth, microorganisms have been shown to occur in extremely specialized niches. For example, in cold dry deserts of Antarctica, cyanobacteria‐dominated communities have adapted to growth between the crystals of porous rocks (cryptoendolithic) where some modulation of the extreme environmental stress occurs (Friedmann 1982; Johnston and Vestal 1991). This portends a variety of mechanisms of community adaptation to these stresses. Three reactions to stressful conditions should be highlighted.
First, if the stress is extreme, there is no adaptation. This is exemplified by soils with temperatures greater than 70 °C around hot springs or adjacent to nuclear reactor water outflows, some extremely acidic mineland reclamation sites, and soils containing excessively high metal concentrations. The existence of such ecosystems underscores the fragile nature of the biotic system and highlights the fact that at least some growth is necessary for implementation of the two following survival mechanisms.
Second, if growth is possible, microbial strains resistant or at least tolerant to the stress factor are selected. This has been observed with osmotic pressure, temperature, oxygen tension, and moisture stress. This adaptation mechanism may result in reduction of the species diversity, especially if the stress is severe, but a functional community develops. Reduction of the microbial diversity, and therefore the genetic diversity, of the community could portend enhanced vulnerability to ecological insult. Soil presents a somewhat unique exception to this principle in that the potential always exists for invasion by propagules from adjacent soils. Thus, the genetic potential required to overcome chemical or physical stress in a particular site could exist in less stressed neighboring soils. In this case, passive or active transfer of propagules from the neighboring soils may result in establishment of a new community in a site that previously lacked the resources necessary for survival.
Lastly, in some situations, the microbial community may possess the capacity to alter the stress factor, thereby relieving the impediment to growth or survival. For example, microorganisms may modulate inhibitory acidic or alkaline conditions or decompose toxic substances. Organic acids or even antibiotics may be decomposed to carbon dioxide by microbial strains resistant to their toxicity. Even nonbiodecomposible toxins such as heavy metals may be transformed to nontoxic states. They may be complexed, precipitated, oxidized or reduced, or even chelated. The products are rendered nonwater soluble, converted to water‐soluble but nontoxic forms, or incorporated into a chelate complex that can be leached from the microbial habitat. Inhibitory inorganic acids may also be rendered ineffective. For example, sulfate is reduced to hydrogen sulfide, which is precipitated by metal ions, such as iron, which are common in soil. In each of these examples, homeostasis is reached by activity of resistant populations conferring benefits to the survivors of the susceptible organisms.
5.4 Concluding Comments
The nature, activity, and future of any soil microbial community are determined by the capacity of its individual members to adapt to or modify negative soil properties. Those members of the community best equipped to cope with the combination of ecological stresses are active, while more stringent organisms succumb or enter resting stages. Modification of the physical, biological, or chemical properties of the site results in induction of microbial processes to restore the homeostasis of the community. This adaptation may result in reemergence of an ecosystem resembling that existing prior to perturbation, development of a new community better suited to the newly developed conditions, or total destruction of the biological activity. The ultimate fate depends upon the genetic diversity of the populations and the severity of the stress imposed, as well as the time allowed for recovery. Some situations may allow recovery in hours to days, whereas in others, full recovery may not be evident for decades or centuries.
References
1. Asghar, H.N., Setia, R., and Marschner, P. (2012). Community composition and activity of microbes from saline soils and non‐saline soils respond similarly to changes in salinity. Soil Biol. Biochem. 47: 175–178.
2. Benoit, R.E. and Hall, C.L. (1970). The microbiology of some dry valley soils of Victoria Land Antarctica. In: Antarctic Ecology, vol. 2 (ed. M.W. Holdgate), 697–701. New York: Academic Press.
3. Biederbeck, V.A. and Campbell, C.A. (1971). Influence of simulated Fall and Spring conditions on the soil system. I. Effect on soil microflora. Soil Sci. Soc. Am. Proc. 35: 474–479.
4. Brady, N. and Weil, R. (2017). Nature and Property of Soils. New Jersey: Prentice Hall.
5. Cameron, R.E. (1972). Microbial and ecological investigations in Victoria Valley, Southern Victoria Land Antarctica. In: Antarctic Terrestrial Biology. Antarctic Research Series, vol. 20 (ed. G.A. Llano), 195–260. Washington, DC: American Geophysical Union.
6. Chowdhury, N., Marschner, P., and Burns, R.G. (2011). Soil microbial activity and community composition: impact of changes in matric and osmotic potential. Soil Biol. Biochem. 43: 1229–1236.
7. Clarholm, M. and Rosswall, T. (1980). Biomass and turnover of bacteria in a forest soil and a peat. Soil Biol. Biochem. 12: 49–57.
8. Csonka, L.N. and Hanson, A.D. (1991). Prokaryotic osmoregulation: genetics and physiology. Annu. Rev. Microbiol. 45: 569–606.
9. Dagley, S. and Johnson, P.A. (1963). Microbial oxidation of kynurenic, xanthurenic, and picolinic acid. Biochim. Biophys. Acta 78: 577–587.
10. Dommergues, Y.R., Reiser, L.W., and Schmidt, E.L. (1978). Limiting factors for microbial growth and activity in soil. In: Advances in Microbial Ecology, vol. 2 (ed. M. Alexander), 49–104. New York: Plenum Press.
11. Friedmann, E.I. (1982). Endolithic microorganisms in the Antarctic cold desert. Science 215: 1045–1053.
12. Gijs Kuenen, J. (2008). Anammox bacteria: from discovery to application. Nat. Rev. Microbiol. 6: 320–326.
13. Heinen, W. and Lauwers, A.M. (1981). Growth of bacteria at 100°C and beyond. Arch. Microbiol. 129: 127–128.
14. Hirschberg, R.L. and Ensign, J.C. (1972). Oxidation of nicotinic acid by a Bacillus species: source of oxygen atoms for the hydroxylation of nicotinic acid to 6‐hydroxynicotinic acid. J. Bacteriol. 108: 757–759.
15. Hochstein, L.I. and Dalton, B.P. (1967). The purification and properties of nicotine oxidase. Biochim. Biophys. Acta 139: 56–64.
16. Jager, G. and Bruins, E.H. (1975). Effect of repeated drying at different temperatures on soil organic matter decomposition and characteristics, and on the soil microflora. Soil Biol. Biochem. 7: 153–159.
17. Johnston, C.G. and Vestal, J.R. (1991). Photosynthetic carbon incorporation and turnover in Antarctic cryptoendolithic microbial communities: are they the slowest‐growing communities on Earth? Appl. Environ. Microbiol. 57: 2308–2311.
18. Kartal, B., van Niftrik, L., Rattray, J. et al. (2008). Candidatus ‘Brocadia fulgida: an autofluorescent anaerobic ammonium oxidizing bacterium. FEMS Microbiol. Ecol. 63: 46–55.
19. Knapp, E.B., Elliott, L.F., and Campbell, G.S. (1983). Microbial respiration and growth during the decomposition of wheat straw. Soil Biol. Biochem. 15: 319–232.
20. Knight, W.G. and Skujins, J. (1981). ATP concentration and soil respiration at reduced water potentials in arid soils. Soil Sci. Soc. Am. J. 45: 657–660.
21. Linn, D.M. and Doran, J.W. (1984). Effect of water‐filled pore space on carbon dioxide and nitrous oxide production in tilled and nontilled soils. Soil Sci. Soc. Am. J. 48: 1267–1272.
22. Lund, V. and Goksoyr, J. (1980). Effects of water fluctuations on microbial mass and activity in soil. Microb. Ecol. 6: 115–123.
23. Lundgren, B. (1982). Bacteria in a pine forest soil as affected by clear‐cutting. Soil Biol. Biochem. 14: 537–542.
24. Magdoff, F.R. and Bartlett, R.J. (1985). Soil pH buffering revisited. Soil Sci. Soc. Am. J. 49: 145–148.
25. Mavi, M.S., Marschner, P., Chittleborough, D.J., and Cox, J.W. (2012). Salinity and sodicity affect soil respiration and dissolved organic matter dynamics differently in soils varying in texture. Soil Biol. Biochem. 45: 8–13.
26. Neilson, J.W. and Pepper, I.L. (1990). Soil respiration as an index of soil aeration. Soil Sci. Soc. Am. J. 54: 428–432.
27. Nelson, L.M. and Parkinson, D. (1978). Effect of freezing and thawing on survival of 3 bacterial isolates from an arctic soil. Can. J. Microbiol. 24: 1468–1474.
28. Orchard, V.A. and Cook, F.J. (1983). Relationship between soil respiration and soil moisture. Soil Biol. Biochem. 15: 447–453.
29. Reddy, K.R. and Patrick, W.H. Jr. (1975). Effect of alternate aerobic and anaerobic conditions on redox potential, organic matter decomposition, and nitrogen loss in a flooded soil. Soil Biol. Biochem. 7: 87–94.
30. Robertson, E.V. and Firestone, M.K. (1992). Relationship between desiccation and exopolysaccharide production in a soil Pseudomonas sp. Appl. Environ. Microbiol. 58: 1284–1291.
31. Rousk, J., Elyaagubi, F.K., Jones, D.L., and Godbold, D.L. (2011). Bacterial salt tolerance is unrelated to soil salinity across an arid agroecosystem salinity gradient. Soil Biol. Biochem. 43: 1881–1887.
32. Schmid, M.C., Maas, B., Dapena, A. et al. (2005). Biomarkers for in situ detection of anaerobic ammonium‐oxidizing (anammox) bacteria. Appl. Environ. Microbiol. 71: 1677–1684.
33. Schmidt, I., Slickers, O., Schmid, M. et al. (2002). Aerobic and anaerobic ammonia oxidizing bacteria – competitors or natural partners. FEMS Microbiol. Ecol. 29: 175–181.
34. Schniirer, J., Clarholm, M., and Rosswall, T. (1985). Microbial biomass and activity in an agricultural soil with different organic matter contents. Soil Biol. Biochem. 17: 611–618.
35. Setia, R., Marschner, P., Baldock, J. et al. (2011). Salinity effects on carbon mineralization in soils of varying texture. Soil Biol. Biochem. 43: 1908–1916.
36. Skopp, J., Jawson, M.D., and Doran, J.W. (1991). Steady‐state aerobic microbial activity as a function of soil water content. Soil Sci. Soc. Am. J. 54: 1619–1625.
37. Sorensen, L.H. (1974). Rate of decomposition of organic matter in soil as influenced by repeated airdrying—rewetting and repeated additions of organic matter. Soil Biol. Biochem. 6: 287–292.
38. Tate, R.L. and Ensign, J.C. (1974). Picolinic acid hydroxylase of Arthrobacter picolinophilus. Can. J. Microbiol. 20: 695–702.
39. Tate, R.L. III (1979). Effect of flooding on microbial activities in organic soils: carbon metabolism. Soil Sci. 128: 267–273.
40. Tate, R.L. III (1980). Microbial oxidation of organic matter of histosols. In: Advances in Microbial Ecology, vol. 4 (ed. M. Alexander), 169–201. New York: Plenum Press.
41. Tate, R.L. III (1987). Soil Organic Matter: Biological and Ecological Effects. New York: Wiley.
42. Thiemann, L.K. and Billings, S.A. (2012). Tracking C and N flows through microbival biomass with increased soil moisture variability. Soil Biol. Biochem. 40: 11–222.
43. Wildung, R.E., Garland, T.R., and Buschbom, R.L. (1975). The interdependent effects of soil temperature and water content on soil respiration rate and plant root decomposition in arid grassland soils. Soil Biol. Biochem. 7: 373–378.
44. Wilson, J.M. and Griffin, D.M. (1975). Water potential and the respiration of microorganisms in the soil. Soil Biol. Biochem. 7: 199–204.
45. Yan, N. and Marschner, P. (2012). Response of microbial activity and biomass to increasing salinity depends on the final salinity, not the original salinity. Soil Biol. Biochem. 53: 50–55.