INTRODUCTION

Interactions between dissimilatory sulphate-reducing bacteria (SRB) and metal(loid) ions have been studied since the first half of the twentieth century and the ability of SRB to bring about changes in the speciation of metal(loid)s has been recognized for much of this time. Early work focused on the role of SRB as nuisance organisms and metal(loid) interactions with SRB were often studied in the context of their use as metabolic poisons to control SRB activity (Postgate, 1952; Newport and Nedwell, 1988; Nemati et al., 2001). With growing awareness of the importance of microorganisms in biogeochemical cycling, the emphasis of research has shifted to the environmental roles of SRB. Their capacity to control the mobility of metals in aqueous sediments by the formation of poorly-soluble metal sulphides is now apparent and SRB-generated metal sulphides constitute an important environmental sink for many metals (Morse et al., 1987; see also Chapter 13, this volume). Bioremediation of dissolved metal(loid)s is an application for which SRB may be particularly suitable, given that sulphate frequently co-occurs with toxic metal ions in, e.g. metal-processing wastes and acid mine-drainage waters. SRB have the apparently unique potential to simultaneously remove metals, sulphate and acidity through the bioprecipitation of metal sulphides – a phenomenon that still represents one of the most successful biological approaches to metal removal from aqueous media. Other abiotic chemical reactions that can effect the removal of metal(loid) ions from solution take place during active sulphate-reduction.

One of the best parasitologists in the early part of the twentieth century for making discoveries involving parasite life cycles was Wendell Krull. The late, and great, Miriam Rothschild referred to him as “a genius who hid his light under a bushel” (Ewing, 2001). Part of her admiration for Krull stemmed from his research on the life cycle of Halipegus occidualis, the hemiurid fluke that lives under the tongues of North American ranid frogs. Others might consider him as a genius for his contribution in resolving the life cycle of Dicrocoelium dendriticum.

Wendell Krull is now dead, so I had to rely on other resources for special information regarding his career and some of the research he accomplished. In addition to a number of Krull's papers, one of the important biographical sources for this effort was a book written by Sidney Ewing, entitled Wendell Krull: Trematodes and Naturalists, published in 2001. It really is a delightful read. Over a period of eleven years, including his three years as a graduate student, Sidney had many conversations with Krull regarding his life's work. I have unashamedly quoted from Ewing's book throughout the essay. I also had the wonderful opportunity of interviewing Sidney Ewing and his wife, Margaret, who provided me with even more insights regarding Krull, including Margaret's experience in Krull's veterinary parasitology course at Oklahoma State University.

INTRODUCTION

Sulphate-reducing bacteria (SRB) are a phylogenetically and physiologically diverse group of bacteria, characterized by their versatile metabolic capability to use various electron acceptors and donors (Widdel, 1988). SRB are therefore universally distributed in diverse environments and play significant ecophysiological roles in anaerobic biomineralization pathways. The degradation of organic matter by a complex microbial community is governed to a large extent by available electron acceptors. The terminal stages of the anaerobic mineralization of organic matter is catalyzed by SRB and methanogens, and their competitive and cooperative interactions have been described previously (Oude Elferink et al., 1994).

Typical domestic wastewaters contain sulphate concentrations of 100–1000 μM and relatively low dissolved oxygen due to the lower solubility and rapid depletion of this gas by biological activity. Thus, sulphate reduction can be the dominant terminal electron accepting process and account for up to 50% of mineralization of organic matter in wastewater biofilms (Kühl and Jorgensen, 1992; Okabe et al., 2003a). Multiple electron donors and electron acceptors are present in the wastewaters. As a result, wastewater biofilms are very complex multispecies biofilms, displaying considerable heterogeneity, with regard to both the microorganisms present and their physicochemical microenvironments. Sulphate reduction is anticipated to take place in the deeper anoxic biofilm strata even though the bulk liquid is oxygenated. It is, therefore, thought that a successive vertical zonation of respiratory processes can be found in aerobic wastewater biofilms with a typical thickness of only a few millimeters (Ito et al., 2002b; Kühl and Jorgensen, 1992; Okabe et al., 1999a; 2003a; Ramsing et al., 1993).

As it turns out, the title for this book is hugely misleading. When I began writing about two years ago, I really thought that many of the discoveries in parasitology were of a serendipitous nature. However, after talking with my group of scholars/parasitologists and seriously evaluating some of the historical aspects of parasitology, I discovered this is simply not the case. I would say, in fact, that of all the major discoveries made in the field of parasitology over the last 150 years, no more than 5–10% can be considered as truly serendipitous. Yes, there were occasions when somebody found something, or saw something, for which they were not looking. For example, in one of my first interviews, Dick Seed pointed out that very early in the twentieth century, there were serendipitous discoveries made regarding antigenic changes in the African trypanosomes. Keith Vickerman observed, quite by accident, some sort of ‘coat’ on trypomastigotes of T. brucei in the salivary gland of a tsetse fly. He put ‘two and two’ together and came up with a hypothesis that, when tested, generated positive evidence for the presence of VSGs. Sidney Ewing discovered erhlichiosis in a North American dog by accident. Arthur Looss, while working in Egypt, accidentally dropped culture medium containing L3 larvae of hookworm on his own skin and this led him to identify the entry route for these parasites into humans.

THE HUMAN LARGE INTESTINAL ECOSYSTEM

The adult human colon typically contains over 200 g of digestive material (Banwell et al., 1981; Cummings et al., 1990; 1992), with the average daily output of faeces in Western countries being approximately 120 g (Cummings et al., 1992). A large proportion of this is microbial cell mass with bacteria comprising approximately 55% of faecal solids in persons living on Western-style diets (Stephen and Cummings, 1980). The large intestine is an open system in the sense that food residues from the small intestine enter at one end, and together with bacterial cell mass, are excreted at the other end. Because of this, the colon is often viewed as being a continuous culture system, although only the proximal bowel really exhibits characteristics of a continuous culture.

The large intestine is a complex microbial ecosystem in which bacteria exist in a multiplicity of microhabitats and metabolic niches. The microbiota comprises several hundred bacterial species, subspecies and biotypes. Microbial cell counts are generally in the region of 1011–1012 per gram of gut contents. Some organisms occur in higher numbers than others, but about 40 species make up approximately 99% of all readily culturable isolates (Finegold et al., 1983). Viable counting indicates that bacteria belonging to the genera Bacteroides, Bifidobacterium and Eubacterium, together with a variety of anaerobic Gram-positive rods and cocci predominate in the gut (Finegold et al., 1983), however, molecular methods of analysis indicate that other groups are also numerically important, including atopobium, faecalibacterium and clostridia belonging to the C. coccoides group (Harmsen et al., 2002).

Lyons (1992) describes sleeping sickness as a “classical disease of the savannah and transitional savannah around river systems where tsetse flies and people are forced into close contact by their shared need for water, especially during dry seasons. It has also been labeled a disease of frontier zones as it more generally occurs on the edges of human settlements where transition from the sylvan, or wild, ecosystem to the domesticated ecosystem of man is in progress.” One can thus easily view sleeping sickness as a ‘pastoral’ problem, which it is. It is certainly not an urban disease.

However, when most of us think of a pastoral setting, we probably conjure up a vision of rolling countrysides, green pastures, and a certain sort of tranquility, if you will. This is as true in sub-Saharan Africa as it is in Kansas, where I grew up as a boy. However, lurking in many of these peaceful African locales are tsetse flies and, as a result, anything but tranquility for humans and their domesticated ungulates. The problem is trypanosomiasis: more specifically, sleeping sickness for humans and nagana for the latter.

When we speak of trypanosomiasis, we are actually talking about a wide range of diseases in mammals caused by several species of Trypanosoma. Included among these diseases are surra, which affects horses, mules, and camels in India and North Africa.

GENERAL INTRODUCTION

Approximately 70% of the Earth's environment is marine, which includes substantial sediment deposits, some of which can be greater than 10 km in depth (Fowler, 1990). Although these sediments contain the largest global organic carbon reservoir (∼15 000 × 1018 g C, Hedges and Keil, 1995), apart from shallow margin sediments (to 200 m water depth), they have been considered to be relatively biogeochemically inactive. For example, Jørgensen (1983) calculated that margin sediments accounted for 83% of global marine sediment oxygen uptake whilst only representing 8.6% of global sediment area. In contrast, deeper sediments (200 to >4000 m water depths), despite being ∼91% of marine sediment area, accounted for only 17% of global oxygen uptake. The situation was considered even more extreme for rates of sulphate reduction, with this being responsible for, respectively, 50% and 0% of all organic matter being degraded in margin and deep water sediments (>4000 m water depths) (Jørgensen, 1983). This low activity was consistent with results demonstrating the limited depth distribution of prokaryotic populations in deep sediments. Morita and ZoBell (1955) concluded that the marine biosphere ended at 7.47 m deep, based on their inability to culture bacteria at this or greater depths. Reports of prokaryotes being isolated from deeper sediments were considered to be contaminants introduced during sampling, or dormant organisms being re-activated (ZoBell, 1938).

INTRODUCTION

Sulphate-reducing bacteria (SRB) derive energy for growth by coupling the oxidation of hydrogen or organic compounds to the reduction of sulphate to sulphide. The bioenergetics and the global topology of energy-conserving reactions have already been discussed in Chapter 1. Understanding the bioenergetics of the coupling of hydrogen oxidation and sulphate reduction is simple, in principle. Four H2 are oxidized by periplasmic hydrogenases and the eight protons and electrons are transferred to the cytoplasm through ATP synthase and transmembrane-electron-transfer complexes for sulphate reduction. This produces approximately three adenosine triphosphates (ATPs), of which two are needed to activate sulphate. Hence a net yield of one ATP is produced per sulphate reduced. Energy conservation by coupling the reduction of sulphate to the incomplete oxidation of lactate is more complex because the primary oxidation reactions are now also cytoplasmic. Because these yield two ATPs by substrate level phosphorylation, the same number as required for the activation of sulphate, a net energetic benefit can only be obtained by hydrogen cycling as proposed by Odom and Peck (Odom and Peck, 1981), cycling of formate or CO (Heidelberg et al., 2004; Voordouw, 2002) or by electrogenic proton translocation associated with the electron transport chain for reduction of sulphate. The components that participate in these anaerobic electron transport pathways will be considered in detail here. Harry Peck and Jean LeGall, the pioneers of the biochemistry of SRB, contributed greatly by purifying and characterizing many of the redox proteins present in these organisms.

Without question, the most speciose group of eukaryotic parasites are the coccidians. For me, the three most significant members are Toxoplasma gondii, Sarcocystis neurona, and Neospora caninum. One way or another, J. P. Dubey has had a direct hand, literally, in discovering and naming two of these organisms, in resolving one of the parasites' life cycle, and in developing a significantly deeper understanding regarding the biology of all three. Two of these three coccidians, namely T. gondii and N. caninum, infect more people, and cause more abortions in sheep and cattle, than any other protozoan parasite. The third, S. neurona, inflicts serious neurological damage to horses within the U.S.A. Because of the significant medical, veterinary, and economic importance of these parasites, I wanted to talk with J. P. and learn what I could about his interest in these organisms and the diseases they cause. Having known J. P. for probably thirty years, I thought I knew the answer, but I wanted to satisfy my curiosity and see if my guesswork was correct, so I headed for Beltsville, Maryland, in early May of 2006 to find out.

His ‘love affair’ for protozoan parasites, especially Toxoplasma gondii, goes back a long way.

According to the most recent estimates, there are almost a billion men, women, and children infected with hookworm. Among the most affected peoples in the world are the poorest populations in sub-Saharan Africa, the Americas, and Asia. For centuries, hookworm has been a major problem among the Chinese. According to Peter Hotez, between 1990 and 1992, parasitologists in various parts of China undertook what I consider to be one of the most intense surveys of the problem ever conducted. They collected more that 1.4 million stool samples from 2848 study sites in 726 counties in every province in the country. Of these, 17% passed eggs of either Necator americanus or Ancylostoma duodenale. This extrapolates to approximately 194 million cases of hookworm disease in just that country, and it does not include those who have had the disease and lost it for one reason or another. The enormity of the problem is exacerbated by Norman Stoll's estimate in 1962 that each day hookworms suck enough blood to cause the total exsanguination of 1.5 million people. The number of infections by hookworm has nearly doubled since the publication of Stoll's paper, so we can safely assume the amount of blood lost on a daily basis has, likewise, doubled!

Hookworm disease has been associated with humans since we changed from being hunter-gatherers to farmers.

The first time I saw live Trichinella spiralis was when I traveled to Chapel Hill, North Carolina, and began an NIH postdoctoral fellowship in the laboratory of Professor John E. Larsh. It was there that I observed probably Jim Hendricks or Norm Weatherly cervically dislocate a mouse, then skin and gut it. I watched as they took a pair of heavy scissors, and cut the mouse into small pieces before dumping everything into a 500-ml Erlenmeyer flask, containing a foul-smelling concoction of hydrochloric acid and pepsin. The flask was placed on a hot plate equipped with a magnetic stirrer, and allowed to stand for a few hours. The flask was then removed and the contents were poured through several layers of cheesecloth before gently centrifuging it and pouring off everything except the pellet at the bottom. A Pasteur pipette was then placed into the pellet and a small amount was removed to a microscope slide to which was added a cover slip. When I gazed through the microscope, I was astonished. There were hundreds of live, first-stage larvae of T. spiralis. It was an amazing site, one that I have not forgotten. As a student, I had seen these larvae in in situ-tissue sections stained with hematoxylin and eosin.

In 1898, Sir Patrick Manson announced to the world (actually to the British Medical Association at their annual meeting in Edinburgh) that Ronald Ross had successfully proven that Plasmodium sp. was transmitted by mosquitoes. In the same speech, Manson said, “in virtue of the new knowledge thus acquired, we shall be able to indicate a prophylaxis of a practical character, and one which may enable the European to live in climates now rendered deadly by this pest.” Of course, he was aware that there was already something available for the treatment of malaria, namely quinine, and that it had been available for a long, long time.

In the previous essay, I wrote about the discovery of ivermectin by Bill Campbell and his colleagues at Merck and the Kitasato Institute in Tokyo. In the case of ivermectin, these folks were looking for it, or at least were looking for something like it. In the process, they developed a plan and research protocol, conducted a vigilant and extensive search, made the discovery, defined the drug's structure, improved it structurally and functionally, tested it under highly controlled conditions, and then marketed it world wide. In other words, it was a carefully configured approach to discovery.

Quinine is a completely different story. In fact, I do not think it was really ‘discovered’, certainly not in the sense of ivermectin. Quinine just sort of appeared.

INTRODUCTION

The microbiota present in the sulphur cycle have been studied since the end of the nineteenth century when the pioneering work of the famous microbiologists Winogradsky and Beijerinck took place. Sulphur conversions involve the metabolism of several different specific groups of bacteria, e.g. sulphate-reducing bacteria (SRB), phototrophic sulphur bacteria and thiobacilli, specialized to use these sulphur compounds in their different redox states (Lens and Kuenen, 2001). Many of these microorganisms possess unique metabolic and ecophysiological features, and to date there are still regular reports of novel microorganisms with extraordinary properties. Several of the microbial conversions of the sulphur cycle can be implemented for pollution control (Table 13.1). This chapter overviews the applications in environmental technology, which utilize the metabolism of SRB as the key process.

Technological utilization of SRB sounds at first somewhat controversial, as sulphate reduction has been considered unwanted for many years in anaerobic wastewater treatment (Hulshoff Pol et al., 1998). Emphasis of the research in the 1970s–1980s was mainly on the prevention or minimalization of sulphate reduction during methanogenic wastewater treatment (Colleran et al., 1995). From the 1990s onwards, interest has grown in applying sulphate reduction for the treatment of specific wastestreams, e.g. inorganic sulphate-rich wastewaters such as acid mine drainage, metal polluted groundwater and flue-gas scrubbing waters. Nowadays, sulphur-cycle-based technologies are not solely considered as “end-of-pipe” applications, but their potential for pollution prevention as well as for sulphur, metal or water recovery and re-use are now fully recognized.

INTRODUCTION

An early focus on the use of molecular techniques to characterize natural populations of sulphate-reducing microorganisms (SRM) derived from the close relationship between their phylogenetic affiliation and their capability to anaerobically respire with sulphate. In other words, all so-far characterized SRM associate with lineages in the tree of life that predominantly consist of sulphate reducers. Known SRM are affiliated with two divisions (phyla) within the Archaea (the euryarchaeotal genus Archaeoglobus species and the crenarchaeotal genera Caldivirga and Thermocladium, affiliated with the Thermoproteales) and five divisions within the Bacteria (the Deltaproteobacteria, endospore-forming Desulfotomaculum, Desulfosporosinus, and Desulfosporomusa species within the Firmicutes division, Thermodesulfovibrio species within the Nitrospira division, and two divisions represented by Thermodesulfobacterium species and the recently isolated Thermodesulfobium narugense, the exact phylogenetic position of the latter is still ambiguous). Most described SRM are either Gram-positive bacteria with a low G+C content or Gram-negative Deltaproteobacteria. However, it is important to note that almost all major physiological properties of cultured and uncultured SRM, such as substrate usage patterns, the ability to completely oxidize a substrate to CO2, and alternative ways of anaerobic energy generation cannot be unambiguously determined from comparative analysis of their 16S rRNA genes.

The generally tight association between phylogenetic affiliation and sulphate-reducing physiology offered a foundation to directly associate the population structure determined by 16S rRNA sequence type and process. These studies have now been complemented by the use of highly conserved genes in the pathway for sulphate respiration.

INTRODUCTION

In the summer of 2003, I finished work on a book entitled Parasites, People and Places: Essays on Field Parasitology. My wife, Ann, and I were in our cabin in Green Mountain Falls, Colorado, and I was trying to tell her the story from the book that had to do with the discovery by William Walter Cort of the cause of swimmer's itch back in 1927. At the same time, she knew I was sort of lamenting the absence of a new project. She must have been impressed by my tale, because out of the blue, she said, “Why don't you write a book about discovery in parasitology?”

This started me thinking about the possibility of doing something along that line. Gradually, over the next several months, I put together an idea. Stories regarding the discovery of the transmission of malaria or sleeping sickness have been told many times over the years, so they are sort of ‘old hat’. But, then I thought, are they really?

I recalled the way I teach my own general parasitology course to undergraduates. I know that I mention Ronald Ross and David Bruce, among others, but I really do not get into much detail about how Ross and Bruce did their work regarding malaria or African sleeping sickness, respectively.

INTRODUCTION

In both natural environments and human-made systems, a phylogenetically diverse and heterogeneous group of anaerobic sulphate-reducing bacteria (SRB) (Barton, 1985; Odom and Singelton, 1993; Postgate, 1984) thrive as members of complex microbial communities, living on surfaces of particles, minerals and manufactured materials, i.e. within biofilms (Characklis and Marshall, 1990; Dar et al., 2005 and references therein).

The presence of biofilms often results in deterioration of colonized substrata. In the case of metallic materials, undesirable change in their properties resulting from material loss under biological influence is termed microbially-influenced corrosion (MIC) or biocorrosion. A number of reviews have been published describing fundamental and practical aspects of MIC (Geesey et al., 2000; Hamilton, 2000; Beech and Coutinho, 2003; Beech and Sunner, 2004).

The annual direct and derived costs of corrosion are estimated to be around 4% of the GNP of developed countries, of which 10–20% are related to biocorrosion (Geesey et al., 2000). In the oil and gas industry, biocorrosion accounts for 15–30% of the corrosion cases, resulting in financial losses in a range of 100 M $ per annum in the USA alone, excluding costs of lost revenues and often necessary remediation treatments. In some cases, for example in the Gulf of Guinea, extremely high rates of pitting-corrosion related to biocorrosion have reduced the life of oil subsea lines to one year (J.-L. Crolet, personal communication).