The Gram-stain technique is used to classify bacteria as either Gram-positive or Gram-negative depending on their colour following a specific staining procedure originally developed by Hans Christian Gram. As the word "Gram" is derived from a name it is always written with an upper case "G".

Following the Gram stain procedure, and on visualization with a microscope Gram-positive bacteria appear dark blue or violet due to the crystal violet stain; Gram-negative bacteria, which cannot retain the crystal violet stain, appear red or pink due to the counterstain. Gram-positive bacteria retain the crystal violet due to a difference in structure of their cell wall, specifically the amount of peptidoglycan.

Gram-negative bacteria do not retain the crystal violet dye in the Gram stain protocol. Gram-negative bacteria will thus appear red or pink following the Gram stain procedure due to the effects of the counterstain (for example safranin).

The cell envelope is defined as the cell membrane and cell wall plus an outer membrane, if one is present. The cell envelope of Gram-negative bacteria contains an outer membrane composed by phospholipids and lipopolysaccharides which face the external environment. The lipopolysaccharides confer an overall negative charge to the Gram-negative cell wall. The chemical structure of the outer membrane lipopolysaccharides is often unique to specific bacterial strains. Many species of Gram-negative bacteria are pathogenic. This pathogenicity is often associated with the lipopolysaccharide layer of the Gram-negative cell envelope.

Gram-negative bacteria have a characteristic cell envelope structure very different from Gram-positive bacteria. Gram-negative bacteria have a cytoplasmic membrane, a thin peptidoglycan layer, and an outer membrane containing lipopolysaccharide. There is a space between the cytoplasmic membrane and the outer membrane called the periplasmic space or periplasm. The periplasmic space contains the peptidoglycan.

Genera of Gram-negative bacteria include:

• Acinetobacter
• Actinobacillus
• Bordetella
• Brucella
• Campylobacter
• Cyanobacteria
• Enterobacter
• Erwinia
• Escherichia coli
• Franciscella
• Helicobacter
• Hemophilus
• Klebsiella
• Legionella
• Moraxella
• Neisseria
• Pasteurella
• Proteus
• Pseudomonas
• Salmonella
• Serratia
• Shigella
• Treponema
• Vibrio
• Yersinia

Labels: bacteria, bacteriology, bacterium, gram negative bacteria, gram stain

from Saraswathi Lanka (University of Illinois College of Veterinary Medicine) writing in "Doodys Reviews":
"This broad overview of many aspects of the family Pasteurellaceae provides excellent coverage of the current status of taxonomy and phylogeny of this diverse group of bacteria. ... This is a much needed information resource for researchers. ... This is a rich source of information and provides well balanced coverage of relevant topics. It is a comprehensive guide that provides critical insight into the current understanding of molecular and genomic aspects of Pasteurellaceae" ... read more

Pasteurellaceae: Biology, Genomics and Molecular Aspects
Publisher: Caister Academic Press
Edited by: Peter Kuhnert and Henrik Christensen
Publication date: 2008
ISBN: 978-1-904455-34-9

Labels: bacteria, bacteriology, book review, pasteurella, Pasteurellaceae

Gram-positive bacteria are generally divided into the Actinobacteria and the Firmicutes.

The Actinobacteria or actinomycetes are a group of Gram-positive bacteria with high G+C ratio. They include some of the most common soil bacteria. Other Actinobacteria inhabit plants and animals and including some pathogens, such as Mycobacterium, Corynebacterium, Nocardia, Rhodococcus and a few species of Streptomyces. Actinobacteria produce secondary metabolites and are important to the pharmacological and biotechnology industries. Streptomyces species, for example, produce important antibiotics. Some Actinobacteria form branching filaments and some Actinomycetes species produce endospores.

The majority of Firmicutes have Gram-positive cell wall structure. However some, the Mollicutes or mycoplasmas, lack cell walls altogether and therefore do not respond to Gram staining. They do however lack the second (outer) membrane found in Gram-negative bacteria. Others members of the group, such as Megasphaera, Pectinatus, Selenomonas, and Zymophilus have a porous pseudo-outer-membrane that causes them to stain Gram-negative. The Firmicutes are generally restricted to a core group of related bacteria, called the low G+C group in contrast to the Actinobacteria. Firmicutes can be cocci or rod-shaped forms. Many produce endospores. They are found in various environments and some members of the group are important pathogens.

Recommended reading:

Clostridia: Molecular Biology in the Post-genomic Era      

Corynebacteria: Genomics and Molecular Biology

Mycobacterium: Genomics and Molecular Biology

Bacillus: Cellular and Molecular Biology

Staphylococcus: Molecular Genetics

Lactobacillus Molecular Biology: From Genomics to Probiotics

Genomics of GC-Rich Gram-Positive Bacteria

Labels: bacillus, bacteriology, bacterium, clostridia, clostridium, corynebacterium, lactobacillus

The genus Xanthomonas consists of 20 plant-associated species, many of which cause important diseases of crops and ornamental plants. Individual species comprise multiple pathovars, characterized by distinctive host specificity or mode of infection. Genomics is at the center of a revolution in Xanthomonas biology. Complete genome sequences are available for nine Xanthomonas strains, representing three species and five pathovars, including vascular and non-vascular pathogens of the important models for plant biology, Arabidopsis thaliana and rice. With the diversity of complete and pending Xanthomonas genome sequences, the genus has become a superb model for understanding functional, regulatory, epidemiological, and evolutionary aspects of host- and tissue-specific plant pathogenesis.
Further reading: Damien F. Meyer and Adam J. Bogdanove Chapter 7 in Plant Pathogenic Bacteria

Furthermore, Xanthomonas strains produce the acidic exopolysaccharide xanthan gum. Because of its physical properties, xanthan gum is widely used as a viscosifer, thickener, emulsifier or stabilizer in both food and non-food industries.
Further reading: Anke Becker and Frank-Jörg Vorhölter Chapter 1 in Microbial Production of Biopolymers and Polymer Precursors

Labels: bacteriology, bacterium, biopolymers, biotechnology, xanthan, xanthomonas

The methods currently available to diagnose Legionnaires' disase are culture, urinary antigen detection, direct fluorescent antibody testing, detection of nucleic acid and detection of specific antibodies in serum samples. Presently, none of the diagnostic tests available offers the desired quality with respect to sensitivity and specificity. Culture should be obligatory, especially when hospitalized patients with underlying diseases are investigated. A positive culture is the prerequisite of molecular epidemiological investigations. Urinary antigen detection is a valuable tool in the majority of community-acquired cases when L. pneumophila serogroup 1 is the causative agent. In cases of nosocomial disease, when Legionella pneumophila serogroups other than sg 1 are frequent, this assay has limitations. The detection of nucleic acid is very useful method of diagnosis but requires further validation. The detection of antibodies in a patient's serum is of little use in the acute phase of the illness. Several molecular subtyping techniques are in use to subtype L. pneumophila strains in epidemiological investigations. Legionella pneumophila is genetically very heterogeneous thus allowing an individual fingerprint of each strain. However, the majority of clinical cases are caused by a limited number of clones that cause disease worldwide. The therapy for Legionnaires' disease requires drugs that can access and are active intracellularly. Currently, fluorochinolones and macrolides are the most active agents.

Further reading: Paul C. Lück in Legionella: Molecular Microbiology

Labels: bacteriology, bacterium, legionella

Recent research suggests that Clostridium-based tumor targeted therapy holds promise for the treatment of solid tumors. Upon systemic administration, various strains of non-pathogenic clostridia have been shown to infiltrate and selectively replicate within solid tumors. This specificity is based upon the unique physiology of solid tumors, which is often characterized by regions of hypoxia and necrosis. Clostridial vectors can be safely administered and their potential to deliver therapeutic proteins has been demonstrated in a variety of preclinical models.

from Asferd Mengesha, Ludwig Dubois, Kim Paesmans, Brad Wouters, Philippe Lambin and Jan Theys in Clostridia: Molecular Biology in the Post-genomic Era

Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Labels: bacteriology, bacterium, clostridia, clostridium, therapy, tumor

Writing in the journal Microbiology Today (Society for General Microbiology, UK), Paul Hoskisson of the University of Strathclyde, UK, reviews a new book on Corynebacteria published by Caister Academic Press:

"it is not surprising that this book has followed the Handbook of Corynebacterium glutamicum (2005, Eggling & Bott, eds). Obviously there is some overlap in these volumes; however, this one is updated and brings in information relevant to other Corynebacteria ... There is a chapter on plasmids and promoters and their applications that researchers will find very useful in a practical sense ... researchers in the field with find this a useful and up to date addition to their library."
For full details please visit Corynebacteria: Genomics and Molecular Biology

Labels: bacteriology, bacterium, book review, corynebacterium

Writing in the journal Microbiology Today (Society for General Microbiology, UK), Madeline Stone & Kathy Bamford from the Imperial College London, UK, review a new book on Staphylococcus published by Caister Academic Press:

"This is an incredibly useful book for anyone with in interest in staphylococci. It provides a broad and in-depth synopsis of up-to-date staphylococcal research. This book is very well suited to its target audiences, researchers who are relatively new to the field and also as a suitable reference for those with greater experience. The first five chapters are particularly informative, providing an excellent overview of the staphylococcal sequencing projects, population structure and evolution of S. aureus, as well as analysis of the methods used ... The chapter on 'Global regulators of Staphylococcus aureus virulence genes' is excellent ... this chapter provides a thorough review of the literature ... We hope that this book will be regularly reviewed and updated in line with this rapidly expanding field."
For full details please visit Staphylococcus: Molecular Genetics

Labels: bacteriology, bacterium, book review, Staphylococcus

Writing in the journal Microbiology Today (Society for General Microbiology, UK), Simon L. Croft of the London School of Hygiene and Tropical Medicine, UK, reviews a new book on Leishmania published by Caister Academic Press:

"The volume is up-to-date; the genome was published in 2005 and the most recent references in the book were published in 2007. There is a richness of information - chapters on gene regulation and the metabolome are particularly engaging ... Let us enjoy a volume that provides a valuable overview of the molecular biology and biochemistry of these fascinating parasites, their metabolic pathways, differentiation process, and their surface molecules"
For full details please visit Leishmania: After The Genome

Labels: bacteriology, bacterium, book review, leishmania

Writing in the journal Microbiology Today (Society for General Microbiology, UK), Edward Bolt of the University of Nottingham, UK, reviews a new book on Archaea published by Caister Academic Press:

"I particularly enjoyed a review on signal transduction in archaea, which captures the frontiersman spirit of some research into Archaea ... The chapter on DNA replication holds it own against several recent review articles in journals ... The book is timely and the publishers promise a 'state-of-the-art overview of Archaea'. In this it mostly works, and its slimness (246 pages) reflects a concise and mostly well-referenced style ... it conveys plenty of the novelty and oddity in Archaea that captures the imagination of students, researchers and PIs."
For full details please visit Archaea: New Models for Prokaryotic Biology

Labels: archaea, bacteriology, bacterium, book review

Writing in the journal International Microbiology, Mercedes Berlanga of the University of Barcelona, Spain, describes a new book on Acinetobacter published by Caister Academic Press as a "useful book ... especially recommended for advanced students in the field, senior researchers, and physicians". She continues to suggest that "all microbiologists will find in the book an exceptional opportunity to extend their understanding of an unusual and unique microbial group."

Other reviewers have also heaped praise on this book, edited by Ulrike Gerischer of the University of Ulm, Germany. It is pleasing that a major new review of Acetinobacter research has been received in such a positive manner by the scientific community. This book will be a major resource for many years to come.

For full details of this book please visit Acinetobacter Molecular Biology

Labels: acinetobacter, bacteriology, bacterium, book review, nosocomial

Mycobacterium tuberculosis, also known as as the "tubercle bacillus" is the bacterium that causes most cases of tuberculosis. The genome has been sequenced and recent research on the genomics and molecular biology of mycobacteria has contributed greatly to our knowledge of this pathogen. Some of the most important recent findings are highlighted here.

Strain Variation and Evolution in Mycobacteria
Mycobacterium tuberculosis appears to be more genetically diverse than generally assumed. There is mounting evidence that this genetic diversity translates into significant phenotypic differences between clinical isolates. M. tuberculosis exhibits a biogeographic population structure and different strain lineages are associated with different geographic regions. Phenotypic studies in the laboratory and in clinical settings suggest that this macro-evolutionary strain variation has implications for the development of new diagnostics and vaccines. Micro-evolutionary variation affects the relative fitness and transmission dynamics of antibiotic-resistant strains. In the light of the emerging epidemic of multidrug-resistant and extensively drug-resistant tuberculosis, there is an urgent need to improve our understanding of the evolution and ecological consequences of strain variation in drug-resistant M. tuberculosis. Further reading: Mycobacterium: Genomics and Molecular Biology

Hypervirulent Mycobacterium tuberculosis
Tuberculosis outbreaks are often caused by hypervirulent strains of Mycobacterium tuberculosis. In experimental animal infections, these clinical isolates elicit unusual immunopathology and may be either hyper- or hypoinflammatory. Similarly, recombinant hypervirulent M. tuberculosis mutants, which exhibit increased bacterial burden or decreased host survival times in model infections, induce a spectrum of inflammatory responses. The majority of hypervirulent mutants identified have deletions in cell wall modifying enzymes or regulators that respond to environmental stimuli. Studies of these mutants have provided insight into the mechanisms that enable M. tuberculosis to mask its full pathogenic potential, inducing a granuloma that provides a protective niche and enables the bacilli to sustain a long-term persistent infection. Further reading: Mycobacterium: Genomics and Molecular Biology

Electron Transport and Respiration in Mycobacteria
Bacteria have evolved a modular respiratory system that enables them to optimize energy production in environments that are variable and may be hostile. By adjusting the composition of the system to suit the specific conditions encountered, the organism is able to thrive in a particular environment. The flexibility conferred by a modular respiratory system is critical to the survival of many bacterial pathogens, including Mycobacterium tuberculosis. The composition of the respiratory systems of sequenced mycobacterial species can be deduced from a comparative analysis of their respiratory gene complements and from the function of specific system components. Common themes have emerged from studies of various models of growth and persistence and can be related to the physiology of this pathogen during infection. Exciting new developments in tuberculosis drug discovery are predicated on targeting respiration and electron transport through inhibition of type II NADH dehydrogenase, ATP synthase, and menaquinone biosynthetic enzymes. Further reading: Mycobacterium: Genomics and Molecular Biology

Lipid biosynthetic machinery of Mycobacterium tuberculosis
Mycobacterium tuberculosis posseses a repertoire of complex lipids. Many of these lipids are crucial to its survival and virulence. Fatty-acyl components of the mycobacterial lipids are synthesised by the concerted action of fatty acid synthases (FASs) and polyketide synthases (PKSs). While the single multifunctional type I FAS carries out de novo biosynthesis from acetyl-CoA, the multicomponent type II FAS generates the very long acyl chains from type I FAS products. Polyketide synthases take over from FAS to complete the biosynthesis of the unusual acyl chains of many exotic lipids like mycolic acids, phthioceroldimycocerosate ester, sulfolipids and mannosyl-beta-1-phophomycoketides. The novel family of fatty acyl-adenylate ligases (FAALs) is crucial to this intricate enzymatic network. FAALs mediate the crosstalk between FAS and PKS by activating long-chain fatty acids to fatty acyl-adenylates which are transacylated onto the PKSs. Further reading: Mycobacterium: Genomics and Molecular Biology

DNA Repair in Mycobacteria
Sequence comparisons indicate that mycobacteria possess the majority of the key DNA repair pathways identified in other bacterial species, including base excision repair, nucleotide excision repair, recombination repair and non-homologous end-joining. However, there are some notable differences such as the absence of a mismatch repair system, as well as variations in the components of other repair pathways. Currently functional studies of DNA repair within mycobacterial species are limited, but this is an expanding area of research. It has been demonstrated that DNA-damage induced mutagenesis is mediated by a different class of DNA polymerase to that used in Escherichia coli. Although the classical SOS system of gene regulation in response to DNA damage is conserved and functional in mycobacteria, many of the DNA repair genes whose expression increases following DNA damage are controlled by an alternative system or systems that are yet to be characterised. The increase in expression observed for a number of Mycobacterium tuberculosis DNA repair genes in infection models suggests that DNA repair might be particularly important during pathogenesis. Further reading: Mycobacterium: Genomics and Molecular Biology

Oxygen, Nitric Oxide, and Carbon Monoxide Signaling in Mycobacteria
Mycobacterium tuberculosis is an aerobe that can survive extended periods of anaerobiosis. The bacillus responds to inhibition of respiration during hypoxic conditions as well as exposure to NO and CO by the induction of over 60 genes, referred to as the "dormancy regulon". Control of the dormancy regulon by NO and CO, not just hypoxia, is mediated by a three component regulatory system composed of two sensors, DosT and DosS and a transcriptional regulator DosR. The dormancy proteins are part of a programmed strategy employed by the bacilli to survive in the absence of aerobic respiration. Further reading: Mycobacterium: Genomics and Molecular Biology

Sulphur Metabolism in Mycobacteria
Sulphur is a key life-supporting element. The recent combined efforts of genomic analysis and laboratory studies have greatly clarified the mycobacterial sulphur metabolic pathways. Sulphur metabolism contributes to intracellular survival and virulence of Mycobacterium tuberculosis. Several enzymes in the sulphur metabolic pathways are essential for mycobacterial survival. Further reading: Mycobacterium: Genomics and Molecular Biology

The Eukaryotic-like Serine/Threonine Protein Kinase Family in Mycobacteria
Mycobacteria have a complex life style comprising different environments and developmental stages. Signal sensing and transduction leading to cellular responses must be tightly regulated to allow survival under variable conditions. Prokaryotes normally regulate their signal transduction processes through two-component systems, however, the genome sequence of Mycobacterium tuberculosis revealed a large number of eukaryotic-like serine/threonine kinases. It is becoming clear that in M. tuberculosis, many of these kinases are involved in the regulation of metabolic processes, transport of metabolites, cell division and virulence. Further reading: Mycobacterium: Genomics and Molecular Biology

Protein Secretion Systems of Mycobacteria
Mycobacteria have a highly complex cell wall. Specialised secretion systems are therefore required to transport proteins across this cell wall. However, genome analysis shows that, apart from the omnipresent Sec and Tat systems, all of the known secretion pathways of other bacteria are absent. Mycobacteria do have a second SecA protein (SecA2) that is involved in the extracellular accumulation of a specific protein subset. In addition, a new secretion pathway was recently identified that is responsible for the secretion of various proteins into the culture supernatant. This pathway is present in multiple copies in the mycobacteria and has been named the type VII secretion pathway. Further reading: Mycobacterium: Genomics and Molecular Biology

Labels: bacillus, bacteriology, bacterium, mycobacteria, mycobacterium, tuberculosis

The genus Clostridium represents a heterogeneous group of anaerobic spore-forming bacteria, comprising prominent toxin-producing species, such as C. difficile, C. botulinum, C. tetani and C. perfringens, in addition to well-known non-pathogens like solventogenic C. acetobutylicum. In the last decade several clostridial genomes have been deciphered and post-genomic studies are currently underway. The advent of newly developed, genetic manipulation tools have permitted functional-based and systems biology analyses of several clostridial strains. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Botulinum and Tetanus Neurotoxins
Botulinum neurotoxins (BoNT) and tetanus toxin (TeNT) are potent toxins which are responsible for severe diseases, botulism and tetanus, in men and animals. BoNTs induce a flaccid paralysis, whereas TeNT causes a spastic paralysis. Both toxins are zinc-dependent metalloproteases, which specifically cleave one of the three proteins (VAMP, SNAP25, and syntaxin) forming the SNARE complex within target neuronal cells which have a critical function in the release of neurotransmitter. BoNTs inhibit the release of acetylcholine at peripheral cholinergic nerve terminals, whereas TeNT blocks neurotransmitter release at central inhibitory interneurons. Only a single form of TeNT is known, but BoNTs are divided in 7 toxinotypes and various subtypes, which differ in amino acid sequences and immunological properties. In contrast to TeNT, BoNTs are associated to non-toxic proteins (ANTPs) to form highly stable botulinum complexes. TeNT is produced by Clostridium tetani, and BoNTs by Clostridium botulinum and atypical strains of Clostridium barati and Clostridium butyricum. The genes encoding the neurotoxin and ANTPs are clustered in a DNA segment, called botulinum locus, which is located on chromosome, plasmid or phage. Neurotoxin synthesis is a highly regulated process, which occurs in late exponential growth phase and beginning of stationary phase, and which is dependent of alternative sigma factors (BotR or TetR). BotR and TetR are related to other clostridial sigma factors, TcdR and UviA, which are involved in the control of Clostridium difficile toxins A and B, and Clostridium perfringens bacteriocin, respectively. BotR, TetR, TcdR and UviA form a new subgroup of RNA polymerase sigma factors. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Clostridium perfringens Enterotoxin
Clostridium perfringens enterotoxin (CPE) causes the intestinal symptoms of a common food-borne illness and ~5-15% of all antibiotic-associated diarrhea cases. In food poisoning isolates, the enterotoxin gene (cpe) is usually present on the chromosome, while cpe is carried by conjugative plasmids in antibiotic-associated diarrhea isolates. CPE action involves its binding to claudin receptors, oligomerization/prepore formation, and prepore insertion to form a functional pore that kills cells by apoptosis or oncosis. The C-terminal half of CPE mediates receptor binding, while its N-terminal half is required for oligomerization. CPE/CPE derivatives are being explored for cancer therapy/diagnosis and improved drug delivery. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

The Cholesterol-dependent Cytolysins and Clostridium septicum α-Toxin
Two classes of pore-forming toxins of the clostridia are represented by the cholesterol-dependent cytolysins (CDCs) and the Clostridium septicum α-toxin. The CDCs are found in a wide variety of clostridial species, but are also found in many species from other Gram-positive genera. As a result, various CDCs have evolved specific traits that appear to enhance their ability to complement the pathogenic mechanism of a specific bacterial species. In contrast, closely related toxins to C. septicum α-toxin (AT) have not been found in other species of the clostridia, although C. perfringens epsilon toxin appears to be distantly related. Remarkably, distant relatives of AT have been found in species of Gram-negative bacteria as well as certain species of mushrooms and the enterolobin tree seed. Although the CDCs appear to be restricted to Gram-positive bacterial pathogens it has recently been shown that the unusual protein fold of their membrane-penetrating domain is present in proteins of the eukaryotic complement membrane attack complex. Both toxins penetrate the membrane by the use of a β-barrel pore but differ significantly in their pore-forming mechanisms. The contribution of both classes of toxins to disease is not yet well understood for the clostridia. It is clear that they play important, but likely different roles in clostridial disease. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Binary Bacterial Toxins
Several proteins from Gram-positive, spore-forming bacilli use a synergistic binary mechanism for intoxicating eukaryotic cells. These toxins include Clostridium botulinum C2 toxin, Clostridium difficile toxin (CDT), Clostridium perfringens iota (ι) toxin, and Clostridium spiroforme toxin (CST). Furthermore, closely related Bacillus species such as Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis produce strikingly similar binary toxins. As per existing literature, these latter proteins have provided a "model" for the clostridial binary toxins. Each of these clostridial and bacillus binary toxins consists of distinct enzymatic "A" and binding "B" proteins that work in concert. Conservation of a basic intoxication theme between different genera clearly suggests retention of an evolutionarily successful mechanism promoting bacterial survival and dissemination throughout Nature. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Group I and II Clostridium botulinum
Clostridium botulinum, producing highly potent botulinum neurotoxin, is a diverse species consisting of four genetically and physiologically distinct groups (Groups I-IV) of organisms. Groups I and II C. botulinum produce A, B, E, and/or F toxins which cause human botulism. In addition, some strains of Clostridium butyricum and Clostridium barati produce type E and F toxins, respectively, and have thus been related to human illness. Human botulism appears in five different forms, such as the classical food botulism, infant botulism, wound botulism, adult infectious botulism, and iatrogenic botulism. Typical of all forms of human botulism is descending flaccid paralysis which may lead to death upon respiratory muscle failure. While the research and diagnostics of botulinum neurotoxigenic clostridia and botulism were based on toxin detection by the mouse bioassay until mid 1990¹s, the subsequent development of molecular detection and typing assays enabled rapid, sensitive, specific, and ethically acceptable molecular epidemiological detection, identification and strain characterization of these organisms, increasing our understanding of the epidemiology of botulinum neurotoxigenic clostridia and botulism. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

C. difficile large clostridial toxins
Clostridium difficile, as all clostridia, is a toxin producing microorganism and the toxins are the main virulence factors. In the early eighties it was clear that two large toxins are produced by the bacterium and epidemiological studies have indicated that strains either produce both toxins (toxin A, TcdA, and toxin B, TcdB) or none of them. Toxigenic strains were usualy associated with the disease, while nontoxigenic were not. This simple situation changed as strains producing only TcdB or strains producing an additional toxin (binary toxin CDT) were described. Such strains with unusual toxin production pattern were subsequently found to have changes in the genomic PaLoc region encoding the toxins TcdA and TcdB. These changes are the basis for a method that distinguish C. difficile strains into toxinotypes. The variability of genes coding for large clostridial toxins (LCTs) has consequences in laboratory diagnosis, changes in understanding of the role of both toxins in pathogenesis, in structure function relationships and in the understanding of the evolutions of LCTs. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Comparative Genomics of Clostridium difficile
The recent emergence of hypervirulent strains of Clostridium difficile and their ability to spread across continents, has caused alarm in both hospitals and the community. This has drawn attention away from other important pathogenic C. difficile strains, which are responsible for significant morbidity and mortality. Little is known about the genetic diversity of these strains and their less pathogenic counterparts. The recent publication of the genome sequence of strain 630 and advances in both microarray and mutagenesis technologies promises to revolutionise our understanding of the pathogenesis and population dynamics of C. difficile. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Surface Structures of Clostridia
The cell wall of Clostridium difficile has an architecture typical of other Gram-positive bacteria. A thick peptidoglycan layer lies external to the cell membrane with many associated cell wall proteins. In C. difficile two major cell wall proteins constitute the S-layer, a paracrystalline two-dimensional array surrounding the entire cell. The sequences of these S layer proteins (SLPs) are variable between strains, perhaps reflecting immunological pressures on the cell. The genome sequence reveals a family of proteins with homology to the high molecular weight SLP; each of these proteins have a second unique domain but their functions remain largely uncharacterised. This family of cell wall proteins is also found in some other species, for example C. botulinum and C. tetani, but not in others such as C. perfringens. Some cell wall proteins of C. difficile, including the SLPs, have properties that imply an involvement in pathogenesis, particularly in binding to host cell tissues. The cell wall proteins of C. difficile may also act as immunogens to induce a partially protective immune response to infection, and may be considered as components of future vaccines against C. difficile associated disease. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Antibiotic resistance determinants in Clostridium difficile
Clostridium difficile, the well known nosocomial pathogen responsible for the majority of antibiotic associated diseases, is increasingly recognised also as the cause of community-associated disease and of enteric disease in animals. The organism is resistant to several antibiotics and can survive disruption of the normal intestinal flora after antibiotic treatment exploiting this advantage to colonize and cause disease. The study of the mechanisms responsible for resistance have highlighted the presence of mobile genetic elements in the C. difficile genome, potentially acquired from other microorganisms. C. difficile might be able to disseminate resistance determinants to other species, thus collaborating to the evolution of the antibiotic resistant patterns that characterise the bacteria circulating worldwide. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Genetic Knock-out Systems for Clostridia
Despite the medical and industrial importance of the genus Clostridium our understanding of their basic biology lags behind that of their more illustrious counterpart, Bacillus. The advent of the genomics era has provided new insights, but full exploitation of the data becoming available is being hindered by a lack of mutational tools for functional genomic studies. Thus, in the preceding decades the number of clostridial mutants generated has been disappointingly low. On the one hand, the absence of effective transposon elements has stymied random mutant generation. On the other hand, the construction of directed mutants using classical methods of recombination-based, allelic exchange has met with only limited success. Indeed, in the majority of clostridial species mutants are largely based on integration of plasmids by a Campbell-like mechanism. Such single crossover mutants are unstable. As an alternative, recombination-independent strategies have been developed that are reliant on retargeted group II intron. One element in particular, the ClosTron, has been devised which provides the facility for the positive selection of mutants. ClosTron-mediate mutant generation is extremely rapid, highly efficient and reproducible. Moreover the mutants made are extremely stable. Its deployment considerably expands current options for functional genomic studies in clostridia. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Clostridia in Anti-tumor Therapy
Although traditional anticancer therapies are effective in the management of many patients, there are a variety of factors that limit their effectiveness in controlling some tumors. These observations have led to interest in alternative strategies to selectively target and destroy cancer cells. In that context, Clostridium-based tumor targeted therapy holds promise for the treatment of solid tumors. Upon systemic administration, various strains of non-pathogenic clostridia have been shown to infiltrate and selectively replicate within solid tumors. This specificity is based upon the unique physiology of solid tumors, which is often characterized by regions of hypoxia and necrosis. Clostridial vectors can be safely administered and their potential to deliver therapeutic proteins has been demonstrated in a variety of preclinical models. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Metabolic Networks in Clostridium acetobutylicum
Clostridia belong to the few bacterial genera able to undergo cell differentiation. They can either grow vegetatively or form endospores, the most resistant survival form of all living organisms. Some species, e. g. Clostridium acetobutylicum, link the metabolic network of sporulation to that of solventogenesis (formation of acetone and butanol). This gives them an ecological advantage by preventing toxic effects of acidic end products from the fermentation and allows them to stay longer metabolically active. In other clostridia, even toxin formation is coupled to sporulation. The key component for these links at the molecular level is the response regulator Spo0A in its phosphorylated form. In contrast to bacilli, clostridia do not possess a phosphorelay for Spo0A activation. Instead, phosphorylation is catalyzed directly by still unknown kinases or by butyryl phosphate. In addition to Spo0A~P, various other regulators are required to control the different metabolic networks. Systems biology is a new approach to understand these processes and their interaction at the molecular level and to adapt them for biotechnological use. Further reading: Clostridia: Molecular Biology in the Post-genomic Era

Labels: bacillus, bacteriology, bacterium, clostridia, clostridium

from Gerard Carter in Aus. J. Med. Sci. (2008) 29: 63-64

Pathogenic Treponema, edited by Radolf and Lukehart, is a comprehensive update of the current state of knowledge of the Treponemes and other spirochaetes ... Thirty five scientists who specialise in molecular biology, epidemiology, entomology and microbiology have pooled their findings of their more recent research efforts to produce an up-to-date account of Treponemal biology ... This hardback text is sturdily bound (less and less common these days) and beautifully printed on very high quality paper. It will make a welcome and useful addition to the libraries of any microbiological research laboratory and pathology establishment.

Further reading: Pathogenic Treponema

Labels: bacteriology, bacterium, book review, treponema

from Paul H. Edelstein in Legionella: Molecular Microbiology

The history of Legionnaires' disease began at least 33 years before the 1976 Philadelphia epidemic, when Legionella micdadei was isolated from human blood. Multiple isolations of several different Legionella spp. were made prior to 1976, and it was known by 1968 that tetracycline therapy prevented deaths in L. pneumophila-infected chicken embryos. The 1976 epidemic provided the scientific focus and resources necessary to determine that L. pneumophila caused epidemic pneumonia and to show that epidemics of Legionnaires' disease had occurred worldwide many years before 1976. Despite a surfeit of available resources and expertise, the effort to isolate the etiologic agent succeeded solely on the basis of one person's determination to solve a scientific problem and his willingness to reexamine his assumptions about prior laboratory results. Pontiac fever, a disease of unknown etiology, is a self-limiting and short duration febrile illness that has been associated with exposure to L. pneumophila. Because of non-specific clinical findings that overlap with other diseases, accurate diagnosis of Pontiac fever in non-outbreak settings is impossible. Legionnaires' disease can be diagnosed specifically through specialized laboratory tests, but not by clinical findings alone. This is because the clinical findings of Legionnaires' disease overlap with those of other more common causes of community acquired pneumonia. Antimicrobial therapy of Legionnaires' disease requires the use of drugs that are active against intracellular Legionellaspp., such as tetracyclines, macrolides, azalides and antibacterial quinolones.

Further reading: Legionella: Molecular Microbiology

Labels: bacteriology, bacterium, legionella

The biochemical capacity to use water as the source for electrons in photosynthesis evolved once, in a common ancestor of extant cyanobacteria. The geological record indicates that this transforming event took place early in our planet's history, at least 2450-2320 million years ago (Ma), and possibly much earlier. Geobiological interpretation of Archean (>2500 Ma) sedimentary rocks remains a challenge; available evidence indicates that life existed 3500 Ma, but the question of when oxygenic photosynthesis evolved continues to engender debate and research. A clear paleontological window on cyanobacterial evolution opened about 2000 Ma, revealing an already diverse biota of blue-greens. Cyanobacteria remained principal primary producers throughout the Proterozoic Eon (2500-543 Ma), in part because the redox structure of the oceans favored photautotrophs capable of nitrogen fixation. Green algae joined blue-greens as major primary producers on continental shelves near the end of the Proterozoic, but only with the Mesozoic (251-65 Ma) radiations of dinoflagellates, coccolithophorids, and diatoms did primary production in marine shelf waters take modern form. Cyanobacteria remain critical to marine ecosystems as primary producers in oceanic gyres, as agents of biological nitrogen fixation, and, in modified form, as the plastids of marine algae.

From: Andrew H. Knoll in The Cyanobacteria: Molecular Biology, Genomics and Evolution

Labels: archaea, bacteriology, bacterium, cyanobacteria

Scientists at the Institute of Food Research, Norwich UK found that probiotic bacteria in a daily drink can modify the immune system's response to grass pollen. Volunteers with a history of seasonal hay fever drank a daily milk drink with or without live Lactobacillus casei over 5 months. The study was double-blinded and placebo controlled, so neither the volunteers nor the scientists knew who had been assigned the probiotic drinks.

Blood samples were taken before the grass pollen season, then again when it was at its peak (June), and 4 weeks after the end of season. There were no significant differences in levels of IgE in the blood between the two groups at the start of the study, but IgE levels were lower in the probiotic group both at the peak season and afterwards. IgE stimulates the release of histamine which produces the symptoms of hayfever.

Further reading: Lactobacillus Probiotics

Labels: bacteriology, biotechnology, lactic acid bacteria, lactobacillus, probiotics

The Quarterly Review of Biology (2008, 83:117) has published a very positive review of a recent book Bacillus: Cellular and Molecular Biology. An extract follows:

"The quality of the chapters is uniformly high. Together they provide a review of significant progress toward a better and deeper understanding of the physical structure and molecular biological organization and function in Bacillus subtilis. As a consequence, a truly intimate grasp of this bacterium is achieved"

Further reading: Bacillus: Cellular and Molecular Biology

Labels: bacillus, bacteriology, bacterium, book review

The Journal of Microbiological Mehods (2008) has published a review of the new Staphylococcus book. An extract is provided below:

Whereas previous books on the molecular biology of the opportunistic human pathogen Staphylococcus aureus used to cover the classical methodologies the timely book by Lindsay et al includes chapters on whole genome sequences, array technologies, the staphylococcal population structure, transciptomics and rapid diagnosis in addition to the more traditional chapters on mobile genetic elements, genetic manipulation, regulation, resistance and environmentally induced responses. This renders the book unique in its kind and as complete as one should expect a modern book on bacterial molecular genetics to be. ...

From the positive perspective: this is a book suitable for both starting and experienced staphylococcal researchers. It is both a referral handbook as explicated by some of the chapters and an adequate introductory text in others. The mixture between "hard-core science" and clinical application also is a balanced one ... the book by Lindsay et al deserves a prominent spot in the personal library of all staphylococcologists!!

Further reading: Staphylococcus: Molecular Genetics

Labels: bacteriology, bacterium, book review, Staphylococcus

from George W. Sundin and Jesús Murillo in Plant Pathogenic Bacteria
The concept of bacterial plasmids as gene traders is illustrative of the role of plasmids in horizontal gene transfer and specifically in the acquisition and distribution of sequences that enable rapid evolution. Plasmids are components of the horizontal gene pool and, as such, their genetic content is potentially accessible by a wide range of organisms. Most plasmids appear to ameliorate any potential negative effect on host fitness by encoding determinants of virulence and ecological fitness that can enhance adaptation to a specific niche or can influence niche expansion. The availability of multiple complete genome sequences of bacterial phytopathogens has shown the importance of horizontally-acquired gene sequences in pathogen evolution. We suspect that plasmids have played a significant role in this gene mobility and also in the delivery of acquired genes to bacterial chromosomes through plasmid integration events. The versatility of plasmids plays a critical role in the evolutionary arms race of bacterial pathogens and plants.

Further reading:

Chapter 14 in Plant Pathogenic Bacteria: Genomics and Molecular Biology

Plasmids: Current Research and Future Trends

Labels: bacteriology, bacterium, plasmids

Vibrio cholerae, the causative agent of cholera belongs to a group of organisms whose natural habitats are the aquatic ecosystems. The strains that cause cholera epidemics have evolved from non-pathogenic progenitor strains by acquisition of virulence genes, and V. cholerae represents a paradigm for this evolutionary process.

Genomics of Vibrio cholerae and its Evolution
The 4.0 Mbp genome of N16961, an O1 serogroup, El Tor biotype, 7th pandemic strain of V. cholerae, is comprised of two circular chromosomes of unequal size that are predicted to encode a total of 3,885 genes. The genomic sequence of this representative strain has facilitated global experimental approaches that have furthered our understanding of the genetic and phenotypic diversity found within the species V. cholerae. Sequence data have been used to identify horizontally acquired sequences, dissect complex regulatory and signaling pathways, and develop computational approaches to predict patterns of gene expression and the presence of metabolic pathway components. In addition, these data have served as a basis for the construction of microarrays to study the evolution of the organism through comparative genomic analyses. Genomic sequencing of additional strains, subtractive hybridization studies and the introduction of new model systems have also contributed to the identification of novel sequences and pathogenic mechanisms associated with other strains. The sequence of strain N16961 has therefore resulted in an expanded view of the genetic repertoire of V. cholerae and focused our attention on the progressive evolution of this marine bacterium that can also be a human pathogen.

Population Genetics of Vibrio cholerae
The influence of evolutionary forces on the genetic diversity of natural populations of living organisms is the subject matter of population genetics. In the case of Vibrio cholerae, data obtained from detailed molecular studies of large populations of these bacteria have allowed for a better understanding of the epidemiology of diseases due to their presence in humans. The species has a high genetic diversity and a complex image of its population structure. There is also evidence of linkage disequilibrium and frequent intragenic and assortative recombination events in their housekeeping genes. Horizontal transfer of genes in V. cholerae is higher than those reported for Escherichia coli and Salmonella enterica. In spite of the frequent horizontal gene transfer, clonal lineages of Vibrio cholerae might persist for decades. The best example of this is the presence and survival of epidemic and pandemic clones over long periods of time. To date, there are four major genetic lines of toxigenic V. cholerae O1 biotype El Tor: an Australian clone (ET 1); the U.S. Gulf Coast clone (ET 2); the seventh pandemic clone isolated in the South East Asia together with the O139 "Bengal" clone (ET 3); and the clone that caused cholera in Latin America in the 1990's (ET 4). There are also isolated clones that have appeared over time under special conditions, e.g., serogroup O37 that was shown to have limited epidemic potential in the 1960's. Given the close evolutionary relationship between V. cholerae O1 and other non-O1 virulent serotypes and the fact that virulence genes can be transferred horizontally, new pathogenic strains of V. cholerae could arise in the future through the modification of existing clones that have the capacity to spread rapidly, and thus cause outbreaks of disease.

Genetics of Vibrio cholerae Colonization and Motility
Survival of Vibrio cholerae either in the aquatic environment or in the human host is mediated by appropriate expression of factors that control motility, colonization, production of virulence factors, as well as sensing the cell density (quorum sensing). Successful transition of the organism between the aquatic and the host intestinal environments thus depends on the coordinated activity of a number of genes and regulatory circuits.

Genetics of O-antigens, Capsules, and the Rugose Variant of Vibrio cholerae
The human pathogen Vibrio cholerae produces three major cell-surface associated polysaccharides, including (i) lipopolysaccharide (LPS), (ii) capsule, and (iii) rugose exopolysaccharide. While LPS and capsule primarily help the bacterium to evade host defense mechanisms, the rugose exopolysaccharide may aid the bacterium in persisting in the nutrient-deficient aquatic environments.

Genetics and Microbiology of Biofilm Formation by Vibrio cholerae
In nature, most bacteria grow as matrix-enclosed, surface-associated communities known as biofilms. Vibrio cholerae, the causative agent of the disease cholera, forms biofilms on diverse surfaces. This ability to form biofilms appears to be critical for the environmental survival and the transmission of V. cholerae. The molecular mechanisms utilized by V. cholerae to form and maintain biofilms have been investigated by molecular genetic and microscopic approaches and these studies should prove useful in the development of future strategies for predicting and controlling cholera epidemics.

Molecular Ecology of Vibrio cholerae
Although Vibrio cholerae causes human disease, aquatic ecosystems are major habitats of V. cholerae, and all V. cholerae are not pathogenic for humans. V. cholerae represents a paradigm for origination of pathogenic bacteria from environmental nonpathogenic progenitor strains by horizontal transfer of genes. Besides environmental factors which are not precisely defined, bacteriophages, and horizontally transmissible genetic elements have a significant role in the epidemiology and evolution of the pathogen. Recent studies are beginning to reveal the mechanisms associated with the occurrence of seasonal epidemics in endemic areas, waterborne spread of cholera, and the factors that enable the organisms to survive unfavorable conditions in the aquatic environment. The emergence of new epidemic strains, and their enrichment during epidemics of cholera appear to constitute a natural system for the evolution of V. cholerae and genetic elements that mediate horizontal transfer of genes among bacterial strains.

Coordinated Regulation of Gene Expression in Vibrio cholerae
Vibrio cholerae, the causative agent of the severe diarrhoeal disease cholera, has evolved with intricate signal transduction and gene regulatory systems to survive and grow under various environmental conditions. The virulence regulon of V. cholerae, which involves multiple genes working in a coordinated manner, represents a regulatory paradigm for extracellular bacterial pathogens. Availability of the whole genome sequence has allowed microarray based transcriptome analyses of V. cholerae cells isolated directly from cholera patients. Such studies indicate that quite a large number of genes are involved in the disease process and their expression pattern changes as the infection progresses. Further understanding of the process came with the recent discoveries of small noncoding RNAs and intracellular signal molecule c-di-GMP as modulators of gene expression in V. cholerae. Transcriptome analysis has also shed light on synchronized gene expression related to chitin utilization and development of natural competence when the organism exists in the natural aquatic environment. Thus, the survival, evolution and pathogenesis of V. cholerae appear to be controlled by several intricate overlapping regulatory circuits.

Evolutionary Relationships of Pathogenic Clones of Vibrio cholerae
Evolution refers to the differentiation of an ancestral genome into recognizably distinct genomes. Understanding the evolutionary history of an organism can provide insight into how it can be expected to evolve in the future and provide predictions that serve as the basis of where to best focus effort to prevent the emergence of new pathogenic variants. In order to accurately understand the evolutionary history, the methods used for interpreting the genetic variation need to reflect the mechanisms of genetic change. The critical mechanism for deciding how to interpret the genetic relatedness is the amount of recombination. If recombination is rare, then the traditional phylogenetic analysis based on bifurcating trees works well. If recombination is common, then a method that incorporates recombination must be used. Evolutionary relationships among pathogenic clones based on these assessments have been presented and discussed.

Emerging Hybrid Variants of Vibrio cholerae O1
Rapid emergence of genetic variants among toxigenic epidemic strains of Vibrio cholerae, contributes to the intricate epidemiological pattern of cholera. A remarkable event in recent years has been the emergence of strains of V. cholerae O1 which possess traits of both the classical and El Tor biotypes.

Antibiotic Resistance in Vibrio cholerae
Antimicrobial resistance has become a major medical and public health problem as it has direct link with the disease management. Vibrio cholerae, the cholera causing pathogen is increasingly developing resistance towards many antimicrobials used for the treatment of diarrhoea. However, the pattern of resistance differs from country to country. The well-known factor responsible for development and spread of resistance is injudicious use of antimicrobial agents, which is directly related to the stimulation of several mechanisms of resistance. In V. cholerae, several resistance mechanisms such as plasmid encoded resistance, mutation in the quinolones resistance determining regions, integrons, efflux pumps and SXT constins have been established. Considering the importance of drug resistance, quick diagnostic assay methods are available for the identification of multidrug resistant (MDR) V. cholerae. Many new generation antimicrobials were discovered, which are effective against V. cholerae in the in vitro studies. The resistance pattern of V. cholerae to several antimicrobials are not always uniform as it depends on the source of isolation. Vibrios can act as reservoirs of antimicrobial resistance as cross-spread is common in in vitro studies. Promotion of indigenous drugs should be considered in the future and studied in detail for their efficacy.

Further reading: Vibrio cholerae: Genomics and Molecular Biology

Labels: antibiotic resistance, bacteriology, bacterium, biofilm, evolution, population genetics, regulation, vibrio