Molecular Marine Microbiology Chapter Abstracts
CHAPTER 1.
Introduction
Douglas H. Bartlett
The following thirteen chapters introduce some of the exciting milestones being covered in molecular marine microbiology today, and in many cases, their biomedical or biotechnological relevance. The issues covered include quorum and anti-quorum sensing, selected symbioses, DNA shuffling in the environment, the evolution and development of novel motility mechanisms, hydrocarbon degradation, the interactions of microbes with metals, and adaptations to extremes of pressure and temperature. Although most of these topics are not exclusive to marine realms, they all have as a common thread a profound relevance to the ocean environment.
One of the great contributions of marine microbiology to all of microbiology has been the discovery of the importance of acylated homoserine lactones in cell density or "quorum" sensing by bacteria. The paradigm for this mode of regulation, the control of bioluminescence in Vibrio fischeri, is described by Paul Dunlap. An entirely new light on V. fischeri is provided by Ned Ruby, who describes how this microorganism, along with a squid host with which it has established a mutualistic association, is providing new clues to bacterial colonization of animal epithelial tissue. A fascinating twist on quorum sensing is presented by Rice et al. who describe the synthesis of an inhibitor of bacterial quorum sensing systems by a species of red algae. The inhibitor appears to represent a chemical strategy used by the algae to limit bacterial colonization of its surfaces. Additional aspects of bioactive metabolites involved in the symbiotic interactions of marine microorganisms is described by Haygood et al. who describe potential biomedical applications of microbes obtained from marine invertebrates.
Of course microbes take in much more than cell-cell signaling molecules. Many prokaryotes can also take in DNA, and thus re-tailor their genetic identies. John Paul describes gene transfer among indiginous microbes in the environment and the significance of transformation, conjugation and transduction to phylogenetics. Additional microbial processing reactions which are of biotechnological relevance concern the applications of marine prokaryotes to environmental clean up and nano-scale biofabrication. Harayama et al. describe the fate of various petroleum components in seawater, and the significance of Alcanivorax in marine bioremediation. Francis and Tebo explore the details of manganese oxidation, progressing from genes to enzyme kinetics and applications to toxic metal removal. Finally, Dirk Schüler describes the bacterial manufacturing of magnetic iron containing organelles (magnetosomes) within certain benthic bacteria and their biotechnological promise in products ranging from magnetic tapes to magnetic resonance imaging devices.
Two chapters deal with mysterious motility mechanisms operating in marine microbes. Linda McCarter introduces surface-regulated phenomena, including swarming motility via lateral flagella, in the human pathogen and marine resident, V. parahaemolyticus. Bianca Brahamsha details the only known case of liquid motility in the absence of flagella, a phenotype displayed by many marine Synechococcus species.
The book end with descriptions of the adaptations of certain deep-sea prokaryotes to extremes of pressure and temperature. Kato and Qureshi present a taxomonic overview of deep-sea bacteria and genetic and biophysical analyses of respiratory chain adaptations to elevated pressure. My article describes genetic evidence for the role of additional membrane components in pressure-sensing and adaptation, the discovery of a DNA recombination enzyme which is also required for life at low volume change, and the potential of deep-sea bacteria as a source of genes for the biosynthesis of polyunsaturated fatty acids. Finally, Robb and Clark present a discussion of the biochemical bases of thermostability in proteins from hyperthermophiles, most of which are members of the domain Archaea.
Although this admittedly ecclectic collection of articles is hardly representative of all of molecular marine microbiology (for example, no articles dealing with viruses or protists are included), the book provides a valuable glimpse into many of the exciting developments taking place in this broad field. Readers will find the book useful both as a reference source and as an introduction to topics yet to be explored.
CHAPTER 2.
Quorum Regulation of Luminescence in Vibrio fischeri
Paul V. Dunlap
Luminescence in Vibrio fischeri is controlled by a population density-responsive regulatory mechanism called quorum sensing. Elements of the mechanism include: LuxI, an acyl-homoserine lactone (acyl-HSL) synthase that directs synthesis of the diffusible signal molecule, 3-oxo-hexanoyl-HSL (V. fischeri auto-inducer-1, VAI-1); LuxR, a transcriptional activator protein necessary for response to VAI-1; GroEL, which is necessary for production of active LuxR; and AinS, an acyl-HSL synthase that catalyzes the synthesis of octanoyl-HSL (VAI-2). The population density-dependent accumulation of VAI-1 triggers induction of lux operon (luxICDABEG; genes for luminescence enzymes and for LuxI) transcription and luminescence by binding to LuxR, forming a complex that facilitates the association of RNA polymerase with the lux operon promoter. VAI-2, which apparently interfers with VAI-1 binding to LuxR, operates to limit premature lux operon induction. Hierarchical control is imposed on the system by 3':5'-cyclic AMP (cAMP) and cAMP receptor protein (CRP), which are necessary for activated expression of luxR. Several non-lux genes in V. fischeri are controlled by LuxR and VAI-1. Quorum regulation in V. fischeri serves as a model for LuxI/LuxR-type quorum sensing systems in other Gram-negative bacteria.
CHAPTER 3.
The Euprymna scolopes -Vibrio fischeri Symbiosis: A Biomedical Model for the Study of Bacterial Colonization of Animal Tissue
Edward G. Ruby
The diversity of microorganisms found in the marine environment reflects the immense size, range of physical conditions and energy sources, and evolutionary age of the sea. Because associations with living animal tissue are an important and ancient part of the ecology of many microorganisms, it is not surprising that the study of marine symbioses (including both cooperative and pathogenic interactions) has produced numerous discoveries of biotechnological and biomedical significance. The association between the bioluminescent bacterium Vibrio fischeri and the sepiolid squid Euprymna scolopes has emerged as a productive model system for the investigation of the mechanisms by which cooperative bacteria initiate colonization of specific host tissues. The results of the last decade of research on this system have begun to reveal surprising similarities between this association and the pathogenic associations of disease-causing Vibrio species, including those of interest to human health and aquaculture. Studies of the biochemical and molecular events underlying the development of the squid-vibrio symbiosis can be expected to continue to increase our understanding of the factors controlling both benign and pathogenic bacterial associations.
CHAPTER 4.
Bacterial Signals and Antagonists: The Interaction Between Bacteria and Higher Organisms
Scott A. Rice, Michael Givskov, Peter Steinberg, and Staffan Kjelleberg
It is now well established that bacteria communicate through the secretion and uptake of small diffusable molecules. These chemical cues, or signals, are often used by bacteria to coordinate phenotypic expression and this mechanism of regulation presumably provides them with a competitive advantage in their natural environment. Examples of coordinated behaviors of marine bacteria which are regulated by signals include swarming and exoprotease production, which are important for niche colonisation or nutrient acquisition (e.g. protease breakdown of substrate). While the current focus on bacterial signalling centers on N-Acylated homoserine lactones, the quorum sensing signals of some Gram-negative bacteria, these are not the only types of signals used by bacteria. Indeed, there appears to be many other types of signals produced by bacteria and it also appears that a bacterium may use multiple classes of signals for phenotypic regulation. Recent work in the area of marine microbial ecology has led to the observation that some marine eukaryotes secrete their own signals which compete with the bacterial signals and thus inhibit the expression of bacterial signalling phenotypes. This type of molecular mimicry has been well characterised for the interaction of marine prokaryotes with the red alga, Delisea pulchra.
CHAPTER 5.
Microbial Symbionts of Marine Invertebrates: Opportunities for Microbial Biotechnology
Margo G. Haygood, Eric W. Schmidt, Seana K. Davidson, and D. John Faulkner
Marine invertebrates are sources of a diverse array of bioactive metabolites with great potential for development as drugs and research tools. In many cases, microorganisms are known or suspected to be the biosynthetic source of marine invertebrate natural products. The application of molecular microbiology to the study of these relationships will contribute to basic biological knowledge and facilitate biotechnol-ogical development of these valuable resources. The bryostatin-producing bryozoan B. neritina and its specific symbiont "Candidatus Endobugula sertula" constitute one promising model system. Another fertile subject for investigation is the listhistid sponges that contain numerous bioactive metabolites, some of which originate from bacterial symbionts.
CHAPTER 6.
Microbial Gene Transfer: An Ecological Perspective
John H. Paul
Microbial gene transfer or microbial sex is a means of exchanging loci amongst prokaryotes and certain eukaryotes. Historically viewed as a laboratory artifact, recent evidence from natural populations as well as genome research has indicated that this process may be a major driving force in microbial evolution. Studies with natural populations have taken two approaches-either adding a defined donor with a traceable gene to an indigenous community, and detecting the target gene in the indigenous bacteria, or by adding a model recipient to capture genes being transferred from the ambient microbial flora. However, both approaches usually require some cultivation of the recipient, which may result in a dramatic underestimation of the ambient transfer frequency. Novel methods are just evolving to study in situ gene transfer processes, including the use of green fluorescent protein (GFP)-marked plasmids, which enable detection of transferrants by epifluorescence microscopy. A transduction-like mechanism of transfer from viral-like particles produced by marine bacteria and thermal spring bacteria to Escherichia coli has been documented recently, indicating that broad host range transduction may be occurring in aquatic environments. The sequencing of complete microbial genomes has shown that they are a mosaic of ancestral chromosomal genes interspersed with recently transferred operons that encode peripheral functions. Archaeal genomes indicate that the genes for replication, transcription, and translation are all eukaryotic in complexity, while the genes for intermediary metabolism are purely bacterial. And in eukaryotes, many ancestral eukaryotic genes have been replaced by bacterial genes believed derived from food sources. Collectively these results indicate that microbial sex can result in the dispersal of loci in contemporary microbial populations as well as having shaped the phylogenies of microbes from multiple, very early gene transfer events.
CHAPTER 7.
Surface Induced Gene Expression in Vibrio parahaemolyticus
Linda McCarter
Vibrio parahaemolyticus is a ubiquitous marine bacterium and human pathogen. The organism possesses multiple cell types appropriate for life under different circumstances. The swimmer cell, with a single polar flagellum, is adapted to life in liquid environments. The polar flagellum is powered by the sodium motive force and can propel the bacterium at fast speeds. The swarmer cell, propelled by many proton-powered lateral flagella, can move through highly viscous environments, colonize surfaces, and form multicellular communities which sometimes display highly periodic architecture. Signals that induce differentiation to the surface-adapted cell type are both physical and chemical in nature. Surface-induced gene expression may aid survival, whether attached to inanimate surfaces or in a host organism. Genetic rearrangements create additional phenotypic versatility for the organism and is manifested as variable opaque and translucent colony morphotypes. Discovery that a LuxR homolog controls the opaque cell type implicates intercellular signaling as an additional survival strategy. The alternating identities of V. parahaemolyticus may play important roles in attachment and detachment, how bacterial populations adapt to growth on surfaces, form structured communities, and develop biofilms.
CHAPTER 8.
Non-Flagellar Swimming in Marine Synechococcus
B. Brahamsha
Certain marine unicellular cyanobacteria of the genus Synechococcus exhibit a unique type of swimming motility characterized by the absence of flagella and of any other obvious organelle of motility. Although the mechanism responsible for this phenomenon remains mysterious, recent advances have included the development of testable models as well as the identification of a cell-surface polypeptide that is required for the generation of thrust. These developments, as well as the future research directions they suggest, are discussed.
CHAPTER 9.
Petroleum Biodegradation in Marine Environments
Shigeaki Harayama, Hideo Kishira, Yuki Kasai and Kazuaki Shutsubo
Petroleum-based products are the major source of energy for industry and daily life. Petroleum is also the raw material for many chemical products such as plastics, paints, and cosmetics. The transport of petroleum across the world is frequent, and the amounts of petroleum stocks in developed countries are enormous. Consequently, the potential for oil spills is significant, and research on the fate of petroleum in a marine environment is important to evaluate the environmental threat of oil spills, and to develop biotechnology to cope with them.
Crude oil is constituted from thousands of components which are separated into saturates, aromatics, resins and asphaltenes. Upon discharge into the sea, crude oil is subjected to weathering, the process caused by the combined effects of physical, chemical and biological modification. Saturates, especially those of smaller molecular weight, are readily biodegraded in marine environments. Aromatics with one, two or three aromatic rings are also efficiently biodegraded; however, those with four or more aromatic ring are quite resistant to biodegradation. The asphaltene and resin fractions contain higher molecular weight compounds whose chemical structures have not yet been resolved. The biodegradability of these compounds is not yet known.
It is known that the concentrations of available nitrogen and phosphorus in seawater limit the growth and activities of hydrocarbon-degrading microorganisms in a marine environment. In other words, the addition of nitrogen and phosphorus fertilizers to an oil-contaminated marine environment can stimulate the biodegradation of spilled oil. This notion was confirmed in the large-scale operation for bioremediation after the oil spill from the Exxon Valdez in Alaska.
Many microorganisms capable of degrading petroleum components have been isolated. However, few of them seem to be important for petroleum biodegradation in natural environments. One group of bacteria belonging to the genus Alcanivorax does become predominant in an oil-contaminated marine environment, especially when nitrogen and phosphorus fertilizers are added to stimulate the growth of endogenous microorganisms.
CHAPTER 10.
Marine Bacillus Spores as Catalysts for Oxidative Precipitation and Sorption of Metals
Chris A. Francis and Bradley M. Tebo
The oxidation of soluble manganese(II) to insoluble Mn(III,IV) oxide precipitates plays an important role in the environment. These Mn oxides are known to oxidize numerous organic and inorganic compounds, scavenge a variety of other metals on their highly charged surfaces, and serve as electron acceptors for anaerobic respiration. Although the oxidation of Mn(II) in most environments is believed to be bacterially-mediated, the underlying mechanisms of catalysis are not well understood. In recent years, however, the application of molecular biological approaches has provided new insights into these mechanisms. Genes involved in Mn oxidation were first identified in our model organism, the marine Bacillus sp. strain SG-1, and subsequently have been identified in two other phylogenetically distinct organisms, Leptothrix discophora and Pseudomonas putida. In all three cases, enzymes related to multicopper oxidases appear to be involved, suggesting that copper may play a universal role in Mn(II) oxidation. In addition to catalyzing an environmentally important process, organisms capable of Mn(II) oxidation are potential candidates for the removal, detoxification, and recovery of metals from the environment. The Mn(II)-oxidizing spores of the marine Bacillus sp. strain SG-1 show particular promise, due to their inherent physically tough nature and unique capacity to bind and oxidatively precipitate metals without having to sustain growth.
CHAPTER 11.
Magnetosome Formation in Magnetotactic Bacteria
Dirk Schüler
The ability of magnetotactic bacteria to orient and migrate along geomagnetic field lines is based on intracellular magnetic structures, the magnetosomes, which comprise nano-sized, membrane bound crystals of magnetic iron minerals. The formation of magnetosomes is achieved by a biological mechanism that controls the accumulation of iron and the biomineralization of magnetic crystals with a characteristic size and morphology within membrane vesicles. This paper focuses on the current knowledge about magnetotactic bacteria and will outline aspects of the physiology and molecular biology of magnetosome formation. The biotechnological potential of the biomineralization process is discussed.
CHAPTER 12.
Pressure Response in Deep-Sea Piezophilic Bacteria
Chiaki Kato, Mohammad Hassan Qureshi, and Koki Horikoshi
Several piezophilic bacteria have been isolated from deep-sea environments under high hydrostatic pressure. Taxonomic studies of the isolates showed that the piezophilic bacteria are not widely distributed in terms of taxonomic positions, and all were assigned to particular branches of the Proteobacteria gamma-subgroup. A pressure-regulated operon from piezophilic bacteria of the genus Shewanella, S. benthica and S. violacea, was cloned and sequenced, and downstream of this operon another pressure regulated operon, cydD-C, was found. The cydD gene was found to be essential for the bacterial growth under high-pressure conditions, and the product of this gene was found to play a role in their respiratory system. Results obtained later indicated that the respiratory system in piezophilic bacteria may be important for survival in a high-pressure environment, and more studies focusing on other components of the respiratory chain have been conducted. These studies suggested that piezophilic bacteria are capable of changing their respiratory system in response to pressure conditions, and a proposed respiratory chain model has been suggested in this regard.
CHAPTER 13.
Microbial Adaptations to the Psychrosphere/Piezosphere
Douglas H. Bartlett
Low temperature and high pressure deep-sea environments occupy the largest fraction of the biosphere. Nevertheless, the molecular adaptations that enable life to exist under these conditions remain poorly understood. This article will provide an overview of the current picture on high pressure adaptation in cold oceanic environments, with an emphasis on genetic experiments performed on Photobacterium profundum. Thus far genes which have been found or implicated as important for pressure-sensing or pressure-adaptation include genes required for fatty acid unsaturation, the membrane protein genes toxR and rseC and the DNA recombination gene recD. Many deep-sea bacteria possess genes for the production of omega-3 polyunsaturated fatty acids. These could be of biotechnological significance since these fatty acids reduce the risk of cardiovascular disease and certain cancers and are useful as dietary supplements.
CHAPTER 14.
Adaptation of Proteins from Hyperthermophiles to Extreme Pressure and Temperature
Frank T. Robb and Douglas S. Clark
Further clarification of the adaptations permitting the persistence of life at temperatures above 100 °C depends in part on the analysis of adaptive mechanisms at the protein level. The hypertherm-ophiles include both Bacteria and Archaea, although the majority of isolates growing at or above 100 °C are Archaea. Newly described adaptive features of hyperthermophiles include proteins whose structural integrity persists at temperatures up to 200 °C, and under elevated hydrostatic pressure, which in some cases adds significant increments of stability.
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