Ecology, Microbial

Microorganisms are often considered to be among the first life forms on earth. Fascination with these beings was initially predisposed to their small size and simple forms. Early observations of microorganisms (in the seventeenth century) by Leeuwenhoek, with the aid of the first microscope, were followed by the demonstration of the role of microorganisms in the process of fermentation and spoilage by Pasteur and, eventually, the development of a means to prevent the growth of microorganisms, that is, pasteurization. 

I. EVOLUTION OF MICROBIAL ECOLOGY AS A DISCIPLINE 

Isolation of causative microorganisms of disease, as well as pure culture techniques, evolved in the late 1800s; Koch’s postulates provided a solid basis for studying microorganisms and their roles. The era that followed provided for the continued isolation of microorganisms, the definition of their metabolic capabilities, and, in turn, their implied roles in important biogeochemical processes, that is, in the nitrogen and sulfur cycle. Attention was given to organisms from specific habitats, that is, soil, water, animal, and plants. The findings demonstrated the vast numbers of diverse microbiological forms and functions found in nearly every location that was sampled. Eventually, microbiological subdisciplines dealing with microbial associations of natural (soil, water) and manmade (food, industrial, and other) environments were established. Within the last 30 years, it has been recognized that common microorganisms are observed in many habitats and common principles are involved in the mechanisms describing the associations of those microbes in varying habitats. The observation that individual populations of microorganisms are rarely found alone suggests potential interactions between populations and with their surrounding physicochemical environments. Additionally, the observation that they associate or align themselves within specific “strata” or gradients of physiochemical parameters points to the large number of metabolic functions of these organisms. They also have been shown to have the ability to sense (e.g. chemotaxis) and move by various means of locomotion within these gradients to maintain selected conditions for their growth. The field of ecology is defined as a discipline of biology which deals with organisms’ interactions with each other and surrounding environments. As one might expect, these aforementioned observations of microorganisms and their habitats led to the development of the subdiscipline known as microbial ecology. This development further emphasizes the need to establish a union between the examinations of the physicochemical nature of a habitat and microbiological investigations. An equally important emphasis in microbial ecology is the structure and activities associated with microbial communities rather than the earlier emphasis in causative populations of disease or specific processes. 

II. MICROBIAL COMMUNITIES 

Although observations of microorganisms and their associations have been made in numerous habitats, a few of these observations are felt to highlight the importance of the interactions between microorganisms which leads to community versus population responses. The rumen ecosystem has received considerable attention, which has been driven principally by economic considerations. Research in this system has, however, defined the syntrophic relationship between members of the microbial fermentative community and the animal’s growth and survival. Technically, approaches to this ecological niche present somewhat greater difficulties than those of soil or natural waters. The ecosystem is internal to the animal and the microbes are strict anaerobes. Mechanistically, however, rumen microbial ecology is relatively easier to discern, inasmuch as both input and output to and from the system are clearly defined, in much a similar relationship as an industrial bioreactor. It is interesting to note that syntrophic community-level interactions are frequently illustrated with the rumen system, in regard to hydrogen transfer and methane production. The classic anaerobic food web first described by Hungate, and later by Wolf, which occurs in the rumen, involves the interpopulation interactions among bacterial communities capable of plant polysaccharide hydrolysis (e.g. cellulose), monomer fermenting communities, fatty acid-oxidizing communities, and, finally, terminal communities (e.g. methanogenesis), which oxidize fatty acids and reduce CO2 to CH4. The hydrogen is derived from previous oxidative steps. The coupling, or syntrophic, relationship between hydrogen producers and hydrogen consumers (interspecies hydrogen transfer) is now recognized as a fundamental relationship in other systems dominated by anaerobic microorganisms. In the rumen, rates and extent of metabolism are controlled by the interdependence of one community on another. The rumen system has also served as an example of a strategy for, an approach to, and methods to conduct similar investigations of animal–microbe associations, be it the crop of tropical birds or the intestinal tract of termites. Similar interactions have been observed in other anaerobic systems (such as sewage sludge, lake and marine sediments), which suggest common controls and mechanisms associated with metabolism of complex organic compounds in all of these systems. These relationships have also been shown to be involved in the metabolism of naturally occurring and manmade halogenated compounds. One such reaction involved dehalogenation by replacement of a halogen substituent of a molecule with a hydrogen atom. The hydrogen is derived from hydrogenproducing fermentative microorganisms. Questions related to the response of these communities and the interactions among populations to disturbance of their physical–chemical environment will rely on tools which allow analyses of change in both the nature of the populations involved and changes in their functions. Do populations adapt and respond phenotypically or do they replace each other? Traditional food web descriptions in aquatic and terrestrial ecosystems have often failed to consider the role of microbial interactions as a contribution to carbon and nitrogen cycling in these systems. Early recognition of these contributions was blurred by the examination of individual populations of organisms and by inadequately examining the interactions and controls of population size and distribution in the surrounding communities. Figure 31.1 illustrates, in its simplest form, the relationship among primary producers in aquatic/marine or terrestrial environments, heterotrophic bacteria, and phagotrophic protozoans or zooplankton. Primary producers release soluble organic compounds (root exudates, algal metabolites), which are consumed by heterotrophic bacteria, which are subsequently grazed by phagotrophic protozoans or zooplankton. The grazers excrete nitrogen and phosphorus, which is used by the primary producers. In sum, these observations have strikingly modified the contemporary view of the structure and mechanistic controls and regulation of growth of higher plant and animal forms in aquatic and terrestrial systems. They also point to the importance of a thorough understanding of these interactions if one is to consider management of these associations in applied applications, for example, sustainable agriculture or aquaculture. Microbial biofilm communities were first described by Zobell and Anderson. Over the past two decades, their significance and ubiquity have been documented. It is now recognized that biofilm communities predominate numerically and metabolically in most ecosystems. Further, it is clear that biofilm cells are fundamentally phenotypically distinct from nonadhering cells. It now appears that the ability to form surface-associated structured biofilms is a common characteristic of, at least, bacteria. Remarkable intercellular and interspecies interactions, facilitated by chemicals released by these organisms, leads to complex structured communities, made up of both prokaryotic and eukaryotic organisms. These examples, however, primarily involve interactions based on an exchange of chemical metabolites. The fact that these described interactions involve high densities of cells in close proximity to each other, for example, biofilm, can also lead to intercellular exchange of genetic information, which can lead to adaptive change in function within communities. Knowledge of naturally occurring phenotypic and genotypic viability of microorganisms has been hampered because the majority of our knowledge comes from laboratory-selected strains, grown under controlled conditions, and on a medium which has no resemblance to the environment from which they were selected. It is important to realize that approaches to microbial ecology must recognize not only isolated individual populations but, more directly, the interactions of populations within a community and the resultant effect on the overall functions of the community. 

III. APPROACHES TO MICROBIAL ECOLOGY 

The fundamental approach to higher plant and animal ecological studies uses quantitative observations of specific populations in various environments. For over 100 years, plant and animal ecologists have observed and detailed the frequency of occurrence of specific plant and animal populations under various environmental conditions. These observations have led to the development of models for predicting the occurrence of, as well as relationship between, the organisms and the associated environments in which they were observed. Recently, these observations have been complemented with physiological and genetic approaches that suggest specific mechanisms of selection which have led to the observed frequency or distribution of organisms. Unfortunately, our understanding of microbial systems is still embryonic in both description and prediction. Kluyver and van Neil, in 1956, estimated that about one half the “living protoplasm” on earth is microbial. This estimate is now considered conservative, and the sheer numbers represent a daunting challenge to the microbial ecologist. Even the most modern techniques do not allow routine quantification of microbial numbers and definition of specific populations. It is principally the size of the microorganism, and, equally important, the size of the local habitat, that creates a considerable constraint on the observation of microorganisms in natural surroundings. Numbers of organisms ranging from 1 million cells per milliliter of water to 10 billion cells per gram of human fecal material. Further, the spatial distribution of these organisms within these habitats makes it difficult to recover a representative sample to examine. In situ observations are further complicated by our inability to observe this environment without disturbing the microorganism’s natural habitat. The various surfaces and potential differences in chemistries of the habitat provide varying degrees of carbon and nitrogen resources and physicochemical environments (e.g. gaseous exchange) for colonizing microorganisms. Additionally, other organisms, micro- and macroalike, have a potential impact on the native organisms. Grazing of microorganisms by other microbes or faunal components of a habitat have a significant impact on the numbers and types of microorganisms present. As noted, microbial ecology evolved as a crossdiscipline of standard microbiology and environmental analytical analysis. The activity of a specific population or community of microorganisms was inferred by estimating their relative numbers by direct microscope counts, viable colonial or turbidimetric determinations, or specific chemical analyses (e.g. chlorophyll for algae). Unfortunately, limitations by both the microbial and the environmental analyses have, in most cases, failed to accurately define the ecology of microbial habitats. Recently, novel analytical and microbiological methodologies have provided tools for expanding our view of microbial habitats, in a fashion that is neither constrained by the microscale of the environment, nor limited by our ability to selectively cultivate members of a community. In situ observation of microorganisms has been expanded through use of laser confocal microscopy, an advancement which provides for a kind of “X-ray” imaging of material requiring neither disturbance nor fixation. Similarly, laser optical trapping allows the removal of individual cells from a habitat. These techniques provide new insights to our understanding of population components of communities. These increases in optical resolution also provide a means of sorting communities either by size or chemical factors through the use of cell sorters. Advancements have also been made in developing methodology to simulate the microhabitats and physiochemical gradients which occur therein. It is interesting to note that one of the pioneering environmental microbiologists, Winogradsky simulated gradients of light, water, sulfide, and oxygen to describe relationships between sulfide-oxidizing photo-autotrophic bacteria, sulfate-reducing, and chemoautotrophic sulfuroxidizing organisms in his Winogradsky column. Advances using gels and gradostats to simulate diffusion barriers allow us to expand our knowledge concerning relationships between the spatial heterogeneity of the physicochemical environment and the distribution of diverse groups of microorganisms. Combined with microelectrode techniques, precise analytical measurements will complement these microbial investigations. Although our abilities to observe organisms and to better understand the physical and chemical nature of their habits have improved, we still are unable to isolate a high percentage of these observable organisms. Nevertheless, within the last 30 years, the number of previously undescribed microorganisms has increased significantly. In an effort to forego the inherent problems of the microscopic and numerical diversity of the microbial world, recent method development has approached the microbial component of the unseen habitats at the macromolecular level. The recognition that diversity is intrinsically related to the organism’s genetics has provided a sound basis for separation and characterization of both microscopic and macroscopic life. Examination of the heterogenity of DNA in soil suggests that as many as 10,000 bacterial species can be harbored in 100 g of soil. This estimate only includes the bacterial fraction and, again, reinforces the challenge presented to understanding the diversity of microorganisms in natural systems. From an ecological perspective, it also presents a challenge to understanding how so many species coexist in such small areas. Recombinant DNA technology has allowed the identification and determination of the specific nucleotide sequence of genes. It has provided ecologists with novel methods to pursue community structure. In practice, microbial community samples can be analyzed without cultivation or microscopic observation of microorganisms. In brief, nucleic acid is isolated from the sample and utilized for hybridization studies with the corresponding gene of interest (i.e. the probe). This probe can represent either a metabolic gene or a systematic determinant, such as the 16S rRNA gene sequence. In turn, as one may infer, the sample can be evaluated in terms of population diversity at the species level or community diversity at the kingdom level (Eukaryotic versus Prokaryotic). All of these analyses have been expanded by inclusion of the polymerase chain reaction (PCR), a method allowing amplification and subsequent detection of as few as 10 microorganisms. Interestingly, these methods have, in fact, relied on previous isolation and characterization of specific microbial populations prior to isolation of a gene for use as a probe. These investigations have, however, illustrated the universal nature of certain sequences, such as from 16S rRNA genes, for random use in community analyses. An example of this technology is the use of various profiling techniques. Separate PCR-amplified 16S rRNA gene-fragment sequences are separated, and the resulting numbers provide an estimate of diversity within the community. Areas that still remain to be explored in microbial ecology are the relationship of microbial diversity to the response of microbial communities to varying degrees of disturbance and the relationship between diversity and the function of the community. In higher plant and animal systems, the relationship of diversity within communities to their resistance to change or recovery after disturbance (e.g. fire, tillage) has been discussed and debated for decades. Various indices have been used to calculate diversity within communities. In their simplest form, these indices represent the number of species found within the community; therefore, communities with many species are described as having high diversity. Other indices relate the dominance of specific species to the total diversity, such that even communities with high diversity can have a few populations which dominate activity within the community. Our current inabilities to adequately isolate all microorganisms and the lack of distinctive morphological characteristics fail to provide us with accurate methods for measuring indices of diversity. Continued improvements in our analytical skills in identifying the macromolecular characteristics of microorganisms will provide a basis for estimating diversity at the phylogenetic level. Profiles of cellular or phospholipid fatty acids from lipids extracted from communities or specific chemical or antigenic determinants may provide a snapshot of changes within microbial communities after disturbance. Community-level assays of the functional diversity of populations based on carbon source utilization also provide indications of changes following disturbance. 

IV. FUTURE DIRECTIONS IN MICROBIAL ECOLOGY 

It has often been said that advancements in microbial ecology are limited by the methods which are available to analyze microbial systems. Some of the advances made in the analytical and molecular methods over the past decade have been illustrated in this overview. Although continued advancements in the use of microelectrodes and optical methods increase our abilities to measure and observe microorganisms emphasis must increase on the refinement of techniques to discern changes in microbial community structure and the impact of these changes on the function of the community. A significant area of contemporary concern is the remediation of habitats contaminated with anthropogenic sources of organic and inorganic compounds. The applied area of bioremediation is, and will continue to be, a timely subject in the decades to come. The use and management of the intrinsic properties of microbial communities will provide stimulus for applied microbial ecology. Although this area is yet to be accurately defined, in principle, the directive is to promote microbial dissimilation of anthropogenic compounds in a controlled manner. In a similar manner, a reduced input of anthropogenic chemicals in agricultural systems to reduce environmental contamination will result in greater reliance on activities of the soil microbial communities. These and other applications require further understanding of the structure and activities of microbial communities. Also required is an understanding of changes in the structure and function of these communities in relation to changes in the physiochemical environments associated with them. This, in fact, is microbial ecology.