Microbiology Blog: The weblog for microbiologists.

A huge variety of biopolymers, such as polysaccharides, polyesters, and polyamides, are naturally produced by microorganisms. These range from viscous solutions to plastics and their physical properties are dependent on the composition and molecular weight of the polymer. The genetic manipulation of microorganisms opens up an enormous potential for the biotechnological production of biopolymers with tailored properties suitable for high-value medical application such as tissue engineering and drug delivery.

Microorganisms naturally produce a wide variety of carbohydrate molecules, yet large-scale manufacturing requires production levels much higher than the natural capacities of these organisms. Metabolic engineering efforts generate microbial strains capable of meeting the industrial demand for high synthesis levels. As both oligosaccharide and polysaccharide synthesis are carbon and energy-intensive processes, improved production of these products require similar metabolic engineering strategies. Metabolically engineered strains have successfully produced many carbohydrate products and many unexplored strategies made available from recent progress in systems biology can be used to engineer better microbial catalysts.

Further reading: Microbial Production of Biopolymers and Polymer Precursors

Labels: biopolymers, biotechnology, genetic engineering

Whatever the route used, horizontal transfer of DNA requires elaborated multi-protein machinery to enable the long and charged nucleic acid polymer to cross the cell envelope barriers. The best-studied system for cell-to-cell DNA translocation is bacterial conjugation. This system can be divided in two discrete specialized modules: the relaxosome, which triggers and takes part in plasmid DNA processing and replication, and a type IV secretion system (T4SS), which impels protein and single-stranded DNA through the membranes. In addition, a coupling protein (CP), linking both modules, and a number of ancillary proteins are needed. Over the last decades research efforts in the field have resulted in the clarification of many aspects of this system and its machinery assembly. In particular, structural biology has provided details of the molecular architecture of several of the pieces involved in this intricate scenario.

Further reading: Plasmids: Current Research and Future Trends

Labels: conjugation, DNA, plasmids

Pasteurellaceae comprise a large and diverse family of Gram-negative bacteria with members ranging from important pathogens such as Haemophilus influenzae to commensals of the animal and human mucosa. Information on the biology of these organisms has mushroomed in recent years, driven by the development of novel genetic and molecular methodologies. Since 1995, the family has been expanded from three genera to the current thirteen through the use of new genetic-based classification and identification technologies. Many members of the Pasteurellaceae family make excellent natural models for the study of bacterial pathogenesis and host-pathogen-interactions thus giving valuable insights into related human diseases. Research in this area is at a very exciting stage.

Further reading: Pasteurellaceae: Biology, Genomics and Molecular Aspects

Labels: Haemophilus, model organism, pasteurella, Pasteurellaceae

A number of published reviews on Real-Time PCR have recently been made available as "open-access" papers. These timely and authoritative reviews are written by experts in the field and can be downloaded at: PCR Papers


Further info: PCR Papers

Labels: PCR, real-time PCR

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

In recent years the development of new molecular biology tools and the elucidation of whole genome sequences have revolutionized research in medical mycology. Such advances have led to the development of faster, more reliable diagnostic techniques for medically important pathogens such as Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans. In addition they have led to a major breakthrough in the approach for the generation of novel anti-fungal agents.

In a recent book on Medical Mycology a panel of expert international mycologists critically review the most important areas. Topics include: gene expression and regulation, heterozygosity in Candida, molecular diagnosis, regulation of the host-fungal interaction, the development of anti-fungals, signal transduction, and mechanisms of multi-drug resistance.

Further reading: Pathogenic Fungi: Insights in Molecular Biology

Analysis of the three completed Helicobacter pylori genomes has confirmed the presence of five major outer membrane proteins (OMPs) families. However, there appears to be a trend within the protein families as to which two of the three orthologs are more closely related. H. pylori 26695 and HPAG1 proteins are the most closely related more often as H. pylori J99 and HPAG1 or H. pylori J99 and 26695. Whether this is because both HPAG1 and 26695 were isolated from gastritis patients and J99 from a duodenal ulcer patient is an intriguing, but untested possibility. It has been demonstrated that several OMPs in the largest family act as adhesions, and these include BabA, SabA, OipA, AlpAB and HopZ. This unusual set of specialized OMPs may be a reflection of the adaptation of H. pylori to the unique gastric environment where it is found. Many of these genes undergo phase variation such that not all strains will produce functional proteins, and the stability of expression with passage varied with OipA > BabA > BabB > SabA. Each OMP appears to have specific functions.

Further reading: Helicobacter pylori

Interest in the microbial biodegradation of pollutants has intensified in recent years as mankind endeavors to find sustainable ways to cleanup contaminated environments. Uta Breuer (Leipzig) has written a review (Eng. Life Sci. 2008: 8, 81-82) of a recent book on this topic:

"This book is very clearly written with an abundance of information on Microbial Biodegradation from the point of view of a microbial molecular biologist. Therefore this book is strongly recommended for scientists working in the field of microbial degradation and bioremediation and certainly of general interest for environmental microbiologists. It should be made available in all libraries at universities, research institutes and industry, and is further strongly recommended to all those who are interested in life sciences."

Further information: Microbial Biodegradation: Genomics and Molecular Biology

Labels: book review

Molecular diagnostic methods are currently being used for the early detection of many viral, bacterial, parasitic and fungal infections. Due to their high specificity and sensitivity, these methods will be inserted in the routine of the clinical laboratories to complement information provided by more conventional methods and above all, to help in the diagnosis of dubious cases. The design of specific probes, coupled to the Polymerase Chain Reaction (PCR) technique provides the required specificity and sensitivity to identify fungal species in a short time. Multicopy genes such as those of ribosomal origin are preferred by many researchers to achieve well defined results. However, because they may generate false positive results, other researchers have looked at single-copy genes of high specificity. This technology is being applied to the identification of frequently found causative agents of systemic mycoses such as Candida albicans, Aspergillus fumigatus and other species within these genera, or agents of less frequent mycoses produced by Coccidioides immitis, Cryptococcus neoformans, Histoplasma capsulatum, Paracoccidioides brasiliensis, among others.

Further reading: Pathogenic Fungi: Insights in Molecular Biology

Labels: diagnosis, fungi, mycology, PCR, real-time PCR

from Future Virol. (2008) 3(2): 119-123.

"The book entitled Coronaviruses: Molecular and Cellular Biology presents a comprehensive review of the field of coronavirus virology. The contributors to the book are many of the experts in coronavirus virology. The contents of the book are particularly useful for those wishing to conduct research on coronaviruses because most of the authors include in their summary or conclusion paragraphs a number of key questions or problems for which research needs to be carried out to further the understanding of coronaviruses and how to treat the infections they cause. Most of the figures are very useful in helping the reader understand the narratives provided by the authors. The few tables provided are excellent in summarizing information ...

recommended for those who want to quickly be brought up to date on coronavirus research or who want to know what research questions remain to be answered in the field of coronavirus virology."

Further reading: Coronaviruses: Molecular and Cellular Biology

Labels: book review, virology, virus