Non-canonical Biosynthesis of the Brexane-Type Bishomosesquiterpene Chlororaphen through Two Consecutive Methylation Steps in Pseudomonas chlororaphis O6 and Variovorax boronicumulans PHE5-4.

A non-canonical biosynthetic pathway furnishing the first natural brexane-type bishomosesquiterpene (chlororaphen, C17 H28 ) was elucidated in the γ-proteobacterium Pseudomonas chlororaphis O6. A combination of genome mining, pathway cloning, in vitro enzyme assays, and NMR spectroscopy revealed a three-step pathway initiated by C10 methylation of farnesyl pyrophosphate (FPP, C15 ) along with cyclization and ring contraction to furnish monocyclic γ-presodorifen pyrophosphate (γ-PSPP, C16 ). Subsequent C-methylation of γ-PSPP by a second C-methyltransferase furnishes the monocyclic α-prechlororaphen pyrophosphate (α-PCPP, C17 ), serving as the substrate for the terpene synthase. The same biosynthetic pathway was characterized in the ß-proteobacterium Variovorax boronicumulans PHE5-4, demonstrating that non-canonical homosesquiterpene biosynthesis is more widespread in the bacterial domain than previously anticipated.

Stone age diseases and modern AIDS.

The great advantage of being a sexually transmitted disease is the ability to survive and specialize solely on a host species that is present in low numbers and widely distributed so that contact between infected and uninfected organisms by chance is rare. Pathogens of a sparse, but widely distributed host species, must either: i) have an alternative host; ii) be able to survive in a dormant state; or iii) be non-destructive to their host. For the pathogens of a diploid there is a particularly effective strategy, that of being sexually transmitted. Then the hosts' themselves transfer the pathogen.

Evolution of temperate pathogens: the bacteriophage/bacteria paradigm.

BACKGROUND: Taking as a pattern, the T4 and lambda viruses interacting with each other and with their Gram-negative host, Escherichia coli, a general model is constructed for the evolution of 'gentle' or temperate pathogens. This model is not simply either pure group or kin selection, but probably is common in a variety of host-parasite pairs in various taxonomic groups. The proposed mechanism is that for its own benefit the pathogen evolved ways to protect its host from attack by other pathogens and this has incidentally protected the host. Although appropriate mechanisms would have been developed and excluded related viral species and also other quite different pathogens, the important advance would have been when other individuals of the same species that arrive at the host subsequent to the first infecting one were excluded. RESULTS: Such a class of mechanisms would not compete one genotype with another, but simply would be of benefit to the first pathogen that had attacked a host organism. CONCLUSION: This would tend to protect and extend the life of the host against the detrimental effects of a secondarily infecting pathogen. This leads to the pathogens becoming more temperate via the now favorable co-evolution with its host, which basically protects both host and virus against other pathogens but may cause slowing of the growth of the primary infecting pathogen. Evolution by a 'gentle' strategy would be favored as long as the increased wellbeing of the host also favored the eventual transmission of the early infecting pathogen to other hosts.

The exocytoskeleton.

The murein or peptidoglycan wall enclosing most bacteria is essential for the life style of most organisms in the Domain of Bacteria. Only in special situations does it not play a role in the bacterial growth cycle. When life first appeared on this planet the cellular osmotic pressure was probably low and a sacculus was probably not relevant, but became necessary as bacterial life evolved from the complex and sophisticated cell called the Last Universal Ancestor. The construction of the murein wall outside of the cytoplasmic membrane is complex and requires elaborate special biochemistry. Growth of the sacculus in some parts of the surface and not in others is important for bacteria cells and allows them to divide and grow without becoming larger and larger and for their being able to maintain a shape characteristic of individual species.

Unidirectional movement of flares of cells of Myxococcus xanthus.

Amongst other modes, Myxococcal cells move in swarms that are flares or columns of cells. It has been argued that this is a strategy allowing a large enough number of them to encounter food bacteria. Then, the combined large amount of extracellular lytic enzymes from the mass of cells can provide adequate nutrient resources from the food bacteria for all the myxococci of the swarm. However, how they move as a coherent column has not been adequately explained. Here based on the idea that a rare cell can experience a special mutation such that it moves only unidirectionally, a proposal to account for this aspect of Myxococcus cell movement is suggested. Although wild type individual organisms of this species engage in forward and back movements, a mutant cell that moves unidirectionally can bias the movement of associated wild type cells and lead to the formation of a column of cells, headed by such a unique mutated cell. The non-mutated cells follow along it is suggested because of the S-motility (or social motility) system. This may link them to this single unidirectionally moving mutant cell to give a coherent movement to the column. This proposed type of mutation back mutates to wild type and the column no longer functions as such and only wild-type cells are present.

Partition of old murein in small patches over the entire wall of E. coli cells forced to grow as a coccoid.

With the development of a technique to visualize the ages of different portions of the sacculus, De Pedro et al. showed that the sacculus of Escherichia coli was tripartite: (i) the establish poles contained only old wall, (ii) the nascent poles (or septa) were composed entirely of new murein, and (iii) the elongating cylindrical wall was a mixture of patches of both old and new peptidoglycan. This short note presents a computer analysis of data files of work presented in the recent paper by De Pedro et al. of the growth pattern of the wall of E. coli forced to grow in a quite unusual morphology as large spheres in the presence of mecillinam. Compared with rod-shaped cells, only very small patches (spikes) of old wall were retained interspersed with new murein during the conversion to large spheroids. This subdivision appeared to be the case for both the previous wall of the poles, which are ordinarily retained intact, and the previous patches retained within the cylindrical wall. These very small patches after the conversion to spheroids were much smaller than the sidewall patches in rod-shaped cells reported previously. This implies that the mechanism that prevents the insertion of new wall into both the wall of the poles and the old wall patches of the sidewall in the presence of mecillinam is superseded by insertion throughout the old wall. The work in the De Pedro et al. paper from 2001 was done with cells of same strain as in the earlier papers with rod-shaped cells, so the results of computer analysis of the fluorescence micrographs can be critically compared.

The first cell.

The First Cell arose in the previously pre-biotic world with the coming together of several entities that gave a single vesicle the unique chance to carry out three essential and quite different life processes. These were: (a) to copy informational macromolecules, (b) to carry out specific catalytic functions, and (c) to couple energy from the environment into usable chemical forms. These would foster subsequent cellular evolution and metabolism. Each of these three essential processes probably originated and was lost many times prior to The First Cell, but only when these three occurred together was life jump-started and Darwinian evolution of organisms began. The replication of informational molecules that made only occasional mistakes allowed evolution to form all the basic components of cellular life. Ribozymes, the first informational molecules, were also catalytic. Energy coupling required the formation of a closed lipid surface to generate and maintain an ion-motive gradient. The closed vesicle partitioned components and avoided dilution within the primordial sea. Closed membranes were essential for the first self-reproducing cell to arise and for its descendants to disperse. Subsequent cellular development after the origin of The First Cell led to the beginnings of intermediary metabolism and membrane transport processes. This long process, subject to strong evolutionary selection, developed the cellular biology that is now shared by all extant organisms.

Shapes that Escherichia coli cells can achieve, as a paradigm for other bacteria.

Because the sacculi of Gram-negative rod-shaped cells are so thin, it is difficult to imagine how they grow and divide and maintain a characteristic shape and size. Abnormal cell shapes can be produced, under special conditions in Escherichia coli. These findings suggest a basis for the variety of bacterial shapes in terms of the Surface Stress Theory. Some proposals are presented to understand the form and function of rods, cocci, fusiform organisms, as well as other bacteria of other shapes using the molecular biology and physiology now known for E. coli.

Catastrophe and what to do about it if you are a bacterium: the importance of frameshift mutants.

Key problems that bacteria have historically faced are the challenges of the lack of essential nutrients and the presence of antibiotics produced naturally, but there are many other challenges. It appears that for many of these challenges the bacteria have mechanisms encoded in their genomes that are not usually functioning, but may be "turned on" when needed, even if the need only occurs once in hundreds of thousands of generations. Such mechanisms at other times somehow need to be "turned off" because they may cause a slight disadvantage, or even a grave disadvantage to the cell compared with wild-type cells during the time the population is not being challenged. On the other hand, a gene cannot simply be discarded because it might be needed again. How do microorganisms solve the problem of responding to challenges that only occur rarely? I suggest that in most cases, the mutation must occur by the existence of a readily reversible mutation. The mutation in likely the result of a frameshift mutation that caused the response and later another frameshift occurs to return the genome to its original state.

Bacterial wall as target for attack: past, present, and future research.

When Bacteria, Archaea, and Eucarya separated from each other, a great deal of evolution had taken place. Only then did extensive diversity arise. The bacteria split off with the new property that they had a sacculus that protected them from their own turgor pressure. The saccular wall of murein (or peptidoglycan) was an effective solution to the osmotic pressure problem, but it then was a target for other life-forms, which created lysoymes and beta-lactams. The beta-lactams, with their four-member strained rings, are effective agents in nature and became the first antibiotic in human medicine. But that is by no means the end of the story. Over evolutionary time, bacteria challenged by beta-lactams evolved countermeasures such as beta-lactamases, and the producing organisms evolved variant beta-lactams. The biology of both classes became evident as the pharmaceutical industry isolated, modified, and produced new chemotherapeutic agents and as the properties of beta-lactams and beta-lactamases were examined by molecular techniques. This review attempts to fit the wall biology of current microbes and their clinical context into the way organisms developed on this planet as well as the changes arising since the work done by Fleming. It also outlines the scientific advances in our understanding of this broad area of biology.

Cell wall-deficient (CWD) bacterial pathogens: could amylotrophic lateral sclerosis (ALS) be due to one?

Recently, a number of diseases that had been thought previously to be caused by something other than an infectious agent are now known to be caused by bacteria. It now appears that it is not uncommon that bacteria, viruses, or fungi can cause diseases even when these organisms have not been detected or cultured. The most recent, well-publicized case is that of stomach ulcers; these are largely due to Helicobacter pylori infections. Here, the possibility is explored that amylotrophic lateral sclerosis (ALS) is caused by a cell wall-deficient microorganism.

Patchiness of murein insertion into the sidewall of Escherichia coli.

This paper extends, with computer techniques, the authors' previous work on the kinetics of pole wall and sidewall synthesis in Escherichia coli. These findings extend the conclusion that the nascent poles are made of entirely new material and that no new material is inserted into old poles. This requires re-evaluation of ideas in the literature about wall growth and cell division. Mechanisms of various types have been suggested for the growth of Gram-negative rod-shaped bacteria and these will also require major re-evaluation because of the finding, reported here, that the sidewall is made in several modes: patches of new murein, bands of new material largely going circumferentially around the cell, and areas of the sidewall that are enlarged by an intimate and regular admixture of new with the old muropeptides.

Were Gram-positive rods the first bacteria?

At some point in the evolution of life, the domain Bacteria arose from prokaryotic progenitors. The cell that gave rise to the first bacterium has been given the name (among several other names) "last universal ancestor (LUA)". This cell had an extensive, well-developed suite of biochemical strategies that increased its ability to grow. The first bacterium is thought to have acquired a covering, called a sacculus or exoskeleton, that made it stress-resistant. This protected it from rupturing as a result of turgor pressure stress arising from the success of its metabolic abilities. So what were the properties of this cell's wall? Was it Gram-positive or Gram-negative? And was it a coccus or a rod?

Why are rod-shaped bacteria rod shaped?

Generally speaking, bacteria grow and divide indefinitely, and as long as the growth conditions are maintained they retain constant dimensions and shapes with little variation. How they do this is a question that I have been considering for three decades. Here, I discuss two hypothetical mechanisms, one for Gram-positive rods and the other for Gram-negative rods. These mechanisms are consistent with what is known, but make some unproven assumptions.

Control of the bacterial cell cycle by cytoplasmic growth.

For free-living single-celled organisms, it can be assumed that it is their success in acquiring resources and converting them into cytoplasm that controls the timing of their cell cycles. Cytoplasm is the sink for the bulk of the environmental resources. It must be the case that this type of control must operate in dilute cultures under adequate nutrition in a constant environment. It follows that there ought to be mechanisms that measure or count the cell's biomass or some component of the cytoplasm to measure their growth success. Besides sensing their biomass, they need to know when a certain value of the cell size has been achieved. When this critical state has been achieved, the cell needs to have an all-or-none trigger that either initiates chromosome replication, the completion of cell replication, cell division, or the process of separating sister cells physiologically or physically. Any of these four different stages, in principle, may be the one triggered in response to cell growth in different species of microorganisms. Alternatively, multiple triggers at different cell sizes may be activated at different cell cycle stages. Although initiation of chromosome replication has been believed to be the event triggered in Escherichia coli, this probably is not generally the case and other control mechanisms may act in other prokaryotes. How the increase in cell biomass is self-assessed and used to carry out critical cell cycle events is not understood in any case. This deficiency in our knowledge of microbial cell physiology is grave. The factor that probably has prevented the elucidation of the mechanisms in any organism is that enzymatic processes deal with concentrations, and a cell cycle trigger must respond to the total amount of material present in a cell. This article discusses the theoretically possible classes of mechanisms for the cell to respond when it has achieved its appropriate critical size. These breakdown into three groups: those mechanisms that assess the total amount of biomass or some special subcellular component, and those that measure the ratio of one component to another component where their two syntheses are differently controlled by cell physiology and morphology, and a third group with some specialized mechanisms.

Diffusion through agar blocks of finite dimensions: a theoretical analysis of three systems of practical significance in microbiology.

A number of experimental methods in biology depend on the kinetics of diffusion of a substance through a gel. This paper reviews the diffusion equations, gives the experimental limitations for some useful cases, and presents computer simulations for cases that cannot be treated analytically. While double diffusion is not considered, three single-diffusion situations are treated. (1) Systems for the study of chemotaxis in the gliding bacterium Myxococcus xanthus. Experimental designs used for this in many cases in the literature were inappropriate and mathematical analysis of these is presented. (2) The development of gradient plates. The time necessary for vertical diffusion to become substantially complete and before diffusion in the direction of the original slant has proceeded significantly is calculated. (3) The application to antimicrobial disk susceptibility tests. The basis of the measurement of antibiotic sensitivities with disks containing antimicrobial agents, as routinely used in clinical microbiological and testing laboratories, is analysed and the limitations are assessed and improvements suggested.