Bacterial Biogeochemistry: The Ecophysiology of Mineral Cycling

By Tom Fenchel, Henry Blackburn and Gary M. King

Bacterial Biogeochemistry, Third Edition focuses on bacterial metabolism and its relevance to the environment, including the decomposition of soil, food chains, nitrogen fixation, assimilation and reduction of carbon nitrogen and sulfur, and microbial symbiosis. The scope of the new edition has broadened to provide a historical perspective, and covers in greater depth topics such as bioenergetic processes, characteristics of microbial communities, spatial heterogeneity, transport mechanisms, microbial biofilms, extreme environments and evolution of biogeochemical cycles.

• Provides up-to-date coverage with an enlarged scope, a new historical perspective, and coverage in greater depth of topics of special interest
• Covers interactions between microbial processes, atmospheric composition and the earth's greenhouse properties
• Completely rewritten to incorporate all the advances and discoveries of the last 20 years such as applications in the exploration for ore deposits and oil and in remediation of environmental pollution

• Language: English
• Publisher: Academic Press
• Release date: Jul 24, 2012
• ISBN: 9780124159747

Tom Fenchel

In 1986, Tom Fenchel was the recipient of the Ecology Institute Prize and the Huntsman Medal for Excellency in Oceanography. He is an honorary member of the Society for General Microbiology and is a Professor of Marine Biology and the Director of the Marine Biological Laboratory at the University of Copenhagen in Denmark. He has authored and co-authored several books and has published about 120 original and review papers on microbial ecology, marine biology and population biology. He holds a Ph.D and a Dr.Sc. from the University of Copenhagen and is a member of the Danish Royal Academy of Sciences and of the Royal Swedish Academy of Sciences.

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Table of Contents

Cover image

Title page

Copyright

Preface

Introduction

Chapter 1. Bacterial Metabolism

1.1 General Considerations: Functional Properties of Bacteria

1.2 Bacterial Metabolism

1.3 Dissimilatory Metabolism

1.4 Assimilatory Metabolism

1.5 Bioenergetics of Microbial Metabolism

Chapter 2. Transport Mechanisms

2.1 Physical Transport Mechanisms

2.2 Bacterial Motility and Sensory Motile Behaviour

Chapter 3. Degradation of Organic Polymers and Hydrocarbons

3.1 Substrates and the Efficiency of Degradation

3.2 Hydrolytic Enzymes

3.3 Mineral Nutrients and Decomposition Rates of Plant Derived Detritus

3.4 Humic Material and Hydrocarbons

Chapter 4. Comparison of Element Cycles

Chapter 5. The Water Column

5.1 The Composition of Planktonic Prokaryote Communities

5.2 Organic Matter: Composition, Origin and Turnover

5.3 Suspended Particles: Formation and Coupling Between Plankton and Sediments

5.4 Bacteria and Cycling of N and P

5.5 The Fate of Bacterial Cells

5.6 Motile Chemosensory Behaviour

5.7 Stratified Water Columns

Chapter 6. Biogeochemical Cycling in Soils

6.1 Soil Water as a Master Variable for Biogeochemical Cycling

6.2 Water Stress Physiology

6.3 Responses to Plant Organic Matter

6.4 Responses of Soil Biogeochemistry to Disturbance and Change

Chapter 7. Aquatic Sediments

7.1 Vertical Zonation, Vertical Transport, and Mixing

7.2 Element Cycling in Sediments

7.3 Sediments in the Light

7.4 Microbial Mats

Chapter 8. Microbial Biogeochemistry and Extreme Environments

8.1 Microbial Biology and Extreme Environments: An Overview

8.2 Biogeochemistry and Extreme Environments

8.3 Hypersaline Microbial Mats as Model Extreme Environments

8.4 Sub-Surface Environments as Extreme Systems

8.5 Thermophiles and Hyperthermophiles in Extreme Environments

8.6 Additional Considerations

Chapter 9. Symbiotic Systems

9.1 Symbiotic Polymer Degradation

9.2 Symbiotic N2 Fixation

9.3 Autotrophic Bacteria as Symbionts

Chapter 10. Microbial Biogeochemical Cycling and the Atmosphere

10.1 The Atmosphere as an Elemental Reservoir

10.2 Atmospheric Structure and Evolution

10.3 Synopsis of Trace Gas Biogeochemistry and Linkages to Climate Change

10.4 Trace Gas Dynamics and Climate Change: An Analysis of Methane Production and Consumption

Chapter 11. Origins and Evolution of Biogeochemical Cycles

11.1 Biogeochemical Cycles and Thermodynamics

11.2 Pre-Biotic Earth and Mineral Cycles

11.3 The Earliest Life and its Origin

11.4 Precambrian Life and Precambrian Biogeochemical Cycling

APPENDIX 1. Thermodynamics and Calculation of Energy Yields of Metabolic Processes

APPENDIX 2. Phylogeny and Function in Biogeochemical Cycles

Index

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Front cover photo credits:

Background (G.M. King): Lake Waiau, a meltwater and ground water derived lake at 3970 meters in the base of Pu’u Waiau, a cinder cone on Mauna Kea, Hawai’i

Insert photos, left to right (T. Fenchel):

An approximately 1 mm long organotrophic bacterium isolated from seawater.

The filamentous colourless sulfur bacterium, Thiothrix, attached to decaying seaweed.

The filamentous cyanobacterium, Oscillatoria, showing autofluorescence in green light due to the presence of phycoerythrin.

A filamentous cyanobacterium showing thick walled heterocysts in which N2-fixation takes place.

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Preface

This book treats the influence of bacterial activity on the chemical environment of the biosphere. Our approach is primarily one based on physiological properties of prokaryotic organisms. The book is the 3rd edition – the previous ones being Fenchel, Blackburn, 1979 and Fenchel, King, Blackburn, 1998. There are no radical changes with respect to the basic structure of the book relative to the 2nd edition, but most chapters have been rewritten and brought up-to-date in the light of the many advances and discoveries that have taken place during the last decade.

The topics treated in the book are central to many aspects of environmental science and to aquatic and terrestrial ecology. The book presupposes some basic knowledge of general microbiology, biological energetics and chemistry. We hope it will serve as a general text for university courses in microbial ecology and environmental sciences and also that the book, or parts of it, will prove useful for professional workers within aquatic and soil sciences, general microbiology, and geochemistry.

Introduction

The surface of Earth is in a state of chemical disequilibrium. If Earth had remained lifeless, its atmosphere and seas would have approached closer to a state of chemical equilibrium perhaps similar to that of Mars or more likely Venus, since Mars was unable to retain an atmosphere. Of course, complete equilibrium on Earth would never be attained due to tectonic processes (volcanism, mountain building and subsequent erosion) and to photochemical processes in the atmosphere and oceans.

Earth is about 4.6 billion years old and life arose perhaps four billion years ago. About half of the time there has been life on Earth it was represented only by bacteria.

In this book, we consider the term bacteria to be synonymous with prokaryotes, that is, the members of the domains Bacteria (Eubacteria) and Archaea (Archaebacteria): organisms that are not eukaryotes. While we are fully aware of the profound differences between Bacteria and Archaea, they are quite similar with respect to basic cellular organization and general functional properties. Thus, we will use the term Archaea when we wish to refer to that group specifically, and Bacteria to refer specifically to their counterparts. The generic term bacteria will refer to both groups collectively that is, the simple unicellular life forms lacking a membrane-bound nucleus classified within the domains Bacteria (Eubacteria)+Archaea (Archaebacteria).

These simple organisms evolved an astonishing diversity of basic types of metabolism based on compounds that were available in the environment and on electromagnetic radiation from the Sun. Tectonic and weathering processes became essential – as they are still−for the maintenance of life through recycling of essential compounds, especially carbon, phosphorus and sulfur, which became buried in sedimentary rocks. The resulting interactions between microbiological processes and various geological and chemical processes are largely responsible for the chemical properties of the extant seas and atmosphere.

Biological processes are in principle driven by chemical oxidation and reduction (or redox) reactions, that is, a coupling of two half cells through an exchange of electrons (see Appendix 1). A modest number of such processes form the basis for microbial metabolism; they mainly involve compounds of carbon, nitrogen, oxygen, iron, and sulfur. They are driven by a number of microbial engines (Falkowski et al., 2008), which have been conserved through evolution. Eukaryotes that arose about two billion years ago have inherited only a subset of processes that occur in different types of bacteria.

Bacterial activity, coupled with geological processes, altered the chemical environment of Earth’s surface so that it was conducive for the evolution of multicellular life forms. The evolution of oxygenic photosynthesis in cyanobacteria was undoubtedly the single most important step, since the origin of oxygenic photosynthesis gave rise to an O2-containing atmosphere without which multicellular organisms cannot survive. Many key processes of the biosphere are still carried out exclusively by bacteria and the major biogeochemical cycling of elements would likely proceed much as they do today – even if eukaryotes had never evolved.

Viewed from an ecological or biogeochemical perspective, metabolic processes tend to come in complementary pairs; for example, in the absence of oxygen some bacteria use sulfate as an electron acceptor in a respiratory process for oxidizing hydrogen or low molecular weight organic compounds; hydrogen sulfide is produced as an end product. When oxygen or nitrate is available, other bacteria in turn oxidize the sulfide back to sulfate, in effect forming a cycle.

The recognition that bacterial activity affects the chemical environment of the surface of Earth developed gradually during the last century. It was first related to the discovery of a wealth of novel bacterial metabolic processes and to the use of enrichment cultures, Winogradsky columns, work that was pioneered by Winogradsky and by Beijerinck before 1900. They discovered chemolithotrophy and nitrogen fixation among many other processes. Their scholarly tradition was initially carried on mainly by the Delft school of microbiology including: Kluyver, van Niel, Baas-Becking and their numerous students in the first half of the twentieth century. Their work and approaches helped establish the research areas of microbial ecology and microbial biogeochemistry, which have become an integral part of modern microbiology. Certain applied problems, especially soil fertility and nitrogen transformations in agricultural soils also played a role in the early development of microbiology. The study of bacterial communities and processes of aquatic habitats was also taken up by Zobell and his students in the middle of the twentieth century and aquatic microbiology has developed rapidly during the last four decades. Geochemists and chemists have also taken interest in how biological processes shape the chemical environment of seawater and atmosphere exemplified by Vernadsky in 1926 (Vernadsky, 2007) who popularized the term biosphere and by Sillén (1966).

Discoveries of novel types of microbial metabolism still take place and remain an inspiration for ecological work. During the last four decades the field has been characterized by the rapid development and deployment of new methods, including increasingly sensitive methods of chemical analysis, the use of radioactive and stable isotopes to determine pathways and rates of processes, the development of microelectrodes and advanced microscopy and imaging tools that all allow for description of chemical zonation and for estimates of process rates at sub-millimeter spatial scales and at very short temporal scales.

Molecular biological methods (genomics and metagenomics) have provided a framework for understanding microbial evolution as well as understanding the extent and distribution of microbial diversity. Molecular approaches also provide detailed information on which specific metabolic processes occur in which habitats (see e.g., DeLong & Karl, 2005). Analyses range from complex communities of organisms to single cells.

The pivotal role of bacterial mass and energy transformations in natural ecosystems has enjoyed growing recognition among ecologists, and microbial ecology has become an important component in the study of aquatic and terrestrial ecosystems. Microbial transformations of the chemical environment play a central role for the understanding of early diagenesis and of palaeoenvironments. Bacterial metabolic processes also impinge on a number of applied problems including the degradation of oil spills and of xenobiotic compounds, sewage treatment, eutrophication of aquatic systems, emission of greenhouse gases, and ore leaching in mines.

Our book is divided into 11 chapters. The first chapter includes various general considerations on basic aspects of the functional biology of bacteria with the main emphasis on the types of metabolism and considerations on bioenergetics. Chapter 2 is devoted to transport mechanisms in the environment (diffusion, advection, turbulence), the role of chemosensory motile behaviour, and the spatial structure of microbial communities. Together, these aspects form the basis for understanding patterns and rates of microbial processes in nature. The following chapter treats the hydrolysis of polymers and degradation of hydrocarbons and includes a discussion of the biologically driven element cycles. The succeeding five chapters describe microbial processes in particular habitats: the water column, soils, the rhizosphere and water saturated soils, marine and freshwater sediments, microbial mats and stratified water columns, symbiotic systems, and extreme environments. These chapters do not provide a comprehensive treatment of all aspects, but mainly emphasize general principles and the control of reaction rates. Chapter 10 considers global element cycling and the role of microbial processes in terms of controlling the abundance of gaseous phases of C, N and S in the atmosphere. The last chapter then is devoted to the early evolution of life and of biogeochemical cycles. An appendix treats thermodynamic principles and redox potentials and an additional appendix provides an overview of bacterial taxonomy.

Chapter 1

Bacterial Metabolism

1.1 General Considerations: Functional Properties of Bacteria

Bacteria are small: typical bacteria measure between 0.5 and 2 μm in diameter. A few are somewhat smaller, the so-called nanoarchaea, represented by Nanoarchaeum equitans, which is about 0.4 µm in diameter is an obligate symbiont of another, larger archaeum (Waters et al., 2008). Structures smaller than about 0.4 µm have been claimed to be bacteria, but in what is regarded as free-living forms, none of these have yet proven to be metabolically active organisms, nor have their fossils.

A few bacteria are considerably larger than 2 µm: some cyanobacterial cells exceed 5 μm and some sulfide oxidizing bacteria may reach a size of 20 μm or more (Thiovulum, Beggiatoa); Achromatium has been recorded to measure up to 0.1 mm, and Thiomargarita has a diameter of 0.75 mm (Schultz & Jørgensen, 2001). It would seem that there is a size-overlap with unicellular eukaryotes, the tiniest of which measure 2–3 μm, but most are five to 100 times larger. In contrast to small eukaryotes most of the volume of very large bacteria is constituted by a vacuole or by inclusions.

Most bacteria are unicellular, although some form colonies that are filamentous or otherwise shaped. Bacterial cells may be rod-shaped (rods), spherical (cocci), comma-shaped (vibrios) or helicoidal (spirilla), but other morphotypes occur as well. Some soil bacteria, in particular, form fungi-like mycelia (actinobacteria, myxobacteria), and myxobacteria have complex life cycles including the formation of sporangia. Bacteria almost always have a rigid cell wall surrounding the cell membrane. Exceptions include obligate intracellular parasites (e.g., Chlamydia) for which the protection against water stress provided by a cell wall is not necessary.

The two important characteristics of bacteria (small size, rigid cell walls) are necessary consequences of the absence of a cytoskeleton, a trait that characterizes eukaryotic cells. These traits explain two additionally important properties of bacteria. One is that bacteria can take up only low molecular weight compounds from their surroundings via the cell membrane and this uptake is brought about either by active (energy-requiring) transport or by facilitated diffusion. Bacteria that utilize polymers or particulate organics can do so only indirectly through extracellular hydrolysis of the substrate catalyzed by membrane-bound or excreted hydrolytic enzymes before the resulting low molecular weight molecules can be transported into the cells (see Chapter 3). Bacteria cannot bring particulate material or macromolecules into their cells; the capability of phagocytosis or pinocytosis is a privilege of eukaryotic cells. Bacterial transformation, which involves uptake of single stranded DNA by bacteria, represents an exception with evolutionary implications. Another consequence of the absence of a cytoskeleton is that all transport within the bacterial cell depends on molecular diffusion and this limits the maximum sizes that bacterial cells can attain. On the other hand, the small size of bacteria renders them extremely efficient in concentrating their substrates from very dilute solutions (see Chapter 2).

Finally, a consequence of small size – when comparing organisms spanning a large size spectrum – is a high rate of living or metabolic rate; that is, small organisms tend to have higher volume-specific metabolic rates and shorter generation times than do larger organisms. Roughly speaking, when comparing organisms of widely different sizes, specific growth rate constants and volume-specific metabolic rates are proportional to (volume)−1/4, notwithstanding that there may be variation in potential growth rates among species of similar size. Under optimal conditions many bacteria have generation times of only 15–30 minutes, with as little as ten minutes the fastest known. Generation times for a 100 μm long protozoan, a copepod and a small fish would be roughly eight hours, 10 days and one year, respectively.

Although the total biomass of bacteria may not be large relative to that of multi-cellular organisms in some habitats (especially terrestrial systems), the impact of bacteria in terms of matter transformations and energy flow may be much greater. For example, seawater typically contains around 10⁶ bacteria per ml of water resulting in a volume fraction somewhat less than 10−6. This is comparable to the volume fraction made up by protozoa; however, the metabolic activity of the bacterial community may be roughly an order of magnitude higher than that of the protozoa.

Another property important for understanding the role of bacteria in nature is that they hold all records as extremophiles. Some bacteria live at temperatures exceeding 80°C or even up to the temperature of an autoclave, 121°C under hyperbaric pressure (extreme thermophiles). Others thrive in concentrated brine (extreme halophiles), at a pH<2 (acidophiles) or pH>10 (alkaliphiles), and some are tolerant to mM concentrations of toxic metal and metalloid ions such as As, Cu, Zn, amongst others (see Chapter 10). Other habitats not usually considered extreme in the senses above, exclude most multi-cellular organisms, and are inhabited almost entirely by bacteria in practice. Such habitats include anoxic, strongly sulfidic waters and sediments (which otherwise harbour only a few types of specialized protozoa) and some sediments rich in clay and silt with small pore sizes that preclude many larger organisms.

In contrast to aquatic systems and extreme environments in which bacteria are dominant, terrestrial systems (soils and the litter layer) often support communities of fungi that rival or exceed bacteria in biomass and activity. This is especially true for the primary decomposition of plant structural compounds (e.g., cellulose and lignocellulose), which fungi typically dominate. One reason for the more limited role of bacteria in terrestrial systems is that among all the possible types of physical and chemical constraints found in nature, bacteria seem to have only one absolute requirement for activity: liquid water. Many bacteria, especially soil isolates, produce desiccation-resistant structures, e.g., cysts and spores. However, metabolic activity and growth require water, and because of this requirement, growth and metabolic activity of terrestrial bacteria is confined to micrometer-thick aqueous films that cover mineral and detrital particles in soils, the surfaces of rocks, litter, and roots, stems and leaves of living plants.

Relative to bacteria, fungi can tolerate water stress to a much greater extent, and are not constrained to aqueous films. Indeed, fungal hyphae can ramify through gas-filled soil pores as well as cellulosic walls of plants, thus exploring the soil space and promoting plant polymer decomposition. In this respect, fungi are better adapted to life in soil and litter. The relation between fungi and bacteria is discussed in more detail in Chapter 5.

Yet another profoundly important reason for the pivotal role of bacteria in all ecosystems is their metabolic diversity. Some species of bacteria are very specialized with respect to their substrates and available metabolic pathways. But the metabolic repertoire of bacteria taken together far exceeds that known from eukaryotes. Examples of processes that are exclusively carried out by certain bacteria include methanogenesis, the oxidation of methane and other hydrocarbons, and nitrogen fixation. These and several others that are carried out exclusively by different kinds of bacteria are all key processes in the function of the biosphere.

Similarly, bacteria collectively have an astonishing capability to hydrolyze virtually all natural polymers as well as many unusual compounds such as secondary plant metabolites, compounds found in crude oil, and many xenobiotics. The degradation of polymers, which is a question of extracellular hydrolysis, is treated in Chapter 3. Here we proceed with a discussion on bacterial metabolic diversity.

1.2 Bacterial Metabolism

Bacteria, like all living organisms, are capable of increasing in size (growth) and dividing (reproduction). Bacterial activities are directed to this end, and this requires energy and a variety of substrates from the environment for the synthesis of cellular material. These two activities, i.e. obtaining energy and obtaining or synthesizing building blocks for growth are referred to as dissimilatory (energy or catabolic) metabolism and assimilatory (anabolic) metabolism, respectively.

It is convenient to discuss these separately, and we do so in the following passages. However, the two types of metabolism are tightly coupled in the sense that microorganisms spend by far the most power they generate on growth due to the high energetic costs of macromolecule synthesis (DNA, RNA and proteins), and of transport of molecules in and out through the cell membrane (see Table 1.1).

Table 1.1. Bacterial Energy Budget for Cells Grown on Glucose (Based on Stouthamer 1973)

Under most normal growth conditions there is, therefore, an almost linear relation between the growth rate constant (measuring the balanced increase in biomass) and the rate of power generation. Furthermore, a particular substrate may serve both as an energy source and as a carbon source. Thus, a bacterium growing aerobically on glucose will use this substrate partly as a source of energy (oxidizing it to CO2) and partly as a source for cell material (largely without changing the oxidation level of the C atoms). In other cases, the energy source and the assimilated materials are different. This is trite in the case of phototrophs, but it also applies, for example, to sulfide-oxidizing bacteria, which must assimilate CO2 or some other material. Finally, the enzymes involved in assimilatory and dissimilatory metabolism may overlap with identical metabolic pathways serving as oxidative, catabolic pathways in some species or under some circumstances – running in reverse – as reductive anabolic pathways in other species or circumstances. For example, the citric acid (TCA) cycle is used in most respiratory organisms for the stepwise oxidation of acetate to CO2. However, in the phototrophic green sulfur bacteria (e.g., Chlorobium), the Aquificales, some Proteobacteria, and in some Archaea, the citric acid runs in reverse and is used as a synthetic, reductive pathway for the assimilation of CO2. In the former case it is an oxidative energy generating pathway, in the latter case it is a reductive energy requiring (ATP-consuming) pathway. In the purple non-sulfur bacteria (e.g., Rhodopseudomonas), the same electron transport system is used for both respiration and for photophosphorylation (Fig. 1.1). These and similar examples are of considerable interest in an evolutionary context because they illuminate the origin and evolution of metabolic pathways; they also show how a relatively small number of basic pathways can lead to a relatively large metabolic repertoire (see Chapter 11). Under all circumstances, it should be kept in mind that while the distinction between dissimilatory and assimilatory metabolism is meaningful in some contexts, the two types of metabolism are in other respects deeply intertwined.

Figure 1.1 Metabolic pathways in a purple non-sulfur bacterium. In the light, the activated bacteriochlorophyll molecule transfer an electron to the electron transport chain and back to the bacteriochlorophyll. In this process protons are expelled from the cell and the return flux is coupled to ATP synthesis. Also, in the light electrons can be passed via NADPH for reducing CO 2 to organics matter. In the dark, external electron donors can be passed through the electron transport chain to an external electron acceptor: O 2 – also acting as a proton pump and resulting in ATP generation.

The terms autotrophy and heterotrophy can be applied to both the dissimilatory and assimilatory metabolism. A heterotroph depends on organic material for energy generation and for precursors for the synthesis of cell material. Autotrophs are independent of organic material and assimilate CO2 as a source of cell carbon.

Photoautotrophs use the energy of electromagnetic radiation (roughly 400 nm to 1100 nm wavelengths) for ATP-generation and for obtaining reducing power for CO2 assimilation. Chemoautotrophs (chemolithotrophs, lithotrophs) obtain energy by oxidizing inorganic substrates (e.g., HS−, Fe²+NH4+) with inorganic electron acceptors, such as O2, NO3− and Fe³+, and obtain cell carbon by assimilatory CO2 reduction.

The terms autotrophy and heterotrophy are rarely absolute. Many organisms can function as autotrophs or heterotrophs under different circumstances. Purple non-sulfur bacteria, for example, metabolize as autotrophs in the light using H2 or HS− as reductants for photosynthesis and assimilating CO2, but they can also photoassimilate acetate and other low molecular weight organics, and in the dark they respire heterotrophically using O2 and various low molecular weight organics as substrates. In addition, some chemoauthotrophs are capable of assimilating organic substrates and some colourless sulfur bacteria can subsist in the absence of O2 through fermentation or sulfate reduction using organic substrates. Some autotrophs also require organic growth factors or vitamins. The concepts of autotrophy and heterotrophy may also be extended to other elements than carbon, such as nitrogen. A bacterium that covers its need for N by assimilation of inorganic N-compounds (NH4+, NO3−, NO2−) is autotrophic with respect to N whereas requirement of organic N (e.g., amino acids) for N- assimilation would be considered heterotrophic.

Finally, while the term autotrophy somehow implies independence of the products of other organisms, this is also a question of context. Purple sulfur bacteria, for example, are photoautotrophs (using HS− as an electron donor in a photosynthetic process and assimilating CO2). In most habitats, however, the reducing power of sulfide derives from plant material originally produced by oxygenic photosynthesis and subsequently degraded under anaerobic conditions. So in an ecosystem context, the purple bacteria are then only a link in a detritus food chain driven by oxygenic photosynthesis. The more exotic situation where phototrophic sulfur bacteria depend on sulfide of geothermal origin would, to a larger extent, justify the term autotrophy.

In this book we are primarily concerned with microbe-mediated chemical transformations of the environment rather than with intracellular physiology. In the following narrative we therefore emphasize what is taken up and what is produced as metabolites, that is, the net results of bacterial metabolism. Our discussion of the metabolic pathways within the cells is therefore limited to what is necessary to understand the energetics of the processes and thus why certain types of metabolism are favoured over others under particular circumstances. In this context the bioenergetics of dissimilatory metabolism is of central significance and is treated in some detail in the following. Accounts of the biochemistry of bacterial metabolism can be found in Madigan et al. (2011); for a general treatment of bacterial biology and diversity, see Dworkin et al. (2006). For phototrophic, anaerobic, chemoautotrophic, and fermentative types of metabolism, in particular, see also: Fenchel & Finlay (1995), Schlegel & Bowden (1989), and Zehnder (1988) can be consulted. During the past two decades a number of novel types of energy metabolism have been discovered and some organisms have proven to have a wider metabolic repertoire than previously believed.

1.3 Dissimilatory Metabolism

Kluyver & Donker (1926) drew attention to what they referred to as unity of biochemistry; they pointed out that in bacteria (and other organisms) energy-yielding metabolic processes seem always to be coupled redox reactions of the type: AH2+B ↔ BH2+A, or in other words, reactions that involve electron transfer between two half-cells. This generalization still holds. The particular coupled reactions must be processes that result in a decrease of free energy of the system (Gibbs free energy ΔG<0); for more details see Chapter 1.5 and Appendix 1). The substrates used by the organisms are taken up from the environment. In some cases the reactions used by bacteria for energy conservation will also occur spontaneously – e.g., the oxidation of sulfide by oxygen. Other reactions, such as the oxidation of ammonia or of many organic substrates will take place very slowly or not at all outside the organisms because these reactions have high activation energies. An important function of energy metabolism is, therefore, to catalyze chemical processes towards equilibrium in addition to conserving the released energy in a form that is useful to the cell.

In living cells energy is conserved first of all as adenosine triphosphate (ATP) in the reaction ADP+Pi+energy→ATP+H2O, where ADP is adenosine diphosphate and Pi stands for inorganic phosphate. The ΔGo’ (Gibb’s free energy change of hydrolysis under standard conditions, 25°C, one atm, molar concentrations and pH=7) of ATP is−29.3 kJ mol−1. The cells in turn use ATP to power vital processes: primarily synthesis of macromolecules and active transport across the cell membrane.

There are two different methods of ATP synthesis: substrate-level phosphorylation and oxidative phosphorylation. In substrate-level phosphorylation ATP is synthesized at specific steps in the catabolism of a substrate so that one mol of ATP is produced from the transformation of one mol substrate, provided the free energy change of this transformation exceeds 29.3 kJ mol−1. In oxidative phosphorylation there is no such strict stoichiometric coupling between transformation of substrate molecules and ATP synthesis. Phosphorylation occurs during both respiration and photosynthesis; it depends on membrane-bound enzymes of electron transport chains, which include H+ (protons) or in some cases Na+- pumps that expel these ions from cells. The resulting electrochemical gradient creates a proton motive force, which leads to a return flux of H+ or Na+ into the cell. This flux is then coupled to ATP synthesis, which is catalyzed by membrane-bound ATP synthase molecules.

The classification of energy-yielding processes is not simple because there are many exceptions and unclear boundaries between some categories. Here we will distinguish between fermentation, respiration, methanogenesis, and phototrophy.

Fermentation

Fermentations are energy-yielding, anaerobic processes in which substrates are sequentially transformed by reduction-oxidation processes. No external electron acceptor is involved; that is, the redox levels of the substrate and the metabolite(s) remain the same. Thus fermentations represent a dismutation of the substrate molecules. In fermentations a relatively low amount of energy is conserved: fermentation of one mol of glucose yields 2–4 moles of ATP depending on the type of fermentation, while respiration with O2 as terminal electron acceptor yields about 32–36 ATP.

A comparison of fermentation and respiration is shown in Table 1.2. None of the criteria, however, are absolute. Thus the dismutation of S2O3²− into sulfide and sulfate – a microbial process that takes place in anaerobic sediments (Bak & Cypionka, 1987) – is technically a type of fermentation, but it is based on an inorganic substrate. In some succinate/propionate fermentations the step in which fumarate is reduced to succinate is coupled to oxidative phosphorylation, rather than to substrate-level phosphorylation. In some cases the excretion of fermentation products results in a proton motive force, which is exploited in membrane-facilitated ATP synthesis. Strictly speaking, some types of fermentations in which externally derived CO2 or H2O are used as electron acceptors do not represent a complete redox balance. Some fermenters can also dump reducing equivalents to external compounds such as nitrate or oxidized Fe as an alternative to H2-excretion.

Table 1.2. Properties of Fermentation and Respiration

Fermenting bacteria may be facultative anaerobes capable of oxidative phosphorylation in the presence of O2 (e.g., Escherichia), aerotolerant anaerobes (e.g., Lactobacillus) or they may be strict (O2-sensitive) anaerobes (e.g., Clostridium).

In the fermentation of glucose to lactate or to ethanol+CO2, the ATP yield is only 2 mol per mol substrate. This is because pyruvate (resulting from glycolysis) is used to re-oxidizing reduced nicotinamide adenine dinucleotid (NADH) produced during glycolysis, thus restoring redox balance and producing lactate or ethanol+CO2 as end products. This is wasteful in the sense that further fermentation of pyruvate to acetate and thus additional generation of ATP would otherwise be possible. In nature, lactate and ethanol+CO2 fermentations are important only where easily degradable sugars occur at high concentrations. In such environments, lactobacilli, which are acid tolerant, lower pH through acid production, and can maintain a competitive advantage over other types of fermenting bacteria once a large population has been established.

In mixed-acid fermentations some reducing equivalents are disposed of as formate, which in most cases is degraded to H2 and CO2. This allows for the oxidation of some pyruvate via acetyl-CoA to acetate. This last step is coupled to substrate-level phosphorylation allowing for the generation of additional ATP relative to homolactic fermentation. Other reducing equivalents from NADH are coupled to acetyl-CoA metabolism resulting in the production of acetate and H2 in addition to a mixture of compounds such as butyrate, succinate, lactate, and ethanol that are less oxidized than acetate. This type of fermentation is characteristic of enterobacteria.

Another way of restoring redox balance is found in clostridial-type fermentations. Clostridia have an