Which of the following is not one of the most common prokaryotic cell shapes?

1. Prokaryotic Cell Structure and Functions

1Even after billions of years of evolution and adaptation to life in different niches of our planet there is only one basic type of cell for all prokaryotes. It differs markedly from the rich variety of cellular forms, sizes and structures found in eukaryotes. The prokaryotic cell is small, its volume and mass being approximately one thousand times less than those of an average eukaryotic cell (Fig. 1). However, it must not be considered merely a diminutive form of the eukaryotic cell but rather as strikingly different and remarkably adjusted to its peculiar and original way of life. Small size has major consequences and it dictates some important biological properties. For instance, the surface to volume ratio is much higher (about 100 times) in prokaryotic cells and this favors more rapid exchanges with the surrounding milieu. In the prokaryotic cell the cytoplasmic contents are more concentrated and in close and free contact with each other. Usually there are no internal compartments as we know them in the eukaryotic cell. High concentrations of ions and molecules combined to free movement and contact generally translate into surprisingly accelerated metabolism, rapid growth and multiplication. These have direct consequences on prokaryotes’ ecology, adaptability and evolution. A prokaryotic cell reproducing at the rate of two divisions per hour, which is by no means exceptional, will give a progeny exceeding one billion cells in less than two days of sustained exponential growth.

figure 1. Comparative size and shape of eukaryotic and prokaryotic cells. A. EUKARYOTES

A.-Examples from the large variety of eukaryotic cells (After 1.5 B.Y. of evolution)
1: Neuron
2: Blood cell
3: Paramecium
4: Animal cells in a tissue
5: Trypanosome
6: Amoeba
7: Yeast cell
8: Sperm cell
9: Dinoflagellate

B. PROKARYOTES

B.-The surprisingly standardized shape and size of the prokaryotic cells (After 3.5 B.Y. of evolution)

2Although unrestrained growth is not the rule in the prokaryotic world, such rapidly metabolizing and multiplying organisms can effect major physiochemical changes in an ecosystem in a short period of time. Also, if the hereditary patrimony of one of these cells is modified by a mutation or a genetic exchange and the new trait happens to be favorable, it will be propagated in a billion copies within a short period of time. Once acquired, a new trait can be spread very rapidly within a bacterial community and, later, in other niches of the biosphere.

3Usually prokaryotic cells are not compartmentalized and the high pressure exerted by the cytosol pushes outwards on the cells’ envelopes. It is generally contained by the outer membrane (the cell wall) which is rigid and resistant. As volume increases, so does the total internal pressure which eventually would physically destroy the cell (lysis). In part because of these physical constrains, during their evolution, the much larger eukaryotic cells have become compartmentalized. The partitioning of the eukaryotic cell’s interior by membranes also plays several other roles like separating different biochemical and physiological functions, providing larger surfaces for respiratory and/or photosynthetic activities, etc. Giant prokaryotic cells are rare exceptions but they are now known to exist (Prescott et al., 1996). Epulopiscium spp. (some are symbionts of fish), the largest known bacteria, are a hundred times the volume of Escherichia coli and they do not seem compartmentalized. It will be interesting to discover how this giant bacterium has solved the problems that are likely to be created by a large volume of cytosol to be contained by external envelopes only.

4In prokaryotes, the basis of cell function is cell structure (Rogers, 1983) as in eukaryotes, but in the latter, important functions are localized in discrete structures called organelles. For example, one of the fundamental distinguishing characteristics of an eukaryotic cell is the organization of its DNA in separate chromosomes contained in a membrane-enclosed bag, the nucleus. This feature does not exist in prokaryotic cells (Fig. 2). Moreover, the latter’s structure does not include an internal cytoskeleton nor true organelles (with the exception of the ribosomes) which characterize the eukaryotic cells. However, several different prokaryotic cells contain intracellular membranes or sac-like membrane components that can be more or less numerous or complicated in arrangement. Their functions vary from one kind of bacterium to another. For example, the cyanobacteria have thylakoids containing chlorophyll and capable of photosynthesis. All nitrogen-fixers have lamellae. For a discussion of inclusions and intracellular membranes, the reader is referred to the many excellent papers on the subject, for example, Jensen (1994). In the present chapter, we will discuss, in very general terms, the main structure of the prokaryotic cell, some of the molecular but mostly the structural aspects that differentiate the prokaryotic cellular body from its eukaryotic counterpart. Our goal here is not to present an exhaustive description of the prokaryotic cell’s anatomy but rather to point out the major differences between the two types of cell.

5The typical prokaryotic cell has the following basic structures: cell wall, also referred to as outer membrane, the cytoplasmic or inner membrane, a cytoplasm or cytosol with ribosomes and a central zone of aggregated, supercoiled circular DNA molecule often called nucleoid, or chromosome, that contains the stable hereditary information. We will hereafter refer to it as the large replicon. Also present are from one to seventeen (Fox, 1998) much smaller replicons (circular DNA molecules) which the cell harbours on a temporary basis (Fig. 2 and 3). Their copies are able to move from one strain to another (Fig. 4 and 5). These small replicons act as visiting genetic information molecules, easily exchangeable groups of genes, playing a major role in the solidarity of prokaryotes through a global communication System (Sonea, 1990).

6Prokaryotes occasionally contain inclusions consisting of reserve material, compounds of carbon, nitrogen, sulfur, or phosphorus. Many of them also possess one or several flagellar structures that enable them to move. The latter generally consist of a single coiled tube of protein and are fundamentally different from the eukaryotes’ undulipodium (Margulis, 1993). Bacterial flagella often move by rotation imparted from a basal body, a motor-like structure moved by protons. Other protruding filamentous proteinic structures that are not essential in all conditions are fimbriae and pili. Fimbriae seem to be involved in the sticking (adherence) of the cells that produce them to inert surface or animal tissues (e.g. epithelial cells). Pili play a role in genetic exchanges, as specific receptors for certain bacteriophages (tailed or filamentous) and also in the “mating” process or transfer of self-transmissible (ST) plasmids (Fig.5). Flagella, fimbriae and pili cannot be visualized under the ordinary light microscope without a special treatment and are not produced under all conditions. It is very significant that all prokaryotic strains studied until now possess surface receptors for temperate bacteriophages, and that about half of the strains also have receptors for self-replicating plasmids (Fig. 4 and 5). The generalized presence of such receptors for exchangeable genetic information is additionnal proof of the importance of the genetic communication System in prokaryotes. Finally, many prokaryotic organisms secrete on their surface slimy materials often referred to as slime layers, capsules or, to use the more general term, glycocalyx. We will return to these in later pages.

a) The cell wall of prokaryotes

7The prokaryotes’ envelopes fulfill several functions (Dijkstra and Keck, 1996). They physically isolate the cytoplasm from the cell’s surroundings, protect it against detrimental environmental influences, allow the maintenance of a high internal osmotic pressure and supply a rigid casing responsible for the shape of the bacterial cell. As noted previously, in prokaryotes mechanical resistance is conferred by the outer layer of the envelope: the cell wall. Basically, only three major specific shapes are adopted by prokaryotic cells and this fact too lends support to the concept of a unified prokaryotic world. These shapes have been recognized ever since the earliest times of examination with the ordinary light microscope (Fig. 1).

8In the cell walls of most eubacteria the layer responsible for strength is a heteropolymer composed of sugar derivatives (N-acetylglucosamine and N-acetylmuramic acid) and a small group of aminoacids connected together to form a repeating structure or network, the peptidoglycan (P.G.). The cross-links in P.G. form a two-dimensional tension-resistant web. It is present in eubacteria and its sugar derivative, N-acetylmuramic acid, has never been found in the cell membranes of eukaryotes. A variety of different types of P.G. exist particularly at the level of the interbridges between the glycan chains, but the point we wish to stress is that its several forms constitute structural polymers clearly different from the cellulose and lignin of plants, the chitin of insects and the keratin of vertebrates.

9In Gram-positive bacteria a large proportion (as much as 80 per cent) of the wall may consist of P.G. to which are attached other kinds of constituents, the teichoic acids. In Gram-negative bacteria, only between 10 to 15 per cent of the wall is P.G., the remainder being composed of additional layers of lipids and lipopolysaccharides in which are inserted different proteins, in particular those serving as membrane-channels, and called porins. They are important in the diffusion of nutrients and other substances. Other proteins present in the envelopes and whose presence and concentrations depend on the milieu and the growing conditions play a role in the entry of DNA during gene exchange processes. Lipopolysaccharides play important roles in the physical interactions (e.g. adherence) of Gram-negative bacteria with epithelial surfaces of animal hosts (Jacques, 1996).

10The synthesis of the P.G. is complex, but of great interest and practical importance because the molecule is exclusive to Eubacteria. It is beyond the scope of this book to review the biochemistry of peptidoglycan formation but it is important to recall that the assembly of its basic constituent molecules synthesized in the cell and organized in a network outside the cytoplasm requires several different biochemical reactions related to transport (through the cytoplasmic membrane) and insertion into the existing web. Some of the steps involved in the transport of these basic units and in the cross-linking between adjacent glycan chains can be inhibited by several antibiotics, of which penicillins (beta-lactams) and cephalosporins are of particular chemotherapeutic importance. Also, the P.G. molecule can be cut into smaller fractions and rendered fragile or lysed by natural enzymes called lysozymes secreted in our lacrymal fluids (tears), saliva, stomach and prostate fluids. These enzymes are useful, non-specific protective agents against bacterial infections and are also used extensively in the laboratory in experimental transformation and the production of cell-wall free bacteria (protoplasts and spheroplasts). Under natural conditions a bacterium that cannot keep its P.G. intact and synthesize it during active growth and division will not resist long internal cytoplasmic pressure and will burst and die. Antibiotics that interfere with the synthesis of the P.G. are not toxic for the eukaryotic cell which has different surface components and therefore these drugs are quite useful in controlling many infections. In addition to penicillins and cephalosporins, a group of substances called the glycopeptide antibiotics (e.g. vancomycin) inhibit P.G. synthesis by forming complexes with P.G. precursors and preventing their incorporation into the wall. Acquisition of resistance to vancomycin results from the transfer of resistance genes able to move from one strain to another by common gene exchange processes (transfection, mobilization and transduction) so that the ability to produce protective protein and resist the lethal effects of glycopeptide antibiotics can easily disseminate from one strain to another (Arthur et al., 1996). The increased use of vancomycin in humans and of avoparcin as a growth promoter for farm animals is certainly a cause for justified concern since it could end up in an increased transfer of resistance genes to a large number of different pathogenic bacterial strains.

11Not all prokaryotes contain muramic acid. Archaea and chlamydia do not have it. However, their walls may contain a heteropolymer somewhat similar to peptidoglycan that lacks muramic acid and is called a pseudopeptidoglycan or, in the case of certain Archaea, be made of special proteins. Many Archaea are found in extreme conditions of temperature or salinity and it is not surprising that they have evolved surface layers different from those of the eubacteria.

12Some cell wall constituents found in bacteria, but not in eukaryotic cells, can trigger a strong defense reaction in the human and animal body; they are powerful antigens and immunostimulatory agents. For example, bacterial lipopoly-saccharides (LPS, also called endotoxins, are well known to activate macrophages in animals and to generate immunoregulatory substances (e.g. tumor necrosis factor, interleukin-1, etc.) involved in the body’s defense and reactions to infections by Gram-negative bacteria. Their role in the triggering and the stimulation of the immune response receives considerable attention and is investigated in many laboratories. They are now considered as modulins or modulators of many of the immune reactions and as mediators of homeostasis and, in large concentrations, of tissue pathology (Henderson et al, 1996). This is also true of peptidoglycan which should not be viewed merely as a biologically inert corset that determines cell shape. Fragments of PG are potent biological effectors which modulate a remarkably diverse set of inflammatory and immune reactions in animals.

b) The bacterial plasmic (inner) membrane is a multi-purpose organ

13The prokaiyotic cell is essentially small, and space saving in its interior is crucial. A currently accepted model for the structure of the bacterial membrane is the fluid mosaic model of Singer and Nicholson. Although it appears simple, the membrane fulfills several functions and can be considered a multipurpose organ. There are no mitochondria in a prokaryotic cell and the role of this eukaryotic energy generator (respiration and electron transport) is assumed by the membrane. It also acts as a boundary layer for the cytosol and as such it pumps metabolites in and catabolites out of the cell and probably plays an essential role in the equal and symmetric genome distribution before separation of the mother cell into two identical daughters. The phospholipid leaflets are arranged in a bilayer structure into which various and numerous proteins are inserted. They play different roles, some of which are enumerated above (electron transport, active transport of metabolites, catabolites, DNA, etc.). Bacterial membranes differ, on the one hand, from the eukaryotic cell membrane and they may also differ among strains both in the occurrence and nature of their basic constituents, the fatty acids. Because cellular fatty acids can be separated and analyzed very rapidly by gas-liquid chromatography, this method can be used to differentiate bacterial strains among themselves (Böttger, 1996) and to show differences with eukaryotes. Another way in which bacterial membranes usually differ from eukaryotic ones is in lacking sterols such as cholesterol. However, many prokaryotic membranes contain sterol-like molecules that are synthesized from the same precursors as steroids. They are called hopanoids and they probably stabilize the two fatty acid sheets. Hopanoids can be isolated from kerosene, an organic precursor of petroleum. It is present in large quantities in some sediments. In fact, it has been estimated that the total mass of hopanoids in all sediments may be around 1011-12 tons, about as much as the total mass of organic carbon in all living organisms (1012 tons) (Prescott et al, 1996). Since in all probability these hopanoids are of prokaryotic origin, this shows the enormous importance of prokaryotes in the formation of fossil fuel, in particular petroleum.

14Let us recall that one important molecular feature that distinguishes the Archaea from the Eubacteria can be found in their membrane which contains aliphatic chains of phytanol instead of the usual fatty acids, and that the bond between phytanol and glycerol is an ether instead of an ester link (Prescott et al., 1996). However, horizontal genetic exchanges are possible despite these differences.

c) Prokaryotic ribosomes differ from those of eukaryotes

15Under the electron microscope a prokaryotic cytoplasm treated with the appropriate reagents shows aggregates and short linear arrays of black dots, the ribosomes. Some may appear randomly distributed, others equally spaced. Ribosomes in both prokaryotes and eukaryotes are an essential part of the translation (in the synthesis of proteins) machinery in cells. They act as the physical support on which the genetic information already transferred from the DNA to the messenger RNA is used to assemble amino acids in the correct order to make functional proteins. In bacteria, ribosomes are smaller than in eukaryotic cells. They sediment less easily in the ultracentrifuge. They consist of about two-thirds ribonucleic acid and one-third protein and, following proper Chemical treatment, they dissociate into two subunits of different size and weight. To fulfill their roles in protein biosynthesis, ribosomal RNA molecules must contain several functionally different regions. The nucleotide sequences in some of these regions are conserved and in others are highly varied. The smaller subunit of the prokaryotic ribosome contains a sequence (16S approximately 1500 nucleotides) that is particularly suitable for genetic and phylogenetic comparison between different bacterial strains and also for studies of evolution. During the very long geologic eons, when bacteria evolved into what taxonomists traditionnally call groups, families, genera or species, changes were imprinted in the sequence of ribosomal RNAs. These imprints or molecular signatures can be used to identify different bacteria and also to assess the most probable evolutionary distance between them (Woese, 1987).

16Since prokaryotic ribosomes differ substantially from those in eukaryotes, many important chemotherapeutic agents have been developed which are useful against pathogenic bacteria, being much more active, at their level, against them than against our own cells. Chloramphenicol, streptomycin, erythromycin and tetracyclines, for example, act as much more potent protein synthesis inhibitors in bacteria than in eukaryotic cells and are clinically useful antibacterial drugs (although they are not entirely devoid of toxicity in eukaryotes).

d) Capsules and glycocalyces

17Some bacteria are surrounded by what appears to be a well-defined halo of material with a smooth surface. Laboratory techniques can be used to stain the bodies of the bacterial cells with a cationic dye (e.g. methylene blue) and to suspend them in liquid with particles that do not penetrate the material of the halo (e.g. India ink). It can be visualized as a defined area surrounding blue dots (bacterial bodies). These transparent zones are usually called capsules and our knowledge of their chemistry is now extensive (Bayer and Bayer, 1994). A large number of polysaccharides have been isolated from capsules and the slime material of bacteria. Size, charge and composition of the capsular material are of primary importance in determining the roles and the usefulness of this structure for the bacteria. It seems that capsular material can be involved in giving some pathogenic bacteria a certain protection against phagocytosis by mononuclear cells of the animal body, and also in sticking (adherence) to the surfaces of the environment. Capsule-producing bacteria such as Streptococcus pneumoniae and Klebsiella pneumoniae are more resistant to phagocytic white blood cells and can invade a tissue or an organ more rapidly than those deprived of capsules. The immune response to polysaccharidic capsular material is often less pronounced and also affords shorter protection than proteinic substances of bacterial origin.

18Glycocalyces are generally less organized surface structures than capsules. They have a very high water content and they too seem to facilitate the attachment of the bacterial cells to solid surfaces. The sticky polysaccharide of the glycocalyx allows some oral bacteria to attach to tooth enamel and plays a role in the formation of dental plaque, the initial phase of tooth decay. Polysaccharides excreted by other types of bacteria (e.g. Xanthomonas, Pseudomonas) find many applications as gelifying agents in shampoos, seasonings, lubricating agents, etc.

e) DNA in the prokaryotic cell

19It is important to recall that prokaryotic cells have the totality of their genes distributed unequally: in most instances, about 99% in the large replicon carrying the genes essential for the survival and reproduction of the host cell, and about 1% in circular, self-replicating molecules, the non-essential, temporary visitors, the small replicons. As noted previously, there is at least one small replicon in each prokaryotic cell and, in some cases, up to seventeen (Fox, 1998a). Recent information obtained from the sequencing of different bacteria’s genomes indicates that in some cases (e.g. Borrelia burgdorferi) a much larger proportion of the hereditary information may be found among the plasmids than was thought initially (Fox, 1998b). Also, it is now known that a few types of bacteria have a different distribution of their genes: two circular large replicons or even a linear one (Jumas-Bilak et al., 1998). Such exceptional arrangements are, however, still much closer to the prokaryotes’ norm and entirely different from the arrangement observed in eukaryotes. The similarity of the general genetic organization (one large replicon and a variable stock of small replicons-associated genes) across the entire prokaryotic super-kingdom after 3.5 B.Y. of existence is striking. It lends further support to our view that the prokaryotic world is organized as a global superorganism whose constituent cells communicate and cooperate through easy and frequent genetic exchanges, with similar mechanisms and structures. Contrary to what we observe in eukaryotes, the free circulation of genes in prokaryotes is not compatible with the notion of species.

f) Prokaryotic spores are a survival form of extraordinary resistance

20In some bacteria (e.g. Bacillus and Clostridium) and mostly under adverse conditions, specialized and extremely resistant cells are formed. They are called spores or endospores, have no metabolic activity of their own (are fully dormant), but can be converted into vegetative (growing and multiplying) cells if favorable conditions prevail. This process is called germination. Various enzymes in spores are much more stable, particularly in relation to heat, than the corresponding enzymes in vegetative cells. The resistance of these prokaryotic endospores to elevated temperatures, to chemical substances and to radiations is much higher than that of any eukaryotic cell. It appears that this striking molecular stability depends more on the internal environment of the sporulated cell than on intrinsic structural or other peculiar properties. Spores contain no more than 20% water and have a high content of calcium dipicolinate, a molecule almost unique to them and which may constitute as much as 10% of their weight. Calcium dipicolinate stabilizes the DNA helices of the spores. Dehydration, calcium dipicolinate and an impervious, bilayered protein (keratin-like) coat all appear to play major roles in protecting the prokaryotic spore.

figure 2. The distinct organization of the intracellular genome in practicaily all prokaryotic cells

E: Equator
LR: The large replicon, also called genophore, nucleoid, and, erroneously, chromosome. It represents the large majority of the cellular DNA, containing the essential and stable genes of prokaryotic cells.
SR: Small replicon (plasmids and prophages). Very small, self-replicating DNA molecules kept ususally as long as the few (converting) genes they carry are useful to their host cell. Easily disposable and replaceable by other SR carrying temporarily better genetic supplements. There is at least one SR per prokaryotic cell but their number may reach seventeen. SR are visiting genes, each one able to multiply in many different prokaryotic strains.
iSR: Inserted small replicon which is not directly attached to the inner membrane of a host cell but inserted in the lenght of the large replicon (LR) of this cell.

21Schematic representation of a transversal section of a prokaryotic cell, at the level of the “equator”, an imaginary line in the inner membrane on which are attached all the cells’ circular DNA molecules and which is the point of origin of its division into two identical daughter cells (scissiparity).

22In a favorable environment germination of such spores leads to a gradual resumption of vegetative growth and the cells recover their typical morphology and content Dormancy of the spores has the same biological function as in plant seeds. It can last almost forever as demonstrated by spores kept in glass containers sealed about 350 years ago, and even more strikingly by Bacillus spores isolated from the gut of extinct bees, fossilized in amber, which apparently have been reactivated after several million years (Cano, 1994; 1995).

2. Dominant role of generalized solidarity in the life of prokaryotes

a) Mutually-supportive associations among different prokaryotic cells in local communities

23The large majority of prokaryotic cells live in mixed local communities, a fundamental feature of most prokaryotes. These communities are opportunistically rearranged and reorganized when necessary. Each type of cell is highly specialized, nonetheless it can fit well with others in metabolically complementary ensembles similar to the ones of a multicellular eukaryotic organism (animal or plant). However, they have the ability to modify or replace constituent cells if needed. These cells practice an efficient sharing of their biochemical activities: exchange of partially modified substances, of enzymes, cross-feeding with metabolic end-products, etc. Such teams form giant ensembles in all fertile soils, ocean floors, the digestive tracts of all animals, etc. Analysis of the heterogeneity of DNA extracted (rom natural environments indicates that there may be, for example, high numbers of different microbial “species” per gram of soil. Abundant populations can also be found at the surface of naturel waters.

24When confronted with challenges such as major changes in the chemical composition of their immediate surroundings, humidity, temperature, the accumulation of toxic or inhibitory substances, prokaryotic communities are also easily modified by borrowing temporarily suitable cells from the neighborhood’s (or planet’s) common stock. Sometimes these cells originate from far-away communities. Since each strain’s contribution represents a small fraction of the immensely rich and varied bio-energetic activities of the entire prokaryotic world, it can be easily interchanged with other types of teams without major disturbances to the latter’s biochemical adaptation and functioning. As is the case for artistic mosaics, the smaller and more varied the stones, the easier it is to obtain a good result; in prokaryotic mixed communities, the cells fall rapidly in their respective, appropriate place. In some cases it is amazing how rapidly new, well-balanced and successful communities of prokaryotic strains become established and persist practically unchanged for long periods of time. In the sterile digestive tract of any newborn animal a prokaryotic community will form, sometimes within a few days, and maintain a surprising resilience during the entire life-time of its host.

25To cope with the challenges of its new environment, a prokaryotic strain that will fit well in a new community will rely on its ability to adjust to a permanent selective pressure arising from constructive competition among all cells. In such communities, different strains do not kill their local competitors but they eventually outbreed the less efficient and poorly adapted ones. Nevertheless, biological solidarity and interdependance among the stable strains in these communities is enormous. In some cases less than one per cent of the strains from well established mixed groups can be grown when artificially isolated and fed in the laboratory. This shows that team partners offer each other support and assistance in the form of substances that we either fail to identify as growth or multiplication requirements or to supply in proper concentrations in laboratory culture media. There are unsuspected similarities between the ways of functioning of our blood cells and those of prokaryotic cells. Both belong to their own multicellular organism and proceed from a common original cell (egg for animals or first viable cell on earth for prokaryotes), they move freely in their liquid or viscous surroundings, have areas of high biochemical specialization and provide services and help (e.g lymphokines) to their mates which in exchange support them. They too are so adapted and specialized for life in a multicellular organism that most of them cannot be easily cultivated as isolated cells in vitro.

26Most mixed prokaryotic communities are able to break down complex molecules and intermediary products to the point where the final, unused waste is in fact a useful, sometimes essential ecological residue, often salts and other inorganic substances necessary for plants and algae. Interestingly, some bacterial teams create their own microclimate adjusted to their need. Sometimes, even the temperature is locally maintained at an optimal level. The compounded effects of prokaryotic communities play a major role in stabilizing the atmospheric nitrogen concentration, the acidity and alkalinity of their environment, etc. This ensures favorable conditions and homeostasis for the entire planet. The significance and importance of prokaryotes as essential components and stabilizers of the biosphere cannot be overemphasized. The totality of their local mixed communities, acting as an ensemble, a single global superorganism, represents the most active and decisive element in the maintenance of planetary homeostasis. Soil fertility and its conservation depend mostly on large, mixed prokaryotic communities, often working in unison with fungi. This activity is one of the most evident confirmations of the Gaia hypothesis (Lovelock and Margulis, 1974) which presents the entire biosphere as a life-supporting entity, modified and maintained in a form most favorable to life, just as an animal maintains a stable internal environment favorable to all its cells: homeostasis. The latter has been created for our biosphere and improved by prokaryotes alone over the first two billion years of life on our planet. As he gradually acquired a better understanding of the prokaryotes, man has recently begun to copy and exploit this potential of prokaryotic communities in sewage treatment, sludge digestion processes and oil spill clean-ups, etc.

27The prokaryotic ability to engage in a variety of associations is not limited to activities within the prokaryotic world but also extends to symbioses with eukaryotes, their estranged offspring. Mycorrhizae in particular and other types of fungi are often participants in favorable associations with bacteria. Together, by their complex common activities, they favor the metabolic exchanges and growth of the associated plants. Many prokaryotic mixed communities contain in their surroundings a variety of eukaryotic species. The simplest example are the protoctists (Margulis, 1993) represented by unicellular organisms or by only slightly differentiated multicellular entities. Often they carry intracellular prokaryotic endosymbionts, and more often ectosymbionts on their surface. The much more complex animals are always associated with prokaryotic endo or ectosymbionts or commensals. For many hosts prokaryotes are essential for survival (Margulis and Fester, 1991). We carry in our own cells descendants of Precambrian bacteria as tiny power plants: mitochondria, and photosynthetic eukaryotic cells carry bacteria (or former bacteria, e.g. cyanobacteria) as symbionts (see Monastersky and Mazzatenta’s The Rise of Life on Earth, 1998). Many prokaryotic cells which became endosymbionts seem incapable of independent life and behave as organelles of the eukaryotic host cell.

28As already stated, the parasitic or pathogenic prokaryotes represent only a particularly small fraction of the giant prokaryotic half of the living world. By contrast, there are several important examples of associations between communities of interdependent bacterial types and higher organisms which are absolutely essential to the survival of both under natural conditions (Margulis and Fester, 1991). One such association that has been and still is, as already stated, intensively studied is the ruminant-bacterial flora ecosystem (Hungate, 1966, 1975; Baldwin, 1984).

Ruminants and the rumen ecosystem

29The bodies of wild and domestic mammals as well as those of humans do not produce the enzymes that can digest insoluble polysacharides, of which cellulose is quantatively the most important, and which constitutes the bulk of organic matter in terrestrial plants. It is now well established that grazing animals like ruminants must resort to microorganisms as digestive agents to subsist on grasses and leafy plants, an enormous source of renewable nutrients. Ruminants are herbivorous animals in whom a special organ, the rumen, has evolved. It accomodates, physically and physiologically, the billions of bacteria (and, in smaller numbers, protozoa) that break down the insoluble polysaccharide of plants. Hydrolysis of cellulose in particular, starch and other substances of plant origin occurs in the rumen through the concerted activities of those specialized, polymer-degrading, extensive (1010 -1011 cells/g of dried rumen contents) microbial populations. They supply the host with essential nutrients and energy derived from plant carbohydrates that would otherwise be unavailable to it. Under natural conditions, ruminants obtain most of their essential nutritive and growth factors as a result of the microbial activity in the rumen (Hobson, 1976). After regurgitation, the massive microbial flora enters a true stomach (abomasum) and the small intestine, where it serves as a nutrient. Thus, in many regions of the world, the human food supply depends to a large extent on baeterial cellulose decomposers and sugar fermenters that have adapted to life in symbiosis with the ruminants.

Dynamics of the rumen ecosystem as a prokaryotic model of association or ectosymbiosis

30The interactions between the various microbial populations of the rumen have been studied extensively. Recent progress in molecular biology has also improved our knowledge of the diversity of ruminal strains and lead to necessary renaming and reclassification. It is certain that the number of different strains that are normal inhabitants of the rumen greatly exceeds (Krause and Russell, 1996) the figure (between 20 and 30) that was the accepted estimate some decades ago. The extent of interdependence and collaboration in this ecosystem is amazing. It also illustrates how successful and efficient different strains, practicing labor and resource sharing, can be. Depending on the respective amounts of cellulose, starch and pectin in the ruminant’s diet, the proportions of cellulose (e.g. Bacteroides succinogenes, Ruminococcus albus), starch (e.g. Bacteroides spp., Selenomonas, Streptococcus spp.) and pectin (e.g. Lachnospira spp.) decomposers fluctuate but the end-result is qualitatively the same: fermentation products, mostly as short chain fatty acids, are made available for the host and for the rumen flora itself. In this very complex community, some strains depend for their growth and multiplication on digestion by-products derived from other strains’ catabolic activities. For example, a rumen inhabitant, Selenomonas ruminantium, which is non-cellulolytic, grows well in vitro on a cellulose substrate, providing it is associated with Bacteroides succinogenes, whereas on its own it will not grow. This is but one example that illustrates the complexity and the collaborative nature of the interrelations within the rumen bacterial community. Constancy of the rumen ecosystem is essential for the good health of the host animal. Under ordinary circumstances it is maintained within narrow limits. The equilibrium between the members of the microscopic flora on the one hand, and the two symbiosis partners on the other, can be broken and this generally results in severe illness as it happens when the animals are switched abruptly from a diet composed predominantly of forage to one that is rich in grain. Can ruminal bacteria exchange genetic material? It is now certain that at least some can, that some harbor broad host range conjugative transposons and that bacteriophages (therefore also prophages) are present in rumen liquid (Morrison, 1996). Three classical methods of DNA transfer: transformation, conjugation, and transduction have been demonstrated under laboratory conditions, although with a limited range of ruminal bacteria (Morrison, 1996). In addition to its complex and essential bacterial flora, the rumen contains a characteristic protozoal fauna that has been estimated in the neighbourhood of 106 cells/ml (Brock et al., 1994). These protozoal cells can hydrolyse plant carbohydrates as well. The rumen is an anaerobic (lack of oxygen) environment. Like the bacteria that grow and multiply in the rumen, the protozoa found in this organ are necessarily anaerobes, that is they live and multiply well only in the absence of oxygen, a property that is exceptional among eukaryotes. Rumen protozoa also play a useful role in controlling the densities of local bacterial populations by feeding upon them. Thus they help balance the dynamics of the ecosystem. Bacterial associations similar to that found in cows are present in sheep, goats, camels and several undomesticated species such as buffalo, deer, elk, etc. Bacterial symbionts are not found only in the digestive tracts of higher animals. Termites, for instance, depend on bacteria to digest their cellulose-containing foods. Also, a considerable body of evidence indicates that many Bacillus spp. are normally present in the digestive cavities of bees and other insects. Bacillus megaterium, B. subtilis, B. pumulus, B. circulans and many others are normal and essential inhabitants of the guts of insects. It is interesting to recall, as already noted, that some of these symbiotic relationships seem to have existed many million years ago (Cano, 1994). Modem techniques of extraction, amplification and sequencing allow systematic studies to determine evolutionary relationships between ancient and present-day organisms.

b) Scope and importance of gene exchanges between different prokaryotic cells: A general way of solving problems and adapting to temporary changes

31In contrast with the norm among eukaryotic cells, hereditary molecules (DNA) of prokaryotes can be transferred from one strain to another of a different type. This gene exchange phenomenon happens on a regular, often predictable basis and at an optimal rate, at least for the bacterial strains already studied. After 3.5 B.Y. of evolution it appears profoundly significant that the mechanisms for transformation and for the active transfer of genes by plasmids and prophages are so similar across the entire spectrum of the different prokaryotes. They play a decisive role as agents of intermittent reshuffling or intermixing of a significant proportion of bacterial genes, particularly at crucial moments, when needed. At the global level, there is a generalized “pulsating” event having, in a different way and with higher frequency and impact, the same essential function as the transfer and mixing of genes that take place by sexuality, in successive generations of eukaryotes. An important point with prokaryotes is that a newly transmitted gene is generally expressed immediately in the cell which has accepted it. Because of its magnitude and efficiency, the exchange of genes in prokaryotes offers a rather rapid and useful means of communication and adaptation. On it are based many complex bacterial functions. The accumulated hereditary information patrimony of prokaryotes is available to other strains, if useful. It is a convincing manifestation of advanced general solidarity among prokaryotes, a benefit deriving from the existence of a common global genome. This again is strikingly different from the familiar eukaryotic genetic processes (the Mendelian ones) which imply sexuality, speciation, and rigorous and restrictive genetic isolation. This fundamental difference between these two superkingdoms has momentous consequences and should therefore be better known and stressed, not relegated to the rank of biological oddity as is sometimes done today.

32The prokaryotic world which has evolved over an estimated period of 3.5 B.Y., still reproduces itself and behaves as one giant entity, a complex System with cells separated but for which collaboration by DNA communication is a basic fact of life. Each known type of bacterium can by analogy be compared with a receiving and broadcasting station with the particularity that genes are used as coded messages or information molecules. All the bacterial strains which have been studied so far in the laboratory are dynamic chimeras. They can change some of their intra-cellular genes acccording to the circumstances. There is no similarity to the rigid genetics of eukaryotic species. Prokaryotes contain in their cells a certain number of genes whose main function is dissemination of hereditary information, i.e. making possible horizontal gene exchanges between themselves and other bacterial types or strains. These genes responsible for the generalized horizontal transfer of genetic information function in two different ways, corresponding to the two basic types of mechanisms involved: transformation and exchange of small replicons.

b1) Transformation

33In nature, genetic transformation is largely distributed among the best known prokaryotic families and is estimated to be present in about half of all existing strains. A large proportion of prokaryotic cells might therefore benefit from it. It consists in the active uptake by a cell of free and soluble “foreign” DNA molecules from its immediate neighborhood and their incorporation into the receptor cell’s hereditary patrimony of genetic information. The genes responsible for the proper functioning of this widespread mechanism are organized in a competence operon (Dreiselkelmann, 1994; Lorenz and Wackemagel, 1994) that encodes about a dozen proteins or “factors”, some of which act as DNA receptor sites at the surface of the cell and actively attract free DNA fragments originating usually from neighbouring dead bacteria. Other guiding proteins associate with these DNA molecules and help them cross the cell wall of the recipient prokaryotic cell, a barrier which normally is not permeable to DNA molecules. A significant fact is that these genes responsible for the mechanism of transformation are normally repressed when a prokaryotic strain benefits from good environmental conditions, growing and multiplying at a satisfactory rate. Prokaryotes not in need of new genes are unable to freely take in soluble fragments of DNA present in their surroundings. They are called “incompetent” cells. Thus in most naturally transformable strains, the ability to take up DNA (competence) is a transient inducible physiological property (Lorenz and Wackernagel, 1994.). If such strains become no longer able to multiply, for instance when the medium is inadequate, nutritionally or otherwise, the “competence” genes are derepressed and they temporarily encode the synthesis of the proteins and other factors involved in the transformation phenomenon. As a consequence, if among the DNA fragments present in the milieu there are copies of a gene able to correct the situation of the bacteria involved, they are likely to be brought in and encode the needed substance. If the acquisition of the new trait results in the growth and multiplication of the cells that have realized this successful transformation, the competence genes will again be repressed. Transformation, available when needed, can be viewed as an economical strategy for the bacteria concerned, since as they resume growth they return to usual functions and become again “incompetent”. Transformation has a long evolutionary history. It is amazing that after possibly some 2.5 B.Y. of evolution following its earliest manifestation, it remains a generalized phenomenon with much similarity among many types of prokaryotes. It has indeed been observed among widely different taxonomic and trophic groups, including Archaea (Grogan, 1996), with its main features preserved. It appears as a fundamental process essential to the genetic solidarity of the entire prokaryotic world.

34Although a successful gene exchange process, transformation presents important limitations since it needs the presence of extracellular “foreign” DNA molecules in the immediate vicinity of a competent strain where DNases are likely to be found as well. Since DNases split DNA into inactive fragments, there is only a short period of time during which naked molecules acting as carriers of hereditary messages can remain intact. For transformation to succeed and be of some significance in prokaryotic life, a minimal pool of soluble DNA molecules must be maintained among the cells of active bacterial populations. The fact that free DNA was considered unstable in soil and water led many microbiologists to believe that transformation would not occur in external environments. However, recent work by several groups (cited by Miller, 1998) show clearly that in many environments (soil, marine sediments, rivers, slime layers on river stones), free DNA can become stable by associating with soil components, for instance, and be taken up by competent cells. We will mention later some of the specific biological ways by which concentrations of soluble DNA molecules in the midst of prokaryotic communities can be kept high enough to make transformation a viable and efficient process. Another limitation to transformation derives from the presence in the surrounding DNA of a predominance of genes originating from the large replicons (“chromosomes”). They represent the vast majority of genes that are candidates for transformation but such fragments cannot replicate on their own. They have to become inserted in the body of the large replicon of the receiving cell to benefit from the activity of the latter’s replicator genes. Their insertion becomes facilitated if, in the recipient cell’s large replicon, they find a zone of homology, that is a sequence of genes rather similar to their own. This, on the one hand, facilitates the base-pairing over the length of the newly arrived, useful gene but, on the other hand, it imposes a constraint which reduces the probability of successful acceptance of entirely different genes. It limits the scope of transformations to minute improvements over short ranges (structural or metabolic), those which have zones of homology in both the donor and the recipient cells. This fact somewhat precludes important genetic innovations through the acquisition in a single event of major and quite distinct traits. However, the exact arrangement of the genes along the large replicon is not permanently set. When the transferred genes are part of a transposon, inside a fragment of the donor’s large replicon, the requirement for a region of homologous sequence in the recipient bacterium is no longer a necessity. Indeed, transposons, genes which are framed between two insertion sequences, may have their copies inserted in different parts of their replicon or in another replicon. Therefore they are usually accepted in the receptor cell’s large replicon without the need for a homology zone, hence the greater possibilities for important genetic innovations by transformation when transposons are involved.

35Conjugative transposons are DNA sequences that are normally integrated in the bacterial host’s genome and that can excise themselves. Following excision, a conjugative transposon can follow different courses:

1.- Reintegrate (elsewhere) the genome of the same cell,
or
2.- Transfer to a recipient cell and integrate into its genome.

36What makes conjugative transposons extremely significant for prokaryotic adaptation and evolution is that they can mobilize co-resident small replicons and effect their transfer with themselves, and some can even carry along with them unlinked DNA sequences from the large replicon (Salyers et al., 1995).

37Conjugative transposons generally have a broad host range, that is they can move readily across “genus” Unes. This ability to transfer themselves and other genes not only to closely related but also to genetically remote strains makes conjugative transposons important elements of communication, exchange and adaptation in bacteria. Their transfer also occurs in nature (Salyers et al., 1995). A rather intriguing observation is that in some bacteria (e.g. Bacteroides spp) certain antibiotics appear to act as stimulants for the conjugative transfer of transposons carrying genes of resistance to antibiotics.

38When the mechanisms involved in transformation operate the transfer of soluble DNA molecules forming small replicons, the latter, once accepted in the recipient cell, can immediately replicate on their own since they carry their replicator genes and they do so, usually, within the same time interval or at the same pace as the large replicon. No longer is there a risk of not finding a matching homology zone. The choice is not biased towards minor genetic change, important innovations are also possible. This kind of transformation based on the acceptance of small replicons by the mechanisms of competence is called transfection. The name was formed by combining the prefix from transformation with the suffix from infection, since for some time the small replicons have (wrongly) been considered to be infectious parasites of bacteria. Even today many biologists still consider the temperate phages merely as viruses, although they are typical self-transmissible small replicons.

39In bacterial cells, generalized transfer of genes by transformation can be viewed as a form of natural genetic engineering available to possibly half the existing strains in the world. By contrast there are no natural horizontal gene transfers between eukaryotes other than by sexuality which does not usually cross the species barrier. Learning from the observations made with bacteria, biologists have been able to realize artificial transformation in eukaryotes, even to perform controlled, selected gene changes with the resulting production of recombinant substances of great commercial and medical value. These developments in biotechnology and genetic engineering have been inspired, greatly helped and accelerated by the knowledge of basic gene exchange processes in prokaryotes.

b2) Small replicons

40Small replicons are normal constituents of prokaryotic cells, dwelling as visiting genes in different strains on a temporary basis. Their replicator genes allow them to be multiplied in different types of cells, without the need to be inserted in the latters’ large replicon. They always bring along, for the possible benefit of the host cell, a few potentially useful accessory genes as hereditary supplements (converting genes) and they are the main participants in the horizontal exchange of genes which may benefit prokaryotic cells. The importance of these essential elements of the prokaryotic world is underestimated by biologists including the majority of microbiologists (Wellington and Van Elsar, 1992). An important difference between eukaryotes and prokaryotes resides in the latter having access to and benefiting from a very active free market of all their genes, a market mostly maintained by small replicons. Hence, in contrast with eukaryotes in which the basic self-replicating sub-units are the cells, the prokaryotes also possess a supplement of much smaller autonomous basic entities, the small replicons. These have a profound influence on the dissemination and the timely use of all prokaryotic hereditary properties.

41Regardless of the mechanism by which a small replicon enters a prokaryotic host cell, it starts immediately replicating itself, usually simultaneously with the large replicon. Prokaryotic cells provide them with the facilities allowing the visiting small replicons to function as if they were their own genes. As far as we know, the number of small replicons varies from one to seventeen (Fox, 1998) in different strains. When a small replicon is no longer useful to its host, being disposable, it is eliminated (curing) and usually replaced by another temporarily more helpful one. The same strain may thus be successively endowed with several different biochemical abilities depending on the type of combination of its small replicons. Until recently, many of these differently modified strains were considered to be different species (Le Minor, 1968). Small replicons and prokaryotic strains are surprisingly polyvalent in their capacities to associate in countless opportunistic combinations, as observed in nature. This contributes to diversity and to adaptable variations, and constitutes a rich reserve of a large and readily available variety of genes in different habitats: soil, oceans, etc. There is a limit to the number of small replicons that a prokaryotic cell can harbor and the cell may become increasingly vulnerable as their number increases (Karska-Wysocki and Sonea, 1973; Sonea et al., 1974). The small replicons may be eliminated from their host cells by some chemical substances, for example the acridines (Damsker and Sonea, 1970; Dobardzic and Sonea, 1971). Small replicons that are very closely related and whose combined presence would not confer any additionnal advantage to the host cell will not generally be kept together in the host cell; one of them will be lost during subsequent cell replications. The giant bank of small replicons in nature is found concentrated where most types of prokaryotic cells are present: in soil, the ocean’s floor, at the protected surface of water expanses, etc. It never remains static because not only do small replicons have their own evolution, but even the same type may present itself with many variations by obtaining converting genes from previous hosts.

42Although small replicons show many similarities among themselves and seem to have a common ancestor, they differ in size, organization and capacities, depending mainly on whether or not they possess transfer genes which make them self-transmissible (ST). In the absence of such transfer genes they are non self-transmissible (NST).

43The non self-transmissible (NST) plasmids are the smallest, simplest and the most numerous types of small replicons. They are formed by two kinds of genes: replicator genes and converting genes (Fig. 3). The replicator genes allow them to replicate autonomously in different types of prokaryotic cells. Added to them are a few converting genes, all of them accessory but possibly useful to some of the visited strains. These genes were very probably picked-up by their NST plasmids on the occasion of visits to former host cells. This could have happened recently or centuries earlier. The NST plasmids do not have active mechanisms to send some of their own copies from one strain to another; they are nonetheless involved in numerous horizontal gene transfers by transfection, when their DNA is liberated by a dying cell close to one receptive and competent prokaryotic cell. Moreover, NST plasmids are often carried along by ST small replicons when these leave their common host cell for another one. The mechanisms involved are called mobilization when performed by ST plasmids and transduction when temperate bacteriophages (the transfer form of the prophages) act as carriers. The frequency of these horizontal exchanges for the genes of NST plasmids is remarkable. This collaboration between the two categories of small replicons added to transformation has raised the importance of gene exchanges in prokaryotes to the level of an efficient free market for genes (a global genome).

figure 3. Proportion of the different types of genes according to their general activities in the small replicons

CG: Converting genes
RG: Replicator genes
TO: Genes of transfer operons
NST: Non self-transmisible plasmid
ST: Self-transmissible plasmid
FP: Filamentous phage
P: Prophage

FIGURE 4. Symbols representing surface receptors for horizontal gene transfer by self-transmissible (ST) small replicons between different types of prokaryotic strains.

LR: large replicon
P: Prophage
SR: Small replicon
SRec: Surface receptors
STP: Self-transmissible plasmid
TPh: Temperate phage

44These receptors are represented here schematically. They accept the tip of the tubules (pili or tails of the phages) necessary for the active transfer of ST small replicons which together perform most of the horizontal transfer of genes between prokaryotic strains. Every strain possesses surface receptors for temperate phages (TPh) which carry prophages and sometimes other genes from the donor cell. Only about half of the strains have surface receptors for the tips of pili which participate in the active transfer of self-transmissible plasmids (STP) and, occasionnally, other genes from the donor cell.

45The self-transmissible (ST) small replicons possess, on the one hand, the same basic structure and functions as the NST plasmids (replicator genes and a variety of converting genes) but, on the other hand, these common features are complemented in the ST small replicons by transfer genes organized as operons (Fig. 3). When ST plasmids and the prophages, the two forms of ST small replicons, arrive in a new host, the first protein encoded by their transfer operon is the repressor which keeps the transfer operon’s genes inhibited. The repressor also prevents the multiplication of any other similar small replicon that would subsequently come into the same cell. This is called immunity for the prophages and exclusion of entry for the ST plasmids. When, for different reasons, the repressor is not rapidly synthesized and the transfer operon is consequently still derepressed when arriving in a new cell, it encodes the synthesis of copies of its ST small replicon independently of the usual division concomitant with that of the large replicon of the host cell. It also encodes very small proteinic tubes (only visible with EM) which will help the copies of the ST small replicons reach a receptive cell and penetrate its cell wall at a receptor site specific for the tip of that tube. Such receptors exist at the surface of all prokaryotic cells for receiving different types of ST small replicons (Fig. 4).

The ST plasmids

46The transfer operon of ST plasmids encode the elements needed for the physical contact between the donor and the recipient cells so that active horizontal transfer of genes may take place. This contact is directly realized by the tubular formation (pilus) while the transfer operon directs a single newly synthesized strand copied from a ST plasmid into the recipient cell. The new copy is guided by the pilus which originates in the donor cell and is attracted to one of the many surface receptors for pili presented by the receiving cell (Fig. 4 and 5). The donor cell, now equipped with one or two pili, functions as a syringe to inject the genes of the ST plasmid and those taken along from the donor cell. The latter process is called conjugation for genes from its large replicon, and mobilization when small replicons are also involved.

47The small replicons thus actively transferred eventually start to divide in the receptor cell. As already, noted they may be eliminated later (curing) if they become useless. The genes originating from the large replicon of the donor cell carried along on such an occasion will, in general, be accepted in the receiving cell, particularly if they find a homology zone on the large replicon of the new host (conjugation), as is also the case in transformation. Usually, only a few genes of the donor’s large replicon are thus transferred. However, under very particular experimental conditions, some ST plasmids may transfer the entire large replicon of the donor cell, a rare mechanism in nature, which has been called bacterial sexuality by François Jacob. The first small replicons that have been widely accepted by biologiste as carriers of useful genes between different types of prokaryotic cells have been the ST plasmids. Their involvement in the spreading of genes of resistance to antibiotics used against pathogenic strains which cause serious, contagious diseases was first shown decades ago and it has been confirmed in hundreds of unchallenged scientific publications afterwards. Soil bacteriologiste and ecologists later discovered similar types of ST plasmids which transferred resistance genes against different toxic substances from one bacterial strain to another. ST plasmids possess a few limitations: they are active mostly in Gram negative strains and they need a very close contact between donor and recipient cells; therefore they are much more successful in very concentrated populations of prokaryotic strains. These are found in animal digestive tracts, in soil, in rich aquatic sediments, at the surface of still water expanses, etc.

The prophages

48The prophages, the most important self-transmissible small replicons, are still the subject of much confusion since they continue to be considered as viruses by the majority of biologiste, including many microbiologiste. As a consequence, their decisive role in gene exchanges among prokaryotes is ignored or underestimated. Their contribution to bacteria’s easy adaptation is often overlooked and the fact that they can confer important properties to their host cell is barely mentioned. The misunderstood “destructive” replication in their host cells when the transfer operon is derepressed (induction) receives most of the attention. The ability of the prophages to carry beneficial hereditary changes is only briefly, if at all, mentioned although it probably plays the most important role in prokaryotic gene transfer.

49We have already pointed out similarities between ST plasmids and prophage, which suggest a common ancestor. They both need specific receptors at the surface of the bacteria they will visit (Fig. 4). The prophages’ receptors on the cell surface have specificity although they often are made of normal molecular components of the host, such as proteins, teichoïc acids, polysaccharides, etc. Most significant is the fact that there are receptors for prophages at the surface of all known strains. Like ST plasmids, prophages also possess complex transfer operons which are usually repressed, but capable of derepression, often when surrounding conditions are not favorable enough for their host cells. In this case, copies of these types of small replicons are synthesized independently from the rythmic division of the large replicon (chromosome) of the host cell. The prophages possess the most complex mechanisms for gene transfer among ST small replicons. Both types of ST small replicons form, however, a family of specialized biological entities with the same general type of results in gene transfer. As mentioned by Dreiselkelmann (1994): “Evident similarities between the different DNA translocation Systems suggest a common evolutionary origin of the assembly, structural subunits, and the structure of a DNA transfer apparatus.” These similarities also support the unitary concept of the prokaryotes.

50A prophage with a derepressed transfer operon encodes in its host cell as many as one hundred copies of itself, and protein envelopes and tubules for each of them. These will assemble and become small biological syringes, the temperate phages (Fig. 5). Soon afterwards they are liberated by the encoded enzymes, (e.g. lysozyme) which by lysing part of the host’s cell wall allow them and all the other cellular DNA to leave the donor cell (lytic phase or cycle). Thus, these newly formed prophages with their proteic capsid and tail have become transfer robots, called temperate phages. They are able to reach even far away receptive cells which present appropriate surface receptors, and to subsequently lysogenize them. This means that they may be accepted as stable small replicons with their transfer operon repressed. In most cases, when the transfer operon is repressed (lysogeny) (Lwoff, 1953) and only the converting genes of the prophages are expressed, the activity of the host cell is modified (Cleckner et Sonea, 1966). In other cases, cells injected with a prophage will be lysed by the activity of its derepressed transfer operon. For the same reason, some lysogenic cells, following derepression of their transfer operon, produce many temperate phages. The other cellular genes liberated by the cell lysis are ready to participate in transformation in the case of the large replicon’s genes, or in transfection in the case of the other small replicons. The majority of the cells in the population of lysogenic strains have their transfer operons repressed, and only in a few of them do they become derepressed in a well-controlled way. This “spontaneous induction” has been viewed for a long time as a perplexing phenomenon. However, when one looks at the prophages as the most complex and efficient of the small replicons, spontaneous induction appears logical and of great biological significance. This shifting from the lysogenic to the lytic phase happens, on every division, at least in one or a few cells in every thousand. This maintains in a lysogenic strain a constant production of temperate phages by spontaneous induction, accompanied by the liberation of all the other DNA of their former host. It is a repetitive and abundant offer of genes for other strains. Any other small replicon of the donor cell is also freed by the lysed cell and may participate in transfer, and the large replicon’s genes are also liberated and made available for transformation. An optimal proportion of all the prophages from a lysogenic strain are thus induced, hence multiplied, and constantly spread as temperate phages, once they are protected by their acquired membrane and endowed with a biological needle (the “tail” of the phage).The temperate phages built in this way are carrying the prophages genes and, often, along with them, other types of genes: other small replicons or stable genes from the large replicon of the donor cell (transduction). This constant liberation of genes freed in nature and available for exchanges is one of the most generalized stable mechanisms of gene spreading in astronomical numbers, all ready to enter a different cell. Specialists in marine microbiology have shown several years ago that ocean waters contain surprisingly high concentrations of tailed bacteriophages more than half of which are carrying prophages (Bergh et al., 1989). Transfer of converting genes by lysogenisation is evidently more frequent than it was believed up to now. Moreover, transduction, the transfer by a temperate phage of genes other than its own, present in the donor cell can happen in lakes, oceans, rivers, soil and sewage treatment facilities. Work by R.V. Miller and his group (1998) has shown that very high concentrations of bacteriophages can be found in some fresh and marine water samples. Again, since temperate bacteriophages can transduce DNA to several different strains their role in broadcasting bacterial genes on a high scale and to faraway places is certainly much greater than formerly estimated. Transduction is also known to occur in soil, plant surfaces, shellfish, etc. (Miller, 1998). There is no reason to exclude the possibility that it often takes place in almost any bacterial community.

51In addition to the general activity favoring gene exchanges mentioned above, prophages have the necessary genetic information to fetch favorable genes when their lysogenic strains are in difficulty. It can start with a provoked induction which may involve up to 90% of the cells of this lysogenic strain, which thus releases in nature many billions of temperate phages. When one of these phages reaches a prokaryotic strain containing the useful gene for the lysogenic strain in need and bearing an adequate receptor, it will inject its prophage into one cell which, generally, will start a lytic cycle with the result that numerous copies of its temperate phage are produced and liberated. Such copies will in turn continue to inject progressively millions of other receptive cells and to produce appropriate quantities of transducing phages carrying the needed gene for the original lysogenic strain. There is a reasonable possibility that a few such temperate phages, containing their prophages enriched with the needed gene, will return to the original strain and help it to solve its problem. Experimentally, such a process can easily be performed and proved possible.

Figure 5

DC: Donor cell
FPh: Filamentous phage
LR: Large replicon
Pr: Prophage
P: Pilus
R: Receptor
RC: Receiving cell
STP: Self-transmissible plasmid
TPh: Temperate phage

52The three known types of self-transmissible (ST) small replicons and the ways in which each of them actively transfers genes for the possible benefit of receiving cells. Active transfer of such small replicons implies fïrst a peculiar multiplication (not the usual one, which is simultaneous with the host cell) and the transfer of copies with the help of tubules characteristic for each of the three types of ST small replicons. In the case of ST plasmids, tubules (pili) remain attached to the donor cell (DC) and can get in contact, by their other end, with receptors (R) on the surface of the receiving cells (RC).

53The filamentous phage (FPh) gets a protective tube from a temporary physical modification of its host cell’s cytoplasmic membrane as its linear DNA crosses it and is freed into the surrounding solutions. When such a liberated phage approaches the pilus of another cell bearing a compatible receptor it sticks to it and injects its DNA into the receiving cell (RC).

54The prophages (Pr) are synthesized in large numbers (ca. 100/cell) when their transfer operon is derepressed. Each is assembled with a capsule (head) and a tube (tail) as a temperate phage and eventually gets out of the donor cell and free in nature following the lysis of the latter by lysozyme; the last action is also directed by the prophage’s transfer operon. Later, when a phage finds cells with appropriate surface receptors, the end of the phage tube (tail) clings to one of them and its DNA content, usually the prophage, is injected into the receiving cell.

55It is evident that prophages are the most active and complex exchangers of prokaryotic genes not only by acting alone but also in association with each type of plasmid or by transduction of genes from the large replicon. When they lyse cells during their synthesis, they also favor to the utmost transformation and transfection in their community. No System or mechanism equivalent to the prophages exists in the eukaryotic world.

56As we have seen, the self-transmissible, free form of the prophage is the temperate phage, which is a tailed one. It is interesting to recall that another type of DNA phage, the filamentous one, may carry converting genes and establish a kind of lysogenic-like situation (fig.5). For example, a whole genetic region on the large replicon of Vibrio cholerae, the bacterium responsible for the serious contagious disease cholera, codes for different substances including the enterotoxin and the factor directing the phage parts assembly. This genetic region (also called a genetic cassette) apparently comes to V. cholerae as elements of a filamentous phage (Fasano, 1997). The filamentous phages have a linear DNA molecule containing replicator genes and other genes, for transfer. The latter encode a tubular envelope which protects this phage’s DNA when it passes into the extracellular environment. It is added to copies of the phage’s DNA before they leave the host cell. Their DNA is also a transposon and it has been shown to contain converting genes. This is a different type of gene exchange. It is based on elements other than those in prophages and plasmids but with a somewhat similar gene transfer activity and a few other characteristics reminescent of ST plasmids and the prophage-temperate phage complex.

Transformation and small replicons support a global free market of genes for all prokaryotes

57Together with transformation (including transfection) and with the concomitant transfer of all the genes of a strain when prokaryotic cells move between different communities of mixed strains, small replicons participate abundantly in the generalized and largely available genetic free market of the global prokaryotic entity. Together they possess and spread a complete collection of all the genetic information carried by prokaryotic genes, a memory of all its biochemical diversity and of the numerous possibilities for combining all the tested ways to cope with different situations. A global communication network has emerged from this arrangement, supported by the gigantic genetic information bank on which superior prokaryotic functions are based. All these genetic blueprints represent the entire genetic patrimony of prokaryotes, kept open for use by any strain (Sonea, 1990). Once we realize this, it becomes less surprising that it took only a few decades for infectious bacteria to acquire resistance genes to dozens of antibiotics, one after another, all around the world.

3. The prokaryotic entity forms one planetary biologic System or superorganism

58The capacities of the complex global entity formed by the prokaryotic world result in a giant efficient System much greater than the sum of all its basic elements. The latter, as well as the mixed prokaryotic communities may temporarily change their composition when needed. These rearrangements are made within the global prokaryotic entity which acts as the reservoir of all these genes and strains, and thus remains the basic stable element of the prokaryotes from which diversity and change originate and spread. In this giant pool are distributed numerous genes of each kind randomly in different sites where they are available for use and exchange. Their survival is ensured.

59Today (and it has probably been so for ca. 2.5 B.Y.) genes of prokaryotic cells may have their copy transferred to other types, and accepted if useful. In the same way, prokaryotic cells may be accepted in different communities of mixed strains if this can participate in division of labor and contribute to the survival of these communities. It follows that there are astronomical numbers of prokaryotic cells and also of separated genes in our biosphere making up a permanently renewed pool of candidates for advantageously replacing the less successful basic elements in the different regions of the prokaryotic world. Therefore, one can talk of a giant common genome of prokaryotes, representing the totality of their genetic patrimony, available to cells or communities of mixed strains which might use it profitably. This allows prokaryotes to perform complex functions similar to those of computers (Sonea, 1988a, 1989). As a result, the blue-prints of hereditary information of the entire prokaryotic world are constantly offered and replenished, supported by the giant reservoir of genes available through the agency of exchange mechanisms. Even if these mechanisms work more frequently between closely related strains, the astronomical numbers of cells and small replicons present in different places and situations in the biosphere will, in time, provide solutions where they are needed. The selection of the best information is performed at the reception site: the mixed community doing it for cells and the individual cell for the genes. If a new addition or replacement helps the receiver and improves its functioning, the latter with its new hereditary stock, will multiply abundantly. The best immediate solutions are, in a sense, constantly selected everywhere and, at the global level, this results in exceptional capacities for the prokaryotic superorganism, a powerful half of the living world. As noted earlier, the latter’s characteristics are surprisingly similar to those of a modem advanced society: free market, generalized competition, high specialization and, often, standardization of the basic units resulting in generalized access to useful information and global communications. Open competition chooses the best solution.

60Prokaryotic cells are present everywhere in our biosphere (Whitman et Wiebe, 1999). They maintain its stability and its life-sustaining qualities. Their realm is mainly, but not exclusively, situated betwen the most external lifeless geologic strata on the one hand, and the lowest part of the atmosphere on the other, but evidence now exists that microbial ecosystems are present even in the earth’s subsurface (Krumholz, 1998; Whitman et Wiebe, 1999). As mentioned previously, the larger concentrations of prokaryotic cells exist as mixed strains in fertile soils, ocean beds, marshes, alimentary tracts of all animals, etc. These communities are not uniform combinations, they are different from one place to another and even vary in time, and with the season. Their cells are often entangled with eukaryotes which live immersed in the prokaryotic world which fills our biosphere. This crowding by prokaryotic cells favors the collaborating strains and discourage the development of less sociable cells. Moreover, such high concentrations make the stabilizing action of the prokaryotes on the biosphere continuous and more efficient.

61The variety of biochemical reactions among the strains studied so far is extremely rich and it seems that all possible specializations for cells of the prokaryotic type have already been accomplished long ago and kept available. The number of enzymes in different types of cells remains relatively small (at a minimum) and each kind of biochemical reaction fïts somewhere in the great variety of local mixed communities. This favours successful settlement and survival in favorable niches and results in major contributions to the cycles of organic and inorganic substances, the production of recuperable waste and the maintenance of homeostasis. The stability of our atmosphere, its chemical composition, results in large part from prokaryotic activities and it has been so for at least 1.5 B.Y.

62Certain features related to their solidarity are common to almost all prokaryotic cells. Small size is one such feature. It allows the production of enormous numbers of different cells, often billions, in the same neighborhood, increasing the likelihood of exchanges of cells and genes, and cross-feeding. Also, the speed of chemical reactions and the pace of growth and cellular division all contribute to the development of stable communities. The larger the number of types and cells, the greater the probability of useful exchanges and collaboration.

63Archaea share with eubacteria basic prokaryotic characteristics. They are highly specialized in their biochemical roles, yet also able to function in associations where division of labor is practiced. Their cells are equally small, surrounded by a resistant wall, and harbor the two types of replicons which function in the same way as those of eubacteria. We believe that the two categories, archaea and eubacteria, taken together, are part of the global prokaryotic superorganism with a lifestyle entirely different from that of eukaryotes.

What are the most common prokaryotic shapes?

What is not common in prokaryotic cells?

What are the 4 types of prokaryotes?

Which of the following is not a structure of a prokaryotic cell?