Microbial diversity of deep-sea extremophiles--Piezophiles, Hyperthermophiles, and subsurface microorganisms]

Knowledge of our Planet's biosphere has increased tremendously during the last 10 to 20 years. In the field of Microbiology in particular, scientists have discovered novel "extremophiles", microorganisms capable of living in extreme environments such as highly acidic or alkaline conditions, at high salt concentration, with no oxygen, extreme temperatures (as low as -20 degrees C and as high as 300 degrees C), at high concentrations of heavy metals and in high pressure environments such as the deep-sea. It is apparent that microorganisms can exist in any extreme environment of the Earth, yet already scientists have started to look for life on other planets; the so-called "Exobiology" project. But as yet we have little knowledge of the deep-sea and subsurface biosphere of our own planet. We believe that we should elucidate the Biodiversity of Earth more thoroughly before exploring life on other planets, and these attempts would provide deeper insight into clarifying the existence of extraterrestrial life. We focused on two deep-sea extremophiles in this article; one is "Piezophiles", and another is "Hyperthermophiles". Piezophiles are typical microorganisms adapted to high-pressure and cold temperature environments, and located in deep-sea bottom. Otherwise, hyperthermophiles are living in high temperature environment, and located at around the hydrothermal vent systems in deep-sea. They are not typical deep-sea microorganisms, but they can grow well at high-pressure condition, just like piezophiles. Deming and Baross mentioned that most of the hyperthermophilic archaea isolated from deep-sea hydrothermal vents are able to grow under conditions of high temperature and pressure, and in most cases their optimal pressure for growth was greater than the environmental pressure they were isolated from. It is possible that originally their native environment may have been deeper than the sea floor and that there had to be a deeper biosphere. This implication suggests that the deep-sea hydrothermal vents are the windows to a deep subsurface biosphere. A vast array of chemoautotrophic deep-sea animal communities have been found to exist in cold seep environments, and most of these animals are common with those found in hydrothermal vent environments. Thus, it is possible to consider that the cold seeps are also one of slit windows to a deep subsurface biosphere. We conclude that the deep-sea extremophiles are very closely related into the unseen majority in subsurface biosphere, and the subsurface biosphere probably concerns to consider the "exobiology".     

Reproduction in marine invertebrates in “stable” environments: the deep sea model

Summary

Studies over the last 15 years have revealed that deep-sea benthic megainvertebrates show a variety of reproductive patterns that are adapted to the deep-sea, an environment in which the fauna occurs at low densities and resources are sparse. In the NE Atlantic the majority of species reproduce year round whilst a limited number of species reproduce on a seasonal basis believed to be entrained by the deposition of surface derived organic material on the deep-sea bed. A third pattern of rapid growth and early reproduction is found in a limited number of species that utilize unpredictable and ephemeral resources in the deep sea. Examination of the fertilization and behavioural biology of species from the bathyal depths suggest some species enhance fertilization success by forming pairs during their breeding season. However, the same concentration of sperm, as seen in shallow water invertebrates, is required for successful fertilization. At least one deep-sea species of echinoid requires high pressure for successful embryogenesis suggesting a depth-related segregation of deep-sea fauna. The origin of megafaunal populations of deep-sea invertebrates in the N. Atlantic is discussed in the light of these new data in relation to varying reproductive patterns and the environmental changes that have occurred during the last deglaciation.     

Piezophilic adaptation: a genomic point of view

Two-thirds of Earth's surface is covered by oceans, yet the study of this massive integrated living system is still in its infancy. Various environmental variables, such as high salinity, low and changeable nutrient availability and depth-correlated gradients of light, temperature, nutrients and pressure shape the diversity, physiology and ecology of marine species. As oceans present an average depth of 3800 m, deep-sea ecosystems represent the most common marine ecological niche. One of the key environment variables that influences the life and evolution of deep-sea organisms is high pressure. This extreme widespread condition requires specific adaptations, the nature of which remains largely unknown. Recent advances in genomic approaches, such as in sequencing technologies and global expression profiling, are rapidly increasing the data available to understand microbial evolution, biochemistry, physiology and diversity. This review summarises the analysis of the results published so far about microbial high pressure adaptation from a genomic point of view. Understanding high pressure adaptation mechanisms is not just a scientific exercise but has important biotechnological implications. For example, hydrostatic pressure is a reality for food science and technology, both for food preparation and preservation. An understanding of the effects of pressure on biomolecules will expand its use in the medical, industrial and biotechnological fields.     

深海真菌多样性及其代谢产物生物活性的研究进展

深海环境复杂多样,深海微生物逐渐进化以适应其生存环境。真菌作为深海环境中的重要微生物类群,是开发海洋生物的新兴资源。综述了近年来深海真菌的物种多样性、活性代谢产物的多样性及其生物学功能等的研究进展,并对未来深海真菌的应用进行了展望。     

A study of deep-sea natural microbial populations and barophilic pure cultures using a high-pressure chemostat

Continuous cultures in which a high-pressure chemostat was used were employed to study the growth responses of (i) deep-sea microbial populations with the naturally occurring carbon available in seawater and with limiting concentrations of supplemental organic substrates and (ii) pure cultures of copiotrophic barophilic and barotolerant deep-sea isolates in the presence of limiting carbon concentrations at various pressures, dilution rates, and temperatures. We found that the growth rates of natural populations could not be measured or were extremely low (e.g., a doubling time of 629 h), as determined from the difference between the dilution rate and the washout rate. A low concentration of supplemental carbon (0.33 mg/liter) resulted in positive growth responses in the natural population, which resulted in an increase in the number of cells and eventually a steady population of cells. We found that the growth responses to imposed growth pressure by barophilic and barotolerant pure-culture isolates that were previously isolated and characterized under high-nutrient-concentration conditions were maintained under the low-nutrient-concentration limiting conditions (0.33 to 3.33 mg of C per liter) characteristic of the deep-sea environment. Our results indicate that deep-sea microbes can respond to small changes in substrate availability. Also, barophilic microbes that are copiotrophic as determined by their isolation in the presence of high carbon concentrations and their preference for high carbon concentrations are versatile and are able to compete and grow as barophiles in the low-carbon-concentration oligotrophic deep-sea environment in which they normally exist.     

A Study of Deep-Sea Natural Microbial Populations and Barophilic Pure Cultures Using a High-Pressure Chemostat

Continuous cultures in which a high-pressure chemostat was used were employed to study the growth responses of (i) deep-sea microbial populations with the naturally occurring carbon available in seawater and with limiting concentrations of supplemental organic substrates and (ii) pure cultures of copiotrophic barophilic and barotolerant deep-sea isolates in the presence of limiting carbon concentrations at various pressures, dilution rates, and temperatures. We found that the growth rates of natural populations could not be measured or were extremely low (e.g., a doubling time of 629 h), as determined from the difference between the dilution rate and the washout rate. A low concentration of supplemental carbon (0.33 mg/liter) resulted in positive growth responses in the natural population, which resulted in an increase in the number of cells and eventually a steady population of cells. We found that the growth responses to imposed growth pressure by barophilic and barotolerant pure-culture isolates that were previously isolated and characterized under high-nutrient-concentration conditions were maintained under the low-nutrient-concentration limiting conditions (0.33 to 3.33 mg of C per liter) characteristic of the deep-sea environment. Our results indicate that deep-sea microbes can respond to small changes in substrate availability. Also, barophilic microbes that are copiotrophic as determined by their isolation in the presence of high carbon concentrations and their preference for high carbon concentrations are versatile and are able to compete and grow as barophiles in the low-carbon-concentration oligotrophic deep-sea environment in which they normally exist.     

Deep-sea hydrothermal vent Epsilonproteobacteria encode a conserved and widespread nitrate reduction pathway (Nap)

Despite the frequent isolation of nitrate-respiring Epsilonproteobacteria from deep-sea hydrothermal vents, the genes coding for the nitrate reduction pathway in these organisms have not been investigated in depth. In this study we have shown that the gene cluster coding for the periplasmic nitrate reductase complex (nap) is highly conserved in chemolithoautotrophic, nitrate-reducing Epsilonproteobacteria from deep-sea hydrothermal vents. Furthermore, we have shown that the napA gene is expressed in pure cultures of vent Epsilonproteobacteria and it is highly conserved in microbial communities collected from deep-sea vents characterized by different temperature and redox regimes. The diversity of nitrate-reducing Epsilonproteobacteria was found to be higher in moderate temperature, diffuse flow vents than in high temperature black smokers or in low temperatures, substrate-associated communities. As NapA has a high affinity for nitrate compared with the membrane-bound enzyme, its occurrence in vent Epsilonproteobacteria may represent an adaptation of these organisms to the low nitrate concentrations typically found in vent fluids. Taken together, our findings indicate that nitrate reduction is widespread in vent Epsilonproteobacteria and provide insight on alternative energy metabolism in vent microorganisms. The occurrence of the nap cluster in vent, commensal and pathogenic Epsilonproteobacteria suggests that the ability of these bacteria to respire nitrate is important in habitats as different as the deep-sea vents and the human body.     

Cell biology of deep-sea multicellular organisms

Establishing tissue cultures derived from deep-sea multicellular organisms has been extremely difficult because of the serious damage they sustain upon decompression and exposure to the high temperature of surface seawater. We developed a novel pressure-stat aquarium system for the study of living deep-sea multicellular organisms under pressure. Using this system, we have succeeded in maintaining a variety of deep-sea multicellular organisms under pressure and atmospheric conditions after gradual, slow decompression. Furthermore, we successfully cultivated and freeze-stocked pectoral fin cells of the deep-sea eel Simenchelys parasiticus collected at a depth of 1,162 m under atmospheric pressure conditions. This review describes novel capture and maintenance devices for deep-sea organisms and cell culture studies of the organisms under atmospheric and pressure conditions.     

Free-living tube worm endosymbionts found at deep-sea vents

Recent evidence suggests that deep-sea vestimentiferan tube worms acquire their endosymbiotic bacteria from the environment each generation; thus, free-living symbionts should exist. Here, free-living tube worm symbiont phylotypes were detected in vent seawater and in biofilms at multiple deep-sea vent habitats by PCR amplification, DNA sequence analysis, and fluorescence in situ hybridization. These findings support environmental transmission as a means of symbiont acquisition for deep-sea tube worms.     

Pressure effects on<Emphasis Type="Italic"> Clostridium</Emphasis> strains isolated from a cold deep-sea environment

Three Clostridium strains were isolated from deep-sea sediments collected at a depth of 6.3–7.3 km in the Japan Trench. Physiological characterization and 16S rDNA analysis revealed that the three isolates were all closely related to Clostridium bifermentans. The spores of all three isolates were resistant to inactivation at high pressure and low temperature. However, despite the fact that the vegetative cells were halotolerant and eurythermal they did not appear to be adapted for growth or viability under the conditions prevailing in the deep-sea sediments from which they were obtained. The results suggest that the isolates had survived as spores in the deep-sea sediments and that the marine benthos could be a source of clostridia originating in other environments.Communicated by K. Horikoshi     

Ancient Origin of the Modern Deep-Sea Fauna

The origin and possible antiquity of the spectacularly diverse modern deep-sea fauna has been debated since the beginning of deep-sea research in the mid-nineteenth century. Recent hypotheses, based on biogeographic patterns and molecular clock estimates, support a latest Mesozoic or early Cenozoic date for the origin of key groups of the present deep-sea fauna (echinoids, octopods). This relatively young age is consistent with hypotheses that argue for extensive extinction during Jurassic and Cretaceous Oceanic Anoxic Events (OAEs) and the mid-Cenozoic cooling of deep-water masses, implying repeated re-colonization by immigration of taxa from shallow-water habitats. Here we report on a well-preserved echinoderm assemblage from deep-sea (1000–1500 m paleodepth) sediments of the NE-Atlantic of Early Cretaceous age (114 Ma). The assemblage is strikingly similar to that of extant bathyal echinoderm communities in composition, including families and genera found exclusively in modern deep-sea habitats. A number of taxa found in the assemblage have no fossil record at shelf depths postdating the assemblage, which precludes the possibility of deep-sea recolonization from shallow habitats following episodic extinction at least for those groups. Our discovery provides the first key fossil evidence that a significant part of the modern deep-sea fauna is considerably older than previously assumed. As a consequence, most major paleoceanographic events had far less impact on the diversity of deep-sea faunas than has been implied. It also suggests that deep-sea biota are more resilient to extinction events than shallow-water forms, and that the unusual deep-sea environment, indeed, provides evolutionary stability which is very rarely punctuated on macroevolutionary time scales.     

Frequency and characteristics of plasmids in bacteria isolated from deep-sea amphipods

Bacterial strains isolated from deep-sea amphipods were identified, classified, and screened for plasmid content. Plasmids were common, with 11 of 16 isolates carrying one or more plasmids; these ranged in size from 2.9 to 63 megadaltons. Several of the strains demonstrated distinctly different phenotypic traits yet contained plasmids of the same molecular weight. Results of agarose gel electrophoresis, DNA hybridization, and restriction analysis indicate that the plasmids detected in these deep-sea isolates are identical, suggesting that transmission may occur in the deep-sea environment and that plasmids are common in some deep-sea habitats.     

Piezotolerance of the respiratory terminal oxidase activity of the piezophilic Shewanella violacea DSS12 as compared with non-piezophilic Shewanella species

The facultative piezophile Shewanella violacea DSS12 is known to alter its respiratory components under the influence of hydrostatic pressure during growth, suggesting that it has a respiratory system that functions in adaptation to high pressure. We investigated the pressure- and temperature-dependencies of the respiratory terminal oxidase activity of the membrane of S. violacea relative to non-piezophilic Shewanella species. We observed that the activity in the membrane of S. violacea was more resistant to high pressure than those of non-piezophilic Shewanella even though DSS12 was cultured under atmospheric pressure. On the other hand, the temperature dependency of this activity was almost the same for all of the tested strain regardless of optimal growth temperature. Both high pressure and low temperature are expected to lower protein flexibility, causing a decrease in enzyme activity, but the results of this study suggest that the mechanism maintaining enzyme activity under high hydrostatic pressure is different from that at low temperature. Additionally, the responses of the activity to the pressure- and temperature-changes were independent of membrane lipid composition. Therefore, the piezotolerance of the respiratory terminal oxidases of S. violacea is perhaps dependent on the properties of the protein itself and not on the lipid composition of the membrane. Our observations suggest that S. violacea constitutively express piezotolerant respiratory terminal oxidases that serve adaptation to the deep-sea environment.     

Taxonomy and biotransformation activities of some deep-sea actinomycetes

Deep-sea soft sediments from trench systems and depths in the northwestern Pacific Ocean ranging from less than 300 to 10 897 m in depth have been analyzed for three target genera of actinomycetes: Micromonospora, Rhodococcus, and Streptomyces. Only culturable strains, recovered at atmospheric pressure on selective isolation media, have been examined to date. Maximum recoveries of culturable bacteria were greater that 10⁷/ml wet g sediment, but actinomycetes comprised a small proportion of this population (usually less than 1%). The target actinomycetes were isolated at all depths except from the Mariana Trench sediments. Actinomycete colonies were defined initially on the basis of colony morphologies, and preliminary identification then was made by chemotaxonomic tests. Pyrolysis mass spectrometry (PyMS) of deep-sea mycolic acid-containing actinomycetes gave excellent correspondence with numerical (phenetic) taxonomic analyses and subsequently was adopted as a rapid procedure for assessing taxonomic diversity. PyMS analysis enabled several clusters of deep-sea rhodococci to be distinguished that are quite distinct from all type strains. 16S rRNA gene sequence analysis has revealed that several of these marine rhodococci have sequences that are very similar to certain terrestrial species of Rhodococcus and to Dietzia. There is evidence for the intrusion of terrestrial runoff into these deep trench systems, and the inconsistency of the phenotypic and molecular taxonomies may reflect recent speciation events in actinomycetes under the high-pressure conditions of the deep sea. The results of DNA-DNA pairing experiments point to the novelty of Rhodococcus strains recovered from hadal depths in the Izu Bonin Trench. Biotransformation studies of deep-sea bacteria have focused on nitrile compounds. Nitrile-metabolizing bacteria, closely related to rhodococci, have been isolated that grow well at low temperature, high salt concentrations, and high pressures, suggesting that they are of marine origin or have adapted to the deep-sea environment. Received: January 22, 1998 / Accepted: February 16, 1998     

Biological patterns and ecological indicators for Mediterranean fish and crustaceans below 1,000 m: a review

The Mediterranean Sea is a relatively deep, closed sea with high rates of fisheries exploitation. In recent years fishing activity has tended to shift towards deeper depths. At the same time, the Mediterranean displays some rather special hydrographic and biogeographic conditions. The present paper reviews the present state of knowledge of the fisheries, biology, and ecology of the deep-sea fish and crustacean species in the Mediterranean dwelling below 1,000 m with potential economic interest, placing special emphasis on the western basin, for which more data are available, as a basis for future studies of the ecology, biodiversity, and effects of climate change and exploitation in this zone. This review reveals that mediterranean deep-sea fishes and crustaceans employ highly conservative ecological strategies, and hence the low fecundity and low metabolic rates in a stable environment like the deep-sea make these populations highly vulnerable. Moreover, ripe females of the main species mentioned here concentrate in the deepest portions of their distribution ranges. Deep-sea fish and crustaceans have high trophic levels and low to medium omnivory index values. The ecological indices discussed here, in combination with the limited knowledge of deep-sea ecosystems, clearly call for an approach based on the Precautionary Principle.     

Antarctic marine biodiversity and deep-sea hydrothermal vents

The diversity of many marine benthic groups is unlike that of most other taxa. Rather than declining from the tropics to the poles, much of the benthos shows high diversity in the Southern Ocean. Moreover, many species are unique to the Antarctic region. Recent work has shown that this is also true of the communities of Antarctic deep-sea hydrothermal vents. Vent ecosystems have been documented from many sites across the globe, associated with the thermally and chemically variable habitats found around these, typically high temperature, streams that are rich in reduced compounds and polymetallic sulphides. The animal communities of the East Scotia Ridge vent ecosystems are very different to those elsewhere, though the microbiota, which form the basis of vent food webs, show less differentiation. Much of the biological significance of deep-sea hydrothermal vents lies in their biodiversity, the diverse biochemistry of their bacteria, the remarkable symbioses among many of the marine animals and these bacteria, and the prospects that investigations of these systems hold for understanding the conditions that may have led to the first appearance of life. The discovery of diverse and unusual Antarctic hydrothermal vent ecosystems provides opportunities for new understanding in these fields. Moreover, the Antarctic vents south of 60°S benefit from automatic conservation under the Convention on the Conservation of Antarctic Marine Living Resources and the Antarctic Treaty. Other deep-sea hydrothermal vents located in international waters are not protected and may be threatened by growing interests in deep-sea mining.     

Small-scale heterogeneity in deep-sea nematode communities around biogenic structures

The unexpected high species richness of deep-sea sediments gives rise to the questions, which processes produce and maintain diversity in the deep sea, and at what spatial scales do these processes operate? The idea of a small-scale habitat structure at the deep-sea floor provides the background for this study. At small scales biogenic structures create a heterogeneous environment that influences the structure of the surrounding communities and the dynamics of the meiobenthic populations. As an example for biogenic structures, small deep-sea sponges (Tentorium semisuberites Schmidt 1870) and their sedimentary environment were investigated for small-scale distribution patterns of benthic deep-sea nematodes. Sampling was carried out with the remotely operated vehicle Victor 6000 at the Arctic deep-sea observatory HAUSGARTEN. In order to investigate nematode community patterns sediment cores around three small sponges and corresponding control cores were analysed. A total of approx. 5800 nematodes were identified. The comparison of the nematode communities from sponge and control samples indicated an influence of the biogenic structure “sponge” on diversity patterns and habitat heterogeneity. The increased number of nematode species and functional groups found in the sediments around the sponges suggest that on a small scale the sponge acts as a gradient and creates a more divers habitat structure. The nematode community from the sponge sediments shows a greater taxonomic variance and species richness together with lower relative abundances of the species compared to those from control sediments. Obviously, the more homogeneous habitat conditions of the control sediments offer less micro-habitats than the sediments around the sponges. This seems to reduce the number of functional groups and species coexisting in the control sediments.     

Culturability and Secondary Metabolite Diversity of Extreme Microbes: Expanding Contribution of Deep Sea and Deep-Sea Vent Microbes to Natural Product Discovery

Microbes from extreme environments do not necessarily require extreme culture conditions. Perhaps the most extreme environments known, deep-sea hydrothermal vent sites, support an incredible array of archaea, bacteria, and fungi, many of which have now been cultured. Microbes cultured from extreme environments have not disappointed in the natural products arena; diverse bioactive secondary metabolites have been isolated from cultured extreme-tolerant microbes, extremophiles, and deep-sea microbes. The contribution of vent microbes to our arsenal of natural products will likely grow, given the culturability of vent microbes; their metabolic, physiologic, and phylogenetic diversity; numerous reports of bioactive natural products from microbes inhabiting high acid, high temperature, or high pressure environments; and the recent isolation of new chroman derivatives and siderophores from deep-sea hydrothermal vent bacteria.