CiteULike: Tag microbiology

The essential Staphylococcus aureus gene fmhB is involved in the first step of peptidoglycan pentaglycine interpeptide formation.

S Rohrer — 2006-11-19T10:44:58-00:00

Proc Natl Acad Sci U S A, Vol. 96, No. 16. (3 August 1999), pp. 9351-9356.

The factor catalyzing the first step in the synthesis of the characteristic pentaglycine interpeptide in Staphylococcus aureus peptidoglycan was found to be encoded by the essential gene fmhB. We have analyzed murein composition and structure synthesized when fmhB expression is reduced. The endogenous fmhB promoter was substituted with the xylose regulon from Staphylococcus xylosus, which allowed glucose-controlled repression of fmhB transcription. Repression of fmhB reduced growth and triggered a drastic accumulation of uncrosslinked, unmodified muropeptide monomer precursors at the expense of the oligomeric fraction, leading to a substantial decrease in overall peptidoglycan crosslinking. The composition of the predominant muropeptide was confirmed by MS to be N-acetylglucosamine-(beta-1,4)-N-acetylmuramic acid(-L-Ala-D-iGln-L-Lys-D-Ala-D-Ala), proving that FmhB is involved in the attachment of the first glycine to the pentaglycine interpeptide. This interpeptide plays an important role in crosslinking and stability of the S. aureus cell wall, acts as an anchor for cell wall-associated proteins, determinants of pathogenicity, and is essential for the expression of methicillin resistance. Any shortening of the pentaglycine side chain reduces or even abolishes methicillin resistance, as occurred with fmhB repression. Because of its key role FmhB is a potential target for novel antibacterial agents that could control the threat of emerging multiresistant S. aureus.

M. tuberculosis and M. leprae Translocate from the Phagolysosome to the Cytosol in Myeloid Cells

Nicole van der Wel — 2007-08-20T16:32:40-00:00

Cell, Vol. 129, No. 7. (29 June 2007), pp. 1287-1298.

Summary M. tuberculosis and M. leprae are considered to be prototypical intracellular pathogens that have evolved strategies to enable growth in the intracellular phagosomes. In contrast, we show that lysosomes rapidly fuse with the virulent M. tuberculosis- and M. leprae-containing phagosomes of human monocyte-derived dendritic cells and macrophages. After 2 days, M. tuberculosis progressively translocates from phagolysosomes into the cytosol in nonapoptotic cells. Cytosolic entry is also observed for M. leprae but not for vaccine strains such as M. bovis BCG or in heat-killed mycobacteria and is dependent upon secretion of the mycobacterial gene products CFP-10 and ESAT-6. The cytosolic bacterial localization and replication are pathogenic features of virulent mycobacteria, causing significant cell death within a week. This may also reveal a mechanism for MHC-based antigen presentation that is lacking in current vaccine strains.

Prediction of transmembrane alpha-helices in prokaryotic membrane proteins: the dense alignment surface method.

M Cserzö — 2008-03-07T06:08:50-00:00

Protein Eng, Vol. 10, No. 6. (June 1997), pp. 673-676.

A new, simple method for predicting transmembrane segments in integral membrane proteins has been developed. It is based on low-stringency dot-plots of the query sequence against a collection of non-homologous membrane proteins using a previously derived scoring matrix [Cserzö et al., 1994, J. Mol. Biol., 243, 388-396]. This so-called dense alignment surface (DAS) method is shown to perform on par with earlier methods that require extra information in the form of multiple sequence alignments or the distribution of positively charged residues outside the transmembrane segments, and thus improves prediction abilities when only single-sequence information is available or for classes of membrane proteins that do not follow the 'positive inside' rule.

PrediSi: prediction of signal peptides and their cleavage positions.

K Hiller — 2008-03-28T19:08:21-00:00

Nucleic Acids Res, Vol. 32, No. Web Server issue. (1 July 2004)

We have developed PrediSi (Prediction of Signal peptides), a new tool for predicting signal peptide sequences and their cleavage positions in bacterial and eukaryotic amino acid sequences. In contrast to previous prediction tools, our new software is especially useful for the analysis of large datasets in real time with high accuracy. PrediSi allows the evaluation of whole proteome datasets, which are currently accumulating as a result of numerous genome projects and proteomics experiments. The method employed is based on a position weight matrix approach improved by a frequency correction which takes in to consideration the amino acid bias present in proteins. The software was trained using sequences extracted from the most recent version of the SwissProt database. PrediSi is accessible via a web interface. An extra Java package was designed for the integration of PrediSi into other software projects. The tool is freely available on the World Wide Web at http://www.predisi.de.

Prediction of lipoprotein signal peptides in Gram-negative bacteria

Agnieszka Juncker — 2008-03-28T19:06:24-00:00

Protein Sci, Vol. 12, No. 8. (1 August 2003), pp. 1652-1662.

A method to predict lipoprotein signal peptides in Gram-negative Eubacteria, LipoP, has been developed. The hidden Markov model (HMM) was able to distinguish between lipoproteins (SPaseII-cleaved proteins), SPaseI-cleaved proteins, cytoplasmic proteins, and transmembrane proteins. This predictor was able to predict 96.8% of the lipoproteins correctly with only 0.3% false positives in a set of SPaseI-cleaved, cytoplasmic, and transmembrane proteins. The results obtained were significantly better than those of previously developed methods. Even though Gram-positive lipoprotein signal peptides differ from Gram-negatives, the HMM was able to identify 92.9% of the lipoproteins included in a Gram-positive test set. A genome search was carried out for 12 Gram-negative genomes and one Gram-positive genome. The results for Escherichia coli K12 were compared with new experimental data, and the predictions by the HMM agree well with the experimentally verified lipoproteins. A neural network-based predictor was developed for comparison, and it gave very similar results. LipoP is available as a Web server at www.cbs.dtu.dk/services/LipoP/. 10.1110/ps.0303703

A Sweet Sensor for Size-Conscious Bacteria

Daisuke Shiomi — 2007-08-02T14:57:38-00:00

Cell, Vol. 130, No. 2. (27 July 2007), pp. 216-218.

Bacteria, like eukaryotic cells, regulate their size by coordinating cell growth and division, growing faster and becoming larger when nutrients are more plentiful. now identify an enzyme in a glucolipid pathway that inhibits assembly of the key cell division protein FtsZ, but only during high nutrient conditions. Delaying cell division during rapid growth allows bacterial cells to become larger.

Metagenomic Analysis of the Human Distal Gut Microbiome

Steven Gill — 2006-06-02T14:00:57-00:00

Science, Vol. 312, No. 5778. (2 June 2006), pp. 1355-1359.

The human intestinal microbiota is composed of 1013 to 1014 microorganisms whose collective genome ("microbiome") contains at least 100 times as many genes as our own genome. We analyzed [~]78 million base pairs of unique DNA sequence and 2062 polymerase chain reaction-amplified 16S ribosomal DNA sequences obtained from the fecal DNAs of two healthy adults. Using metabolic function analyses of identified genes, we compared our human genome with the average content of previously sequenced microbial genomes. Our microbiome has significantly enriched metabolism of glycans, amino acids, and xenobiotics; methanogenesis; and 2-methyl-D-erythritol 4-phosphate pathway-mediated biosynthesis of vitamins and isoprenoids. Thus, humans are superorganisms whose metabolism represents an amalgamation of microbial and human attributes. 10.1126/science.1124234

Environmental Shotgun Sequencing: Its Potential and Challenges for Studying the Hidden World of Microbes

Jonathan Eisen — 2007-03-16T09:31:59-00:00

PLoS Biology, Vol. 5, No. 3. (1 March 2007), e82.

Intestinal Bacteria Trigger T Cell-Independent Immunoglobulin A2 Class Switching by Inducing Epithelial-Cell Secretion of the Cytokine APRIL

Bing He — 2007-06-28T16:56:10-00:00

Immunity, Vol. 26, No. 6. (22 June 2007), pp. 812-826.

Summary Bacteria colonize the intestine shortly after birth and thereafter exert several beneficial functions, including induction of protective immunoglobulin A (IgA) antibodies. The distal intestine contains IgA2, which is more resistant to bacterial proteases than is IgA1. The mechanism by which B cells switch from IgM to IgA2 remains unknown. We found that human intestinal epithelial cells (IECs) triggered IgA2 class switching in B cells, including IgA1-expressing B cells arriving from mucosal follicles, through a CD4+ T cell-independent pathway involving a proliferation-inducing ligand (APRIL). IECs released APRIL after sensing bacteria through Toll-like receptors (TLRs) and further increased APRIL production by activating dendritic cells via thymic stromal lymphopoietin. Our data indicate that bacteria elicit IgA2 class switching by linking lamina propria B cells with IECs through a TLR-inducible signaling program requiring APRIL. Thus, mucosal vaccines should activate IECs to induce more effective IgA2 responses.

The genetics of glycosylation in Gram-negative bacteria

Power — 2008-04-29T09:24:55-00:00

FEMS Microbiology Letters, Vol. 218, No. 2. (2003), pp. 211-222.

Abstract In recent years there has been a dramatic increase in reports of glycosylation of proteins in various Gram-negative systems including Neisseria meningitidis, Neisseria gonorrhoeae, Campylobacter jejuni, Pseudomonas aeruginosa, Escherichia coli, Caulobacter crescentus, Aeromonas caviae and Helicobacter pylori. Although this growing list contains many important pathogens (reviewed by Benz and Schmidt [Mol. Microbiol. 45 (2002) 267-276]) and the glycosylations are found on proteins important in pathogenesis such as pili, adhesins and flagella the precise role(s) of the glycosylation of these proteins remains to be determined. Furthermore, the details of the glycosylation biosynthetic process have not been determined in any of these systems. The definition of the precise role of glycosylation and the mechanism of biosynthesis will be facilitated by a detailed understanding of the genes involved.

A Sanger/pyrosequencing hybrid approach for the generation of high-quality draft assemblies of marine microbial genomes.

Susanne M D Goldberg — 2006-07-17T11:54:16-00:00

Proc Natl Acad Sci U S A (13 July 2006)

Since its introduction a decade ago, whole-genome shotgun sequencing (WGS) has been the main approach for producing cost-effective and high-quality genome sequence data. Until now, the Sanger sequencing technology that has served as a platform for WGS has not been truly challenged by emerging technologies. The recent introduction of the pyrosequencing-based 454 sequencing platform (454 Life Sciences, Branford, CT) offers a very promising sequencing technology alternative for incorporation in WGS. In this study, we evaluated the utility and cost-effectiveness of a hybrid sequencing approach using 3730xl Sanger data and 454 data to generate higher-quality lower-cost assemblies of microbial genomes compared to current Sanger sequencing strategies alone.

Communication modules in bacterial signaling proteins.

JS Parkinson — 2008-02-21T23:01:16-00:00

Annu Rev Genet, Vol. 26 (1992), pp. 71-112.

Two-component signal transduction.

AM Stock — 2008-02-21T23:00:26-00:00

Annu Rev Biochem, Vol. 69 (2000), pp. 183-215.

Most prokaryotic signal-transduction systems and a few eukaryotic pathways use phosphotransfer schemes involving two conserved components, a histidine protein kinase and a response regulator protein. The histidine protein kinase, which is regulated by environmental stimuli, autophosphorylates at a histidine residue, creating a high-energy phosphoryl group that is subsequently transferred to an aspartate residue in the response regulator protein. Phosphorylation induces a conformational change in the regulatory domain that results in activation of an associated domain that effects the response. The basic scheme is highly adaptable, and numerous variations have provided optimization within specific signaling systems. The domains of two-component proteins are modular and can be integrated into proteins and pathways in a variety of ways, but the core structures and activities are maintained. Thus detailed analyses of a relatively small number of representative proteins provide a foundation for understanding this large family of signaling proteins.

Modulated receptor interactions in bacterial transmembrane signaling

Daniel Webre — 2008-02-08T15:17:11-00:00

Trends in Cell Biology, Vol. 14, No. 9. (September 2004), pp. 478-482.

Bacteria can detect and respond to a remarkably diverse set of environmental conditions. This ability enables motile species to integrate stimuli, to compare current surroundings with those of the recent past, and to adjust swimming behavior to move up gradients of attractants and avoid repellents. Many of the molecular details involved in the bacterial chemotaxis system have been elucidated. Several models have been proposed recently to explain how cells process external information through a patch of highly interactive transmembrane receptors and transduce this information to other components in the cytoplasm that, in turn, function to regulate motility.

A Putative [alpha]-Helical Porin from Corynebacterium glutamicum

Karin Ziegler — 2008-07-08T16:10:21-00:00

Journal of Molecular Biology, Vol. 379, No. 3. (6 June 2008), pp. 482-491.

The cell wall of Corynebacterium glutamicum contains a mycolic acid layer, which is a protective nonpolar barrier similar to the outer membrane of Gram-negative bacteria. The exchange of material across this barrier requires porins. Porin B (PorB) is one of them. Recombinant PorB has been produced in Escherichia coli, purified, crystallized and analyzed by X-ray diffraction, yielding 16 independent molecular structures in four different crystal forms at resolutions up to 1.8 Å. All 16 molecules have the same globular core, which consists of 70 residues forming four [alpha]-helices tied together by a disulfide bridge. The 16 structures vary greatly with respect to the 29 residues in the N- and C-terminal extensions. Since corynebacteria belong to the group of mycolata that includes some prominent human pathogens, the observed structure may be of medical relevance. Due to the clearly established solid structure of the core, the native porin has to be oligomeric, and the reported structure is one of the subunits. An [alpha]-helical porin in a bacterial outer envelope is surprising because all presently known structures of such porins consist of [beta]-barrels. Since none of the four crystal packing arrangements was compatible with an oligomeric membrane channel, we constructed a model of such an oligomer that was consistent with all available data of native PorB. The proposed model is based on the required polar interior and nonpolar exterior of the porin, on a recurring crystal packing contact around a 2-fold axis, on the assumption of a simple Cn symmetry (a symmetric arrangement around an n-fold axis), on the experimentally established electric conductivity and anion selectivity and on the generally observed shape of porin channels.

Reconstructing the ancestor of Mycobacterium leprae: The dynamics of gene loss and genome reduction

Laura Gomez-Valero — 2007-08-07T15:38:48-00:00

Genome Res., Vol. 17, No. 8. (1 August 2007), pp. 1178-1185.

We have reconstructed the gene content and order of the last common ancestor of the human pathogens Mycobacterium leprae and Mycobacterium tuberculosis. During the reductive evolution of M. leprae, 1537 of 2977 ancestral genes were lost, among which we found 177 previously unnoticed pseudogenes. We find evidence that a massive gene inactivation took place very recently in the M. leprae lineage, leading to the loss of hundreds of ancestral genes. A large proportion of their nucleotide content ([~]89%) still remains in the genome, which allowed us to characterize and date them. The age of the pseudogenes was computed using a new methodology based on the rates and patterns of substitution in the pseudogenes and functional orthologous genes of closely related genomes. The position of the genes that were lost in the ancestor's genome revealed that the process of function loss and degradation mainly took place through a gene-to-gene inactivation process, followed by the gradual loss of their DNA. This suggests a scenario of massive genome reduction through many nearly simultaneous pseudogenization events, leading to a highly specialized pathogen. 10.1101/gr.6360207

Identification of polymorphonuclear leukocyte and HL-60 cell receptors for adhesins of Streptococcus gordonii and Actinomyces naeslundii.

S Ruhl — 2006-01-09T15:10:59-00:00

Infect Immun, Vol. 68, No. 11. (November 2000), pp. 6346-6354.

Interactions of oral streptococci and actinomyces with polymorphonuclear leukocytes (PMNs), mediated by sialic acid- and Gal/GalNAc-reactive adhesins, respectively, result in activation of the PMNs and thereby may contribute to the initiation of oral inflammation. Sialidase treatment of PMNs or HL-60 cells abolished adhesion of Streptococcus gordonii but was required for adhesion of Actinomyces naeslundii. The same effects of sialidase were noted for adhesion of these bacteria to a major 150-kDa surface glycoprotein of either PMNs or undifferentiated HL-60 cells and to a 130-kDa surface glycoprotein of differentiated HL-60 cells. These glycoproteins were both identified as leukosialin (CD43) by immunoprecipitation with a specific monoclonal antibody (MAb). Adhesion of streptococci and actinomyces to a 200-kDa minor PMN surface glycoprotein was also detected by bacterial overlay of untreated and sialidase-treated nitrocellulose transfers, respectively. This glycoprotein was identified as leukocyte common antigen (CD45) by immunoprecipitation with a specific MAb. CD43 and CD45 both possess extracellular mucinlike domains in addition to intracellular domains that are implicated in signal transduction. Consequently, the interactions of streptococci and actinomyces with the mucinlike domains of these mammalian cell surface glycoproteins result not only in adhesion but, in addition, may represent the initial step in PMN activation by these bacteria.

Advanced sequencing technologies and their wider impact in microbiology

Neil Hall — 2008-01-10T21:05:28-00:00

J Exp Biol, Vol. 210, No. 9. (1 May 2007), pp. 1518-1525.

In the past 10 years, microbiology has undergone a revolution that has been driven by access to cheap high-throughput DNA sequencing. It was not long ago that the cloning and sequencing of a target gene could take months or years, whereas now this entire process has been replaced by a 10 min Internet search of a public genome database. There has been no single innovation that has initiated this rapid technological change; in fact, the core chemistry of DNA sequencing is the same as it was 30 years ago. Instead, progress has been driven by large sequencing centers that have incrementally industrialized the Sanger sequencing method. A side effect of this industrialization is that large-scale sequencing has moved out of small research labs, and the vast majority of sequence data is now generated by large genome centers. Recently, there have been advances in technology that will enable high-throughput genome sequencing to be established in research labs using bench-top instrumentation. These new technologies are already being used to explore the vast microbial diversity in the natural environment and the untapped genetic variation that can occur in bacterial species. It is expected that these powerful new methods will open up new questions to genomic investigation and will also allow high-throughput sequencing to be more than just a discovery exercise but also a routine assay for hypothesis testing. While this review will concentrate on microorganisms, many of the important arguments about the need to measure and understand variation at the species, population and ecosystem level will hold true for many other biological systems. 10.1242/jeb.001370

[Predictive microbiology and risk assessment]

G Hildebrandt — 2008-05-09T06:36:52-00:00

DTW. Deutsche tierärztliche Wochenschrift, Vol. 111, No. 5. (May 2004), pp. 195-200.

Predictive microbiology (predictive modelling PM), in spite of its limits and short-comings, may often contribute to a reduction of the problems arising when HACCP systems are established or microbiological risk assessment is done. Having identified the agents which constitute a risk and the contamination rate and density in the raw material, the influences of production steps and storage on these microorganisms have to be examined. Finally, there should be an exposure assessment, i.e. an estimate of the contamination density in the final product at the time of consumption. Should the exposure assessment together with data from dose response assessments reveal a potential for intake of inacceptable numbers of organisms, the risk identified has to be characterized. As a consequence, risk management should result in a modification of the composition of the product and/or of the production process so that the risk does not surpass an acceptable limit. For this approach it is indispensable to have product- and process-specific information on the multiplication of pathogens prior to heat treatment, on reduction of their density by thermal treatment and on growth or dying of organisms having survived heat treatment or penetrated into the product after heat treatment as post-process contaminant. Commonly, challenge tests are conducted to provide such information. But they are time consuming and, as their results are only valid for the specific product tested and the conditions prevailing during the experiment, the have to be repeated if there is any modification of intrinsic or extrinsic factors. At least partially, the PM may replace the challenge tests. The efficiency of the models is rated particularly high if they are used already at the stage of product development when the question has to be answered whether a planned recipe or process of production are already save or have to be modified to become save.

When mutualists are pathogens: an experimental study of the symbioses between Steinernema (entomopathogenic nematodes) and Xenorhabdus (bacteria).

M Sicard — 2007-01-03T10:34:33-00:00

J Evol Biol, Vol. 17, No. 5. (September 2004), pp. 985-993.

In this paper, we investigate the level of specialization of the symbiotic association between an entomopathogenic nematode (Steinernema carpocapsae) and its mutualistic native bacterium (Xenorhabdus nematophila). We made experimental combinations on an insect host where nematodes were associated with non-native symbionts belonging to the same species as the native symbiont, to the same genus or even to a different genus of bacteria. All non-native strains are mutualistically associated with congeneric entomopathogenic nematode species in nature. We show that some of the non-native bacterial strains are pathogenic for S. carpocapsae. When the phylogenetic relationships between the bacterial strains was evaluated, we found a clear negative correlation between the effect a bacterium has on nematode fitness and its phylogenetic distance to the native bacteria of this nematode. Moreover, only symbionts that were phylogenetically closely related to the native bacterial strain were transmitted. These results suggest that co-evolution between the partners has led to a high level of specialization in this mutualism, which effectively prevents horizontal transmission. The pathogenicity of some non-native bacterial strains against S. carpocapsae could result from the incapacity of the nematode to resist specific virulence factors produced by these bacteria.

The regulation of pathogenicity and mutualism in Photorhabdus.

SA Joyce — 2007-01-03T10:32:51-00:00

Curr Opin Microbiol, Vol. 9, No. 2. (April 2006), pp. 127-132.

Photorhabdus is a genus of insect-pathogenic bacteria that also maintains a mutualistic interaction with Heterorhabditid nematodes. Bacteria in this genus are members of the family Enterobacteriaceae and are, therefore, closely related to many important mammalian pathogens. This bacteria-nematode complex has been exploited as a biocontrol agent that is active against several insect pests. However, this model system is also uniquely placed to address important fundamental questions about pathogenicity and mutualism. Indeed, recent genetic studies have suggested that there is a significant overlap in the genetic requirements of Photorhabdus for these contrasting interactions. In addition, the identification of key regulators of pathogenicity and symbiosis only serves to highlight the similarities between Photorhabdus, a genus of bacteria that infects invertebrate hosts, and closely related mammalian enteric pathogens.

Genomics, environmental genomics and the issue of microbial species

DM Ward — 2007-06-07T09:42:37-00:00

Heredity, Vol. aop, No. current.

A statistical toolbox for metagenomics: assessing functional diversity in microbial communities

Patrick Schloss — 2008-01-24T03:54:24-00:00

BMC Bioinformatics, Vol. 9 (23 January 2008), 34.

Approaches to prokaryotic biodiversity: a population genetics perspective

Francisco Valera — 2008-04-04T12:05:22-00:00

Environmental Microbiology, Vol. 4, No. 11. (2002), pp. 628-633.

Summary The study of prokaryotic diversity has blossomed during the last 10-15 years as a result of the introduction of molecular identification, mostly based on direct 16S rRNA gene polymerase chain reaction (PCR) amplification and sequencing from natural samples. A large amount of information exists about the diversity of this specific gene. However, data from the field of bacterial population genetics and genomics make questionable the value of information regarding just one gene. Even if we accept 16S rRNA genes as useful for species identification, intraspecific variation in bacteria is so high that species catalogues are often of little value. The gene pools represented by an operational species are yet impossible to predict. On the other hand, adaptive features in prokaryotes are often coded in gene clusters (genomic islands) that can be cloned directly from the environment, sequenced and even expressed in a surrogate host. Thus, the study of the environmental genome or metagenome appears as an alternative that could eventually lead to a more realistic understanding of prokaryotic biodiversity, provide biotechnology with new tools and maybe even contribute to develop a model of prokaryotic evolution.

Interpreting the unculturable majority

Lior Pachter — 2007-06-01T00:55:14-00:00

Nature Methods, Vol. 4, No. 6., pp. 479-480.

Genomic sequencing of single microbial cells from environmental samples.

Thomas Ishoey — 2008-06-18T22:46:58-00:00

Current opinion in microbiology (10 June 2008)

Recently developed techniques allow genomic DNA sequencing from single microbial cells [Lasken RS: Single-cell genomic sequencing using multiple displacement amplification. Curr Opin Microbiol 2007, 10:510-516]. Here, we focus on research strategies for putting these methods into practice in the laboratory setting. An immediate consequence of single-cell sequencing is that it provides an alternative to culturing organisms as a prerequisite for genomic sequencing. The microgram amounts of DNA required as template are amplified from a single bacterium by a method called multiple displacement amplification (MDA) avoiding the need to grow cells. The ability to sequence DNA from individual cells will likely have an immense impact on microbiology considering the vast numbers of novel organisms, which have been inaccessible unless culture-independent methods could be used. However, special approaches have been necessary to work with amplified DNA. MDA may not recover the entire genome from the single copy present in most bacteria. Also, some sequence rearrangements can occur during the DNA amplification reaction. Over the past two years many research groups have begun to use MDA, and some practical approaches to single-cell sequencing have been developed. We review the consensus that is emerging on optimum methods, reliability of amplified template, and the proper interpretation of 'composite' genomes which result from the necessity of combining data from several single-cell MDA reactions in order to complete the assembly. Preferred laboratory methods are considered on the basis of experience at several large sequencing centers where >70% of genomes are now often recovered from single cells. Methods are reviewed for preparation of bacterial fractions from environmental samples, single-cell isolation, DNA amplification by MDA, and DNA sequencing.

The Microbial Engines That Drive Earth's Biogeochemical Cycles

Paul Falkowski — 2008-05-23T16:18:23-00:00

Science, Vol. 320, No. 5879. (23 May 2008), pp. 1034-1039.

Virtually all nonequilibrium electron transfers on Earth are driven by a set of nanobiological machines composed largely of multimeric protein complexes associated with a small number of prosthetic groups. These machines evolved exclusively in microbes early in our planet's history yet, despite their antiquity, are highly conserved. Hence, although there is enormous genetic diversity in nature, there remains a relatively stable set of core genes coding for the major redox reactions essential for life and biogeochemical cycles. These genes created and coevolved with biogeochemical cycles and were passed from microbe to microbe primarily by horizontal gene transfer. A major challenge in the coming decades is to understand how these machines evolved, how they work, and the processes that control their activity on both molecular and planetary scales. 10.1126/science.1153213

Antimicrobial effect of crude extracts of Nigella sativa on multiple antibiotics-resistant bacteria.

NM Morsi — 2004-11-27T07:00:23-00:00

Acta Microbiol Pol, Vol. 49, No. 1. (2000), pp. 63-74.

Different crude extracts of Nigella sativa were tested for antimicrobial effectiveness against different bacterial isolates. These isolates comprised 16 gramnegative and 6 grampositive representatives. They showed multiple resistance against antibiotics, specially the gramnegative ones. Crude extracts of Nigella saliva showed a promising effect against some of the test organisms. The most effective extracts were the crude alkaloid and water extracts. Gramnegative isolates were affected more than the grampositive ones.

Quantitative Phylogenetic Assessment of Microbial Communities in Diverse Environments

C von Mering — 2007-02-26T09:46:57-00:00

Science, Vol. 315, No. 5815. (23 February 2007), pp. 1126-1130.

The taxonomic composition of environmental communities is an important indicator of their ecology and function. We used a set of protein-coding marker genes, extracted from large-scale environmental shotgun sequencing data, to provide a more direct, quantitative, and accurate picture of community composition than that provided by traditional ribosomal RNA-based approaches depending on the polymerase chain reaction. Mapping marker genes from four diverse environmental data sets onto a reference species phylogeny shows that certain communities evolve faster than others. The method also enables determination of preferred habitats for entire microbial clades and provides evidence that such habitat preferences are often remarkably stable over time. 10.1126/science.1133420

The universal ancestor and the ancestors of Archaea and Bacteria were anaerobes whereas the ancestor of the Eukarya domain was an aerobe

Di — 2007-02-19T11:08:30-00:00

Journal of Evolutionary Biology, Vol. 20, No. 2. (2007), pp. 543-548.

Abstract The use of an oxyphobic index (OI) based on the propensity of amino acids to enter more frequently the proteins of anaerobes makes it possible to make inferences on the environment in which the last universal common ancestor (LUCA) lived. The reconstruction of the ancestral sequences of proteins using a method based on maximum likelihood and their attribution by means of the OI to the set of aerobe or anaerobe sequences has led to the following conclusions: the LUCA was an anaerobic 'organism', as were the ancestors of Archaea and Bacteria, whereas the ancestor of Eukarya was an aerobe. These observations seem to falsify the hypothesis that the LUCA was an aerobe and help to identify better the environment in which the first organisms lived.

Identification of novel non-coding RNAs as potential antisense regulators in the archaeon Sulfolobus solfataricus

Thean-Hock Tang — 2005-01-13T07:48:04-00:00

Molecular Microbiology, Vol. 55, No. 2., 469.

Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs

Lee Lim — 2005-01-31T21:29:42-00:00

Nature, Vol. aop, No. current. (30 January 2005)

Synergy and contingency as driving forces for the evolution of multiple secondary metabolite production by Streptomyces species.

GL Challis — 2004-12-02T17:15:28-00:00

Proc Natl Acad Sci U S A, Vol. 100 Suppl 2 (25 November 2003), pp. 14555-14561.

In this article we briefly review theories about the ecological roles of microbial secondary metabolites and discuss the prevalence of multiple secondary metabolite production by strains of Streptomyces, highlighting results from analysis of the recently sequenced Streptomyces coelicolor and Streptomyces avermitilis genomes. We address this question: Why is multiple secondary metabolite production in Streptomyces species so commonplace? We argue that synergy or contingency in the action of individual metabolites against biological competitors may, in some cases, be a powerful driving force for the evolution of multiple secondary metabolite production. This argument is illustrated with examples of the coproduction of synergistically acting antibiotics and contingently acting siderophores: two well-known classes of secondary metabolite. We focus, in particular, on the coproduction of beta-lactam antibiotics and beta-lactamase inhibitors, the coproduction of type A and type B streptogramins, and the coregulated production and independent uptake of structurally distinct siderophores by species of Streptomyces. Possible mechanisms for the evolution of multiple synergistic and contingent metabolite production in Streptomyces species are discussed. It is concluded that the production by Streptomyces species of two or more secondary metabolites that act synergistically or contingently against biological competitors may be far more common than has previously been recognized, and that synergy and contingency may be common driving forces for the evolution of multiple secondary metabolite production by these sessile saprophytes.

Acinetobacter sp. ADP1: an ideal model organism for genetic analysis and genome engineering.

D Metzgar — 2005-05-01T17:55:30-00:00

Nucleic Acids Res, Vol. 32, No. 19. (2004), pp. 5780-5790.

Acinetobacter sp. strain ADP1 is a naturally transformable gram-negative bacterium with simple culture requirements, a prototrophic metabolism and a compact genome of 3.7 Mb which has recently been sequenced. Wild-type ADP1 can be genetically manipulated by the direct addition of linear DNA constructs to log-phase cultures. This makes it an ideal organism for the automation of complex strain construction. Here, we demonstrate the flexibility and versatility of ADP1 as a genetic model through the construction of a broad variety of mutants. These include marked and unmarked insertions and deletions, complementary replacements, chromosomal expression tags and complex combinations thereof. In the process of these constructions, we demonstrate that ADP1 can effectively express a wide variety of foreign genes including antibiotic resistance cassettes, essential metabolic genes, negatively selectable catabolic genes and even intact operons from highly divergent bacteria. All of the described mutations were achieved by the same process of splicing PCR, direct transformation of growing cultures and plating on selective media. The simplicity of these tools make genetic analysis and engineering with Acinetobacter ADP1 accessible to laboratories with minimal microbial genetics expertise and very little equipment. They are also compatible with complete automation of genetic analysis and engineering protocols.

Microbial analysis at the single-cell level: tasks and techniques

Howard Shapiro — 2006-05-19T16:43:04-00:00

Journal of Microbiological Methods, Vol. 42, No. 1. (September 2000), pp. 3-16.

The heterogeneity of microorganisms themselves is orders of magnitude greater than the heterogeneity of perspectives from which they are contemplated by human observers. Even closely related species may exhibit marked differences in biochemistry and behavior, and, under many conditions, similar, striking heterogeneity may exist within a clonal population of organisms which, in the aggregate, occupy too small a region of space to be visible to the unaided human eye. Using methods of microscopy, microspectrophotometry, and cytometry developed and refined since the 1960s, it is now possible to characterize the physiology and pharmacology of individual microorganisms, and, in many cases, to isolate organisms with selected characteristics for culture and/or further analysis. These methods include fluorescent and confocal microscopy, scanning and image cytometry, and flow cytometry. Fluorescence measurements are particularly important in single-cell analysis; they allow demonstration and quantification of cells’ nucleic acid content and sequence, of the presence of specific antigens, and of physiologic characteristics such as enzyme activity and membrane potential. Multiparameter cytometry, combined with cell sorting, provides insight into population heterogeneity and allows selected cells to be separated for further analysis and culture. The technology is applicable to a wide range of problems in contemporary microbiology, including strain selection and the development of antimicrobial agents.

Single-cell microbiology: tools, technologies, and applications.

BF Brehm-Stecher — 2005-03-16T17:14:49-00:00

Microbiol Mol Biol Rev, Vol. 68, No. 3. (September 2004)

The field of microbiology has traditionally been concerned with and focused on studies at the population level. Information on how cells respond to their environment, interact with each other, or undergo complex processes such as cellular differentiation or gene expression has been obtained mostly by inference from population-level data. Individual microorganisms, even those in supposedly "clonal" populations, may differ widely from each other in terms of their genetic composition, physiology, biochemistry, or behavior. This genetic and phenotypic heterogeneity has important practical consequences for a number of human interests, including antibiotic or biocide resistance, the productivity and stability of industrial fermentations, the efficacy of food preservatives, and the potential of pathogens to cause disease. New appreciation of the importance of cellular heterogeneity, coupled with recent advances in technology, has driven the development of new tools and techniques for the study of individual microbial cells. Because observations made at the single-cell level are not subject to the "averaging" effects characteristic of bulk-phase, population-level methods, they offer the unique capacity to observe discrete microbiological phenomena unavailable using traditional approaches. As a result, scientists have been able to characterize microorganisms, their activities, and their interactions at unprecedented levels of detail.

Evaluation of molecular methods used for establishing the interactions and functions of microorganisms in anaerobic bioreactors

G Talbot — 2008-01-28T17:39:28-00:00

Water Research, Vol. 42, No. 3. (February 2008), pp. 513-537.

Molecular techniques have unveiled the complexity of the microbial consortium in anaerobic bioreactors and revealed the presence of several uncultivated species. This paper presents a review of the panoply of classical and recent molecular approaches and multivariate analyses that have been, or might be used to establish the interactions and functions of these anaerobic microorganisms. Most of the molecular approaches used so far are based on the analysis of small subunit ribosomal RNA but recent studies also use quantification of functional gene expressions. There are now several studies that have developed quantitative real-time PCR assays to investigate methanogens. With a view to improving the stability and performance of bioreactors, monitoring with molecular methods is also discussed. Advances in metagenomics and proteomics will lead to the development of promising lab-on chip technologies for cost-effective monitoring.

Towards an evolutionary ecology of life in soil

John Crawford — 2008-02-26T18:32:16-00:00

Trends in Ecology & Evolution, Vol. 20, No. 2. (February 2005), pp. 81-87.

The soil-microbe system is one of the most diverse components of the terrestrial ecosystem. The origin of this diversity, and its relation to the life-sustaining processes that are mediated by the resident microbial community, is still poorly understood. The inherent complexities necessitate a theoretical framework that integrates ecological and evolutionary approaches and which embraces the physical heterogeneity of the soil environment. Such a framework is currently lacking, although recent advances in theory and experimentation are beginning to identify the essential ingredients. Here, we review and evaluate the relevance of current modelling approaches, and propose a new synthesis of an evolutionary ecology of life in soil. Key elements include an account of dispersal, horizontal gene transfer, and the consideration of the physical and biological components of soil as an integrated complex adaptive system.

In situ detection of protein-hydrolysing microorganisms in activated sludge.

Y Xia — 2008-02-04T13:36:18-00:00

FEMS Microbiol Ecol, Vol. 60, No. 1. (April 2007), pp. 156-165.

Protein hydrolysis plays an important role in the transformation of organic matter in activated sludge wastewater treatment plants, but no information is currently available regarding the identity and ecophysiology of protein-hydrolysing organisms (PHOs). In this study, fluorescence in situ enzyme staining with casein and bovine serum albumin conjugated with BODIPY dye was applied and optimized to label PHOs in activated sludge plants. A strong fluorescent labeling of the surface of microorganisms expressing protease activity was achieved. Metabolic inhibitors were applied to inhibit the metabolic activity to prevent uptake of the fluorescent hydrolysates by oligopeptide-consuming bacteria. In five full-scale, nutrient-removing activated sludge plants examined, the dominant PHOs were always different morphotypes of filamentous bacteria and the epiflora attached to many of these. The PHOs were identified by FISH using a range of available oligonucleotide probes. The filamentous PHOs belonged to the candidate phylum TM7, the phylum Chloroflexi and the class Betaproteobacteria. In total they comprised 1-5% of the bacterial biovolume. Most of the epiflora-PHOs hybridized with probe SAP-309 targeting Saprospiraceae in the phylum Bacteroidetes and accounted for 8-12% of the total bacterial biovolume in most plants and were thus an important and dominant part of the microbial communities.

Carbon fluxes within the pelagic food web in the coastal area off Antofagasta (23[degree sign]S), Chile: The significance of the microbial versus classical food webs

Hector Paves — 2007-11-30T11:01:21-00:00

Ecological Modelling, Vol. In Press, Corrected Proof

Trophic interactions and the relevance of the "classical" (CFW) versus the "microbial" (MFW) food webs were studied in the upwelling system of Antofagasta (23[degree sign]S), northern Humboldt Current System (HCS) off Chile. Biological and ecological data gathered from the study area during 1996 and 1999-2002 and complementary data from the literature were analysed using the Ecopath with Ecosim software version 5.0 (EwE). The model includes the following functional groups: Detritus, dissolved organic matter (DOM), bacteria, phytoplankton, appendicularians, salps, calanoid copepods, cyclopoid copepods, chaetognaths, ctenophores, clupeiform fishes. The Antofagasta's model indicate that only 18% of the total trophic relationships up to clupeiforms include only organisms of the CFW, while it is increased up to 82% when trophic relationships with organisms of the MFW are included. Removing the MFW from a complex, multi-level, trophic pathways, the energy transfer efficiency that characterise the study area either decreased or increased their value from detritus (from 14.4% to 13.6%) and primary production (from 11.0% to 15.1%), respectively. The total net primary production (PP) of the system with and without the MFW is similar (2.3 grC m-2 d-1 and 2.2 grC m-2 d-1), however, the total energy throughput increased in a system with MFW from ~4.7 grC m-2 d-1 to ~9.8 grC m-2 d-1. Thus, although the energy transfer efficiency is higher in a system without the MFW, the total system throughput is two times higher when it is considered. This might be due to (1) the inclusion of large new compartments in different trophic levels (such as DOM, bacteria), (2) the inclusion of the MFW would enhance the recycling of materials and energy and (3) the inclusion of new pathways such as bacteria - appendicularians/salps - anchovy (clupeiforms), that might produce a close link between the base of the microbial loop and clupeiforms, bypassing most of the MFW. The exclusion of the microbial loop from trophic models should strongly affect different planktonic groups such as salps, small cyclopoid copepods and appendicularians because they feed mainly (or partially, such as euphausiids) on small-size particles such as bacteria, microflagellates and ciliates. Thus, both CFW and MFW constitute highly complementary and interconnected trophic pathways from microbes up to fishes, and where more resolution at lower trophic level would be required to derive sensible model results that satisfactorily represent the system behaviour (i.e. ecosystem approach).

Effects of particulate and soluble substrates on microfauna populations and treatment efficiency in activated sludge systems

Jaume Puigagut — 2008-02-04T13:24:38-00:00

Water Research, Vol. 41, No. 14. (July 2007), pp. 3168-3176.

To determine the effects of particulate and soluble compounds on microfauna populations and treatment efficiency in activated sludge systems, two experimental wastewater treatment plants were set up and evaluated for a period of five months. The plants were fed with pre-flocculated domestic sewage enriched with starch or glucose as model substrates of particulate and soluble organic matter, respectively. It was observed that the starch-enriched system presented lower abundance of filamentous bacteria that turned into a better sludge sedimentation. Mean sludge volume index (SVI) values for the starch and glucose-enriched systems were 54+/-24 and 885+/-845 mL g-1, respectively. Although no differences in organic matter removal were detected between the systems, nitrification and denitrification were higher in the starch-enriched system, which is likely to have been the result of its more compact flocs. The mean ammonia-N effluent concentrations for the starch and glucose-enriched systems were 4.7+/-5.7 and 16.2+/-9.7 mg L-1, respectively, whereas the nitrate-N concentrations were 20.1+/-10.8 and 30.8+/-12.2 mg L-1, respectively. Concerning microfauna analysis, ciliated protozoa--specifically, attached ciliates--were the dominant microfauna group in both treatment systems, whereas metazoa, particularly Lecanidae rotifera, were more abundant in the starch-enriched system. Lecanidae rotifera abundances above 400 ind mL-1 reduced the mean floc area from 60 to 20 mm2 without affecting sludge settleability in the starch-enriched system. Finally, the reduction in floc area caused by metazoa feeding led to effluents of lower nitrogen quality, although no differences in sludge production were detected.

Microbial biosensors

Yu Lei — 2008-02-11T11:28:02-00:00

Analytica Chimica Acta, Vol. 568, No. 1-2. (24 May 2006), pp. 200-210.

A microbial biosensor is an analytical device that couples microorganisms with a transducer to enable rapid, accurate and sensitive detection of target analytes in fields as diverse as medicine, environmental monitoring, defense, food processing and safety. The earlier microbial biosensors used the respiratory and metabolic functions of the microorganisms to detect a substance that is either a substrate or an inhibitor of these processes. Recently, genetically engineered microorganisms based on fusing of the lux, gfp or lacZ gene reporters to an inducible gene promoter have been widely applied to assay toxicity and bioavailability. This paper reviews the recent trends in the development and application of microbial biosensors. Current advances and prospective future direction in developing microbial biosensor have also been discussed.

Nitrogen removal performance and microbial community structure dynamics response to carbon nitrogen ratio in a compact suspended carrier biofilm reactor

Siqing Xia — 2008-02-14T16:05:49-00:00

Ecological Engineering, Vol. 32, No. 3. (3 March 2008), pp. 256-262.

A compact suspended carrier biofilm reactor (SCBR) was developed for simultaneous nitrification and denitrification (SND) in a single reactor and the performance of nutrient removal was investigated. Microbial community structure response to different ratio of carbon to nitrogen (C/N) was determined by denaturing gel gradient electrophoresis (DGGE) profiles of 16S rDNA V3 region and amoA gene amplifications. In addition, the population dynamics of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) were estimated by fluorescence in situ hybridization (FISH) with 16S rDNA-targeted oligonucleotide probes. Results showed that the compact SCBR was efficient in nutrient removal with CODCr removal efficiency over 90% and SND efficiency (ESND) about 83.3%. The diversity of microbial community structure was positively correlated with C/N ratio, while the three communities of amoA gene were relativity homogenous. The population of nitrifiers was in inverse proportions to C/N ratio with the average fraction of AOB and NOB to all bacteria 5.4, 4.8, 3.1% and 4.6, 3.5, 2.7% respectively as C/N ratio changing from 3:1, 5:1 to 10:1. Therefore we could reach a conclusion that the compact SCBR was practical to treat municipal wastewater and the shift of microbial community monitored by molecular technologies could offer guidance to the process optimization in engineering.

Biological nitrogen and phosphorus removal and changes in microbial community structure in a membrane bioreactor: Effect of different carbon sources

Zubair Ahmed — 2007-12-31T10:39:05-00:00

Water Research, Vol. 42, No. 1-2. (January 2008), pp. 198-210.

Bacterial community structures in four sequencing anoxic/anaerobic-aerobic membrane bioreactors (SAMs) that were fed with synthetic medium composed of different organic compounds in substrate as carbon source; acetate-dominant (acetate/propionate=4/1), propionate-dominant (acetate/propionate=1/4), glucose-dominant (glucose/acetate=4/1) and methanol-dominant (methanol/acetate/propionate=6/3/1) were analyzed by respiratory quinone profile and fluorescent in situ hybridization (FISH) techniques. The SAMs were operated at controlled pH range 7-8.5 and at constant temperature 25 [degree sign]C. Total nitrogen (TN), total phosphorus (TP) and COD removal performances were also evaluated and compared. In addition, trans-membrane pressure was monitored to observe the impact of substrate composition on membrane fouling. The dominance of the mole fraction of ubiquinone (UQ-8) in the SAMs indicated dominance of the [beta]-subclass of Proteobacteria; however, its population comparatively decreased when the substrate was glucose dominant or methanol dominant. A relatively higher and stable enhanced biological phosphorus removal performance was observed when methanol-dominant substrate was used concurrently with an increase in the [gamma]-subclass of Proteobacteria. The population of the [alpha]-subclass of Proteobacteria slightly increased along with a decrease in phosphate removal activity when the substrate was glucose-dominant. Results from FISH analysis also supported the findings of the quinone profile. The trans-membrane pressure variation in the SAMs indicated that fouling was relatively rapid when propionate-dominant or methanol-dominant substrate was used and most stable when glucose-dominant substrate was used. A combination of methanol and acetate would be a better choice as an external carbon source when nutrients removals, as well as fouling, are considered in the membrane bioreactor- (MBR-) coupled biological nutrients removing (BNR) process.

Cellulase digestibility of pretreated biomass is limited by cellulose accessibility

Tina Jeoh — 2007-07-27T10:48:11-00:00

Biotechnology and Bioengineering, Vol. 98, No. 1. (2007), pp. 112-122.

Attempts to correlate the physical and chemical properties of biomass to its susceptibility to enzyme digestion are often inconclusive or contradictory depending on variables such as the type of substrate, the pretreatment conditions and measurement techniques. In this study, we present a direct method for measuring the key factors governing cellulose digestibility in a biomass sample by directly probing cellulase binding and activity using a purified cellobiohydrolase (Cel7A) from Trichoderma reesei. Fluorescence-labeled T. reesei Cel7A was used to assay pretreated corn stover samples and pure cellulosic substrates to identify barriers to accessibility by this important component of cellulase preparations. The results showed cellulose conversion improved when T. reesei Cel7A bound in higher concentrations, indicating that the enzyme had greater access to the substrate. Factors such as the pretreatment severity, drying after pretreatment, and cellulose crystallinity were found to directly impact enzyme accessibility. This study provides direct evidence to support the notion that the best pretreatment schemes for rendering biomass more digestible to cellobiohydrolase enzymes are those that improve access to the cellulose in biomass cell walls, as well as those able to reduce the crystallinity of cell wall cellulose. Biotechnol. Bioeng. 2007; 98: 112-122. © 2007 Wiley Periodicals, Inc.

Bacteria Subsisting on Antibiotics

Gautam Dantas — 2008-04-04T07:31:31-00:00

Science, Vol. 320, No. 5872. (4 April 2008), pp. 100-103.

Antibiotics are a crucial line of defense against bacterial infections. Nevertheless, several antibiotics are natural products of microorganisms that have as yet poorly appreciated ecological roles in the wider environment. We isolated hundreds of soil bacteria with the capacity to grow on antibiotics as a sole carbon source. Of 18 antibiotics tested, representing eight major classes of natural and synthetic origin, 13 to 17 supported the growth of clonal bacteria from each of 11 diverse soils. Bacteria subsisting on antibiotics are surprisingly phylogenetically diverse, and many are closely related to human pathogens. Furthermore, each antibiotic-consuming isolate was resistant to multiple antibiotics at clinically relevant concentrations. This phenomenon suggests that this unappreciated reservoir of antibiotic-resistance determinants can contribute to the increasing levels of multiple antibiotic resistance in pathogenic bacteria. 10.1126/science.1155157

Theory and the microbial world

Curtis — 2007-01-09T12:47:20-00:00

Environmental Microbiology, Vol. 9, No. 1. (January 2007), pp. 1-1.

Residence time and food contact time effects on transfer of Salmonella Typhimurium from tile, wood and carpet: testing the five-second rule

Dawson — 2007-03-24T18:57:17-00:00

Journal of Applied Microbiology, Vol. 102, No. 4. (April 2007), pp. 945-953.

Increased subaerial volcanism and the rise of atmospheric oxygen 2.5 billion years ago

Lee Kump — 2007-08-30T16:31:32-00:00

Nature, Vol. 448, No. 7157., pp. 1033-1036.

Recent advances in sea-ice microbiology

Thomas Mock — 2005-04-09T20:43:14-00:00

Environmental Microbiology, Vol. 7, No. 5. (May 2005), pp. 605-619.

Over the past 50 years there has been much effort invested in the investigation of the ecology of sea ice. Sea ice is an ephemeral feature of the Arctic and Southern Oceans and smaller water bodies such as the Baltic and Caspian Seas. The semisolid ice matrix provides a range of habitats in which a diverse range of microbial organisms thrive. In the past 5 years there has been considerable steps forward in sea-ice research, in particular regarding the analysis of sea-ice microstructure and the investigation of the diversity and adaptation of microbial communities. These studies include: (i) controlled simulated and _in situ_ studies on a micrometer scale to unravel the dynamic of the microhabitat with consequences for the organisms; (ii) the introduction of molecular approaches to uncover the diversity of uncultured still unknown microorganisms; and (iii) studies into the molecular adaptation of selected model organisms to the extreme environment. This minireview presents some of the most recent findings from sea-ice studies within the framework of these aims.