There have been many important recent developments in our knowledge of the breadth of prokaryote diversity, our understanding of the driving forces behind that diversity, and of its significance for our lives and for fundamental processes upon Earth. It has become clear that the microbes we know about are actually just the tip of a biological iceberg. In fact, the majority of microbes are unculturable on laboratory media at present. Much of our attention has been focused on pathogens, understanding their interaction with the host and how to prevent disease. However, there is a growing appreciation that without microbes fundamental ecological processes would not be balanced. For example, microbes in the ocean have a direct influence on the composition of the atmosphere we breathe.

A major advance in allowing us to understand the extent and nature of microbial diversity has been the development of genome sequencing. In parallel, there has been the development of tools to allow whole-genome comparisons. This has facilitated the study of microbial diversity and evolution, such as allowing the tracking of unculturable organisms, the study of organisms from extreme environments, and of medical and environmental bacteria and interactions between them. It has given us insights into the exchanges of genes between organisms, resulting in an understanding of the emergence of pathogens, a process which involves both gene acquisition and gene loss. Genomic comparison has helped to identify core genes, to the point where we can predict a minimal genome needed for life, which can be supplemented by the horizontal transfer of genomic islands, phenotypic innovation and catabolic pathway evolution.

INTRODUCTION

Even the simplest unicellular organisms on Earth display an amazing degree of complexity. The question is whether such complexity is a necessary attribute of cellular life or whether, instead, cellular life could also be possible with a much smaller number of molecular components, in the form of what has been called a minimal cell (Luisi et al., 2002; Islas et al., 2004). The first step to envisage such a minimal cell implies the identification of the necessary and sufficient features of life, leading to a clear definition of what ‘life’ is in this context. This is an extremely complex question with a long tradition in theoretical biology because, in addition to the understanding of the essential properties of a living system, the definition of cellular life is also related to the debated issue of the origin of life, since in the early stages of cellular evolution cells must have been close to the simplest possible life forms. Nowadays, there is a reasonable degree of consensus in defining life as the property of a system that displays simultaneously three features: homeostasis, self-reproduction and evolution (Luisi et al., 2002).

Life can be considered an emergent property: it is a quality that arises from the assembly of non-living elements, properly arranged in space and time (Luisi, 2002). Therefore, to understand life it is necessary to understand first the main non-living components.

INTRODUCTION

The human mouth is heavily colonized with bacteria. There are around 100 million bacteria in every millilitre of saliva, and it is estimated that around 800 bacterial species are present as part of the commensal microflora. Unlike the normal microflora at other body sites that live in harmony with the host, the oral microflora, at this stage of man's evolution, has to be controlled. If plaque accumulations are not removed by brushing or by other mechanical or chemical means, the gums become inflamed, a condition known as gingivitis. Around some teeth in some individuals, a more serious condition arises, known as periodontitis, which is an inflammatory condition leading to loss of attachment between the gums and teeth and destruction of the supporting structures of the teeth, which can eventually lead to their loss. Individuals with high sucrose intake in their diets are also at risk of dental caries where certain species, notably Streptococcus mutans and lactobacilli, ferment the sucrose to produce acids which demineralize the enamel layer. If left untreated, the bacteria invade first the dentine and then the pulp, rendering it non-vital, which in turn can lead to the formation of an abscess around the apex of the tooth. Such abscesses can spread via the tissue planes or the bloodstream to cause serious infections elsewhere in the body, such as abscesses of the liver or brain, which can be fatal.

INTRODUCTION

Evolution of catabolic pathways in bacteria is most often equivalent to ‘catabolic pathway expansion’ or ‘new acquisition of catabolic properties’, although in essence it could also mean loss or deletion. Even if we limit ourselves to this narrow interpretation, changes in the repertoire of catabolic functions of a bacterium are largely attributable to the activity of mobile genetic elements (van der Meer, 1997, 2002). Typically, mobile genetic elements may create new recombinations between previously disconnected DNA fragments, even from different bacterial origins, thus assembling bits and pieces together. Such recombinations are not necessarily needed to result in a single, smoothly transcribed new operon; any workable gene or fragment of genes within the boundaries of the new host cell may contribute to the catabolic expansion of functions (Dogra et al., 2004; Müller et al., 2004). Classical examples of evolutionarily ancient pathway expansions are the formation of the operons for toluene and xylene degradation in pseudomonads (Harayama et al., 1987; Harayama & Rekik, 1993; Greated et al., 2002). Observations of (most likely) very recent pathway expansions include the formation of pathways for chlorobenzene degradation, such as in Pseudomonas sp. strain P51 (van der Meer et al., 1991) and Ralstonia sp. strain JS705 (van der Meer et al., 1998; Müller et al., 2003), or pathways for 2,4-dichloro-phenoxyacetic acid degradation in Ralstonia eutropha (now Cupriavidus necator) JMP134 (Laemmli et al., 2000; Trefault et al., 2004).

INTRODUCTION

Soil is heterogeneous in nearly all respects and contains a huge diversity of micro-organisms. The availability of carbon and other energy sources, mineral nutrients and water varies considerably over space and time, as does temperature. Adaptations to nutrient poverty including oligotrophy and zymogeny (upsurge in growth when nutrients are available) are common. The water films essential for microbial life in soil are discontinuous, and only clay particles have the necessary charges to hold water against the pull of gravity. Clay-coated soil particles cluster together to form aggregates, and these aggregates or clusters of aggregates with their adjacent water form the microhabitats in which bacteria function (Stotzky, 1997). The result of the discrete microhabitats in soil is that microbial population dynamics and interactions are very different from those in well-mixed substrates such as some aquatic environments. Soil is also a reservoir for pesticides and other chemical and microbiological inputs from slurry application, all of which will have a selective impact on the indigenous bacteria.

Bacterial evolutionary histories are difficult to untangle. Different scales of evolution occur simultaneously, from events possible over a few generations (chromosomal rearrangement, gene deletion and acquisition of genes via horizontal transfer) to the eon-scale generative evolution which creates diversity from which the novel functional genes of the future will be selected. In the age of genomics we are developing the tools to study the ecology of microbes in soil.

CHLAMYDIAE

Chlamydiae are Gram-negative bacteria that are obligately intracellular parasites (Moulder et al., 1984). Chlamydia trachomatis and Chlamydia pneumoniae are human pathogens. C. trachomatis is a leading cause of preventable blindness in developing nations and sexually transmitted disease in the Western world (Moulder et al., 1984). C. pneumoniae is an aetiological agent of acute respiratory diseases (Grayston et al., 1998) and causes approximately 10% of pneumonia cases and 5% of bronchitis and sinusitis cases in adults (Grayston et al., 1993). Chlamydia psittaci is an avian pathogen. Humans can be infected with C. psittaci, through direct contact with an infected bird or by inhalation of dust contaminated with excreta of infected birds. This disease, known as psittacosis, is a severe pneumonia with systemic symptoms (Moulder et al., 1984; Everett et al., 1999). Other chlamydial species infect animals (Fukushi & Hirai, 1992; Everett et al., 1999). Although rare, humans may be infected through contact with lower mammals.

Chlamydial infection often becomes chronic and induces chronic inflammatory responses leading to tissue destruction, fibrosis and scarring and, thus, has been termed a disease of immunopathology (Grayston et al., 1985). For example, repeated/chronic ocular infection with C. trachomatis can result in blindness and genital tract infection can result in obstruction of the fallopian tube and infertility. Considerable recent attention has been focused on the association of C. pneumoniae with atherosclerosis, a disease of chronic inflammation and the leading cause of morbidity and mortality in the Western world.

INTRODUCTION

Biogeography, or the spatial distribution of biological diversity, has been studied since Darwin and Wallace in the 1800s. Their studies, and most later studies, concentrated on macroscopic organisms, mostly animals and plants, and many differences between species were observed, often correlated with geographical isolation. The theoretical basis for these differences was established later and is still being refined. The theories of island biogeography have been extremely influential in many fields of biology (Bell et al., 2005). Critical to biogeographical studies are comparable organisms from different locations with quantifiable diversity, often sequence diversity.

Microbial biogeography

More recently, micro-organisms have been studied (Finlay, 2002), especially with the advent of molecular tools. Studies using enrichment cultures indicated that identical micro-organisms were present wherever they were collected (Smith et al., 1991); however, this is clearly biased due to the relatively small number of micro-organisms that can be cultivated (Pace, 1997). The advent of small-subunit (SSU) rRNA gene sequence analysis indicated that ‘everything is everywhere’, particularly for spore-forming bacteria (Roberts & Cohan, 1995). It was unclear whether this indicated that there was so much dispersal of these spore-forming organisms that they were identical throughout the world or whether it was general for bacteria due to their extremely large population sizes. For the most part, however, only one gene, generally the SSU rRNA gene, was investigated. Extremophiles are thought to have more barriers to dispersal than mesophilic organisms.

INTRODUCTION

Introducing horizontal (or lateral) gene transfer (HGT) to first-year undergraduates used to be straightforward. A whistle-stop tour through the basic mechanisms of transformation, transduction and conjugation (with an honourable mention of homologous recombination) and descriptions of rough and smooth mutants of Streptococcus pneumoniae, the life cycles of phages lambda and T4 and the sexual proclivities of the ‘F’ factor was, until recently, little changed over the intervening years since their discovery. These processes (described in more detail below) also form the cornerstone of much of bacterial genetics, albeit from a predominantly Escherichia coli-centric perspective. This year celebrates the sixtieth anniversary of Joshua Lederburg and Edward Tatum's groundbreaking discovery of sexual transfer in bacteria (Lederburg & Tatum, 1946), yet to many the concept of HGT as a driving force in evolution has only come of age with the advent of comparative genomics (Kurland, 2000), which has revealed that far more is involved in HGT than plasmid-mediated conjugation. This review begins by providing an overview of the significance of HGT, defined here as ‘the acquisition and stable maintenance of foreign DNA into the genome of a recipient cell’ and introduces the key mobile genetic elements (MGEs) that are the agents of HGT. The roles of comparative genomics and bioinformatics and of the emerging field of metagenomics are discussed in relation to changes in perception of the significance of HGT in prokaryotic evolution, and the review concludes by highlighting key examples of recently recognized HGT-mediated phenotypes in the prokaryotic world.

INTRODUCTION

The extravagant diversity of microbes has only been fully appreciated with the development of comparative genomics. Comparisons of gene repertoires among prokaryotes have revealed striking differences among species and even among strains of the same species. For example, the genomes of three Escherichia coli strains have been shown to share only 40% of their genes, with most of the remaining genes being strain-specific (Welch et al., 2002). More generally, although most prokaryotic genomes contain thousands of genes, only a handful can be identified as truly ubiquitous in modern organisms. This so-called ‘core’ of universal genes has received much interest from evolutionary biologists because it probably represents a relic of the last universal common ancestor (LUCA) and provides valuable information for reconstructing the tree of life. It has also been viewed as the sine qua non condition of life, since no living organism seems able to survive without it. However, perhaps more interesting is the paucity of these ubiquitous genes, as it shows the formidable evolutionary plasticity of biological systems and points to the mechanisms necessary for acquiring and generating new genes.

WHAT'S IN A GENOME?

An inventory

All cellular organisms have in common the use of DNA as the support of genetic information, RNA as an intermediate of protein expression and the same genetic code (with only a few exceptions) as well as catabolism and metabolism based on a limited number of amino acids and sugars.

COMMON FEATURES OF GENOMIC ISLANDS

The genome of a bacterium consists of a core that is common to all strains of a taxon and an accessory part that varies within and among clones of a taxon. The accessory genome represents the flexible gene pool that frequently undergoes acquisition and loss of genetic information and hence plays an important role for the adaptive evolution of bacteria (Dobrindt et al., 2004). The flexible gene pool is made up of accessory elements such as bacteriophages, plasmids, IS elements, transposons, conjugative transposons, integrons and genomic islands (GEIs).

GEIs are chromosomal regions that are typically flanked by direct repeats and inserted at the 3′ end of a tRNA gene. They contain transposase or integrase genes that are required for chromosomal integration and excision and further mobility-related genes. GEIs are clone- or strain-specific and are never found in all clones of a taxon. Most GEIs are easily differentiated from the core genome by their atypical G+C contents and atypical oligonucleotide composition, with steep gradients thereof at their boundaries (Reva & Tümmler, 2005). First identified in pathogenic bacteria (‘pathogenicity islands’), GEIs have since been detected in numerous non-pathogenic species. GEIs may confer fitness traits, increase metabolic versatility or adaptability or promote bacterium–host interaction in terms of symbiosis, commensalism or virulence (Dobrindt et al., 2004).

GEIs have been found in the majority of all currently completely sequenced bacterial genomes, but there is a bias towards Gram-negative bacteria and life in microbial communities.

EVOLUTIONARY EMERGENCE OF DIVERSITY

The majority of phenotypic and ecological diversity on the planet has arisen during successive adaptive radiations, that is, periods in which a single lineage diverges rapidly to generate multiple niche-specialist types. Microbiologists tend not to think of bacteria as undergoing adaptive radiation, but there is no reason to exclude them from this general statement – in fact, rapid generation times and large population sizes suggest that bacteria may be particularly prone to bouts of rapid ecological diversification. Indeed, there is evidence from both experimental bacterial populations (Korona et al., 1994; Rainey & Travisano, 1998) and natural populations (Stahl et al., 2002). This being so, insight into the evolutionary emergence of diversity requires an understanding of the causes of adaptive radiation.

The causes of adaptive radiation are many and complex, but at a fundamental level there are just two: one genetic and the other ecological. Put simply, heritable phenotypic variation arises primarily by mutation, while selection working via various ecological processes shapes this variation into the patterns of phenotypic diversity evident in the world around us.

The ecological causes of adaptive radiation are embodied in theory that stems largely from Darwin's insights into the workings of evolutionary change (Darwin, 1890), but owes much to developments in the 1940s and 1950s attributable to Lack (1947), Dobzhansky (1951) and Simpson (1953). Recent work has seen a reformulation of the primary concepts (Schluter, 2000).

INTRODUCTION

Thomas D. Brock defined extreme environments, considering that there are environments with high species diversity and others with low species diversity. Those environments with low species diversity, in which whole taxonomic groups are missing, are called ‘extreme’ (Brock, 1979). It is not easy to find a definition that is completely acceptable for all environments that are considered as extreme, but we observe that in some habitats environmental conditions such as pH, temperature, pressure, nutrients or saline concentrations are extremely high or low and that only limited numbers of species (that may grow at high cell densities) are well adapted to those conditions.

Hypersaline environments are typical extreme habitats, in which the high salt concentration is not the only environmental factor that may limit their biodiversity; they have low oxygen concentrations, depending on the geographical area, high or low temperatures, and are sometimes very alkaline. Other factors that may influence their biodiversity are the pressure, low nutrient availability, solar radiation or the presence of heavy metals and other toxic compounds (Rodriguez-Valera, 1988). With a few exceptions, most inhabitants of these environments are micro-organisms that are called ‘halophiles’. However, different groups can be distinguished on the basis of their physiological responses to salt. Several classifications have been proposed; one that is very well accepted considers the optimum growth of the micro-organisms at different salt concentrations.

THE EXAMPLE OF YERSINIA PESTIS

Plague is a disease that has shaped the social, genetic and industrial makeup of many populations and especially those in Europe. The pandemic of disease which had the greatest impact on Europe occurred during the 14th to 16th centuries and is usually referred to as the ‘Black Death’ (Perry & Fetherston, 1997). During this pandemic it is estimated that 30% of the population of Europe died. The Justinian plague occurred in AD 541 to 544, originally in Egypt but then spreading through the Mediterranean and Middle East eventually to involve most of the known world (Perry & Fetherston, 1997). The third pandemic of plague originated in China in the 1850s but the impact of this pandemic was minimal in Europe. However, during the peak of the outbreak in India at the end of the 19th century, the disease killed a million individuals a year (Perry & Fetherston, 1997). Nowadays, the 2000–3000 cases of plague which are reported to the World Health Organization each year (Anonymous, 2000) are considered to be the vestigial remnants of the third plague pandemic.

The aetiological agent of plague is Yersinia pestis, one of three human-pathogenic Yersinia species. The human pathogenic Yersinia are closely related but cause very different diseases (Brubaker, 1991). Both Yersinia enterocolitica and Yersinia pseudotuberculosis cause relatively mild, self-limiting infections of the gastrointestinal tract, whereas Y. pestis causes an acute systemic infection which is often fatal.