PDF-AGU Monograph BAB2

185

Microbial Ecology of Mid-Ocean Ridges and Back-Arc Basins

Ken Takai1, Satoshi Nakagawa1,2, Anna-Louise Reysenbach2, and Joost Hoek2

Over the past two decades, microbiologists have gained significant insights into the diversity and physiology of microbial communities associated with deep-sea hydrother-mal systems. Much of the initial research focused on mid-ocean ridge (MOR) systems; however, because of the greater heterogeneity of vent fluid chemistry and sulfide structures from back-arc basin (BAB) systems, recent studies have begun to explore the linkages between geochemistry and microbial diversity in these systems. The impact of microbes on local fluid chemistry and mineralogy has been recognized, and local fluid physical-geochemical states and mineralogical properties have significant impacts on the formation and composition of local microbial communities. Data are being accu-mulated that enable microbiologists not only to link phylogenetic and physiological diversity of microbial communities to geological and geochemical settings within a hydrothermal field, but also to review microbial ecosystems in global deep-sea hydro-thermal systems with comparison of representative MOR and BAB systems. In this review, we briefly outline methods in microbial ecology and microbial ecophysiology at vents and discuss the patterns of diversity (phylogenetic and physiological) emerging from studies in the global hydrothermal systems. As this volume is dedicated to BABs, we highlight several case studies on hydrothermal vent microbial communities in BAB systems that are comparable to the communities in MOR systems.

1. INTRODucTION

Deep-sea hydrothermal vents represent one of the most physically and chemically diverse habitats on Earth for microbial growth. The geochemical and thermal gradi-ents (e.g., >350°c across distances as small as 1 to 3 cm in active sulfide chimneys) provide a wide range of niches for microbial colonization. Psychrophiles, mesophiles, ther-mophiles, and hyperthermophiles (organisms growing best

from 4°c to above 80°c) thrive by chemolithoautotrophy or heterotrophy, utilizing the abundant available inorganic and organic chemical energy and carbon sources. These organisms exist as free-living forms within the hydrother-mal plumes, within mixed diffuse fluids, and as microbial mats on sediments; as attached biofilms on invertebrates and on sulfide mineral particles; and as obligate symbionts within invertebrate hosts. The effect of microbes on local fluid chemistry and mineralogy has also been recognized [Juniper and Tebo, 1995; Karl, 1995], and recent isolation of vent-related microbes that produce filamentous elemen-tal sulfur [Taylor et al., 1999] and of other microbes that are actively associated with weathering of extinct sulfides [Edwards et al., 2003a] has highlighted the importance and impact that microbial activity has on the geology, geochem-istry, and ecology of hydrothermal vent ecosystems at the seafloor as well as in subseafloor environments.

Over the past two decades, we have gained significant insights into the diversity and physiology of microbial com-

1 Subground Animalcule Retrieval (SuGAR) Project, Japan Agency for Marine-Earth Science and Technology (JAMSTEc), Yoko-suka, Japan

2 Department of Biology, Portland State university, Portland, Ore-gon, uSA

Back-Arc Spreading Systems: Geological, Biological, chemical, and Physical InteractionsGeophysical Monograph Series 166Published in 2006 by the American Geophysical union10.1029/166GM10

186 MIcROBIAL EcOLOGY Of MID-OcEAN RIDGES AND BAcK-ARc BASINS

munities associated with these unique ecosystems. Much of the initial research has focused on mid-ocean ridge (MOR) systems; recently, however, in light of the greater heterogeneity of the geochemistry of vent fluids from back-arc basins (BABs), several studies have begun to explore the linkages between geochemistry and microbial diversity in these systems.

In this review, we briefly overview methods in microbial ecology and microbial ecophysiology at vents and discuss the patterns of diversity (phylogenetic and physiological) that are emerging from studies in global hydrothermal systems includ-ing MOR, BAB, and volcanic arcs (VAs). Because this volume is dedicated to BABs, the microbiology of MOR systems sets the stage for comparative studies with BAB systems. Additional aspects of the microbiology of deep-sea vents and the ecology of thermophiles are addressed in numerous reviews [Jannasch, 1995; Karl, 1995; Prieur et al., 1995; Huber et al., 2000; Jeanthon, 2000; Takai and Fujiwara, 2002; Baross et al., 2003, 2004; Cary et al., 2004; Holland et al., 2004; Miroshnichenko, 2004; Takai et al., 2004a; Tivey, 2004].

2. METHODS IN MIcROBIAL EcOLOGY IN HYDROTHERMAL SYSTEMS

Microbiologists have specific challenges to enumerate microbial diversity. unlike most biologists, who can initially use morphological traits to distinguish new species, microbi-ologists need to rely on physiological or phylogenetic traits, and even then, the definition of the microbial species is much debated. Diversity has traditionally been assessed by culturing microorganisms. However, recreating the physical and chemi-cal properties of the environment accurately in a culture tube is rare and frequently results in a culture-biased view of diver-sity. This bias is particularly a problem for trying to culture microbes from deep-sea hydrothermal vents, because develop-ing appropriate media and recreating the extreme conditions in the laboratory are extremely challenging. consequently, it is generally accepted that the majority of microorganisms from vents are not detectable by methods that rely solely on labora-tory cultivation. Nevertheless, these approaches have been crucial in our understanding of the physiological diversity at deep-sea vents. for example, the upper temperature limits for life have been challenged by the characterization of Archaea growing at temperatures above 105°c [Blöchl et al., 1997; Kashefi et al., 2003], and microbes that use a wide range of electron donors and acceptors for growth have been described [Miroshnichenko et al., 2004, for review].

Most of the diversity at deep-sea vents is currently being described by using molecular phylogeny–based approaches that circumvent the need to cultivate microbes for assessing micro-bial diversity [Haddad et al., 1995; Moyer et al., 1995; Polz and Cavanaugh, 1995; Cary et al., 1997; Takai and Horikoshi,

1999; Reysenbach et al., 2000; Campbell and Cary, 2001; Corre et al., 2001; Longnecker and Reysenbach, 2001; Takai et al., 2001; Alain et al., 2002; Lopez-Garcia et al., 2002; Teske et al., 2002; Hoek et al., 2003; Dhillon et al., 2003; Huber et al., 2003; Alain et al., 2004; Nakagawa et al., 2004a, 2004b; Higashi et al., 2004; Takai et al., 2004b, 2004c; Nakagawa et al., 2005a; Nercessian et al., 2004, 2005]. This approach relies on the universal and highly conserved small subunit rRNA molecule (16S rRNA or 18S rRNA), by which all life can be organized within the domains Bacteria, Archaea, and Eukarya. Cary et al. [2004] reviewed these molecular phylogenetic approaches and elaborated more on some of the new advances in molecular ecological techniques being applied to deep-sea vent micro-bial communities. Briefly, as illustrated in figure 1, DNA and RNA are extracted from environmental samples colonized by microbes, and the 16S rRNA genes from the genomic DNA and cDNA assemblages of all microorganisms in the environment are amplified by polymerase chain reaction (PcR). These prod-ucts are then sorted by numerous methods such as cloning or using denaturing gradient gel electrophoresis (DGGE). unique clones or bands on the gel, respectively, are sequenced. using phylogenetic analysis methods, the sequences can be placed in a phylogenetic tree or other phylogenetic context. The differ-ent sequences (phylotype) represent the range of phylogenetic diversity in a sample, and their taxonomic position can be inferred from their closest cultured, known relative. Because the 16S rRNA gene sequence does not provide information on the metabolic function or other traits that the microbes might harbor in the environment, one has to be very careful if making inferences about the role these organisms play in the environ-ment. In some cases, however, inferences can be made; for example, if a sequence falls within a lineage that is occupied only by methanogens, then it is likely that the phylotype can produce methane. furthermore, the sequence information can be used to develop molecular probes and primers that can spe-cifically identify the microorganism in the original sample and explore the distribution and abundance of the microorganisms and their rRNA in multiple samples by using fluorescent in situ hybridization (fISH) and quantitative PcR and hybridization techniques. figure 1 illustrates the above processes.

functional diversity can be explored in a number of dif-ferent molecular approaches [Cary et al., 2004, for review] and can be compared to molecular phylogenetic diversity. The potential metabolic function of a community can be inferred from detecting genes or mRNA that are diagnostic for a specific metabolism. for example, a key enzyme for dissimilatory sulfate reduction, dissimilatory sulfite reduc-tase, is encoded by the dsrAB gene, which has been explored at deep-sea vents [Cottrell and Cary, 1999; Dhillon et al., 2003; Hoek et al., 2003; Nakagawa et al., 2004a, 2004b; Nercessian et al., 2005]. Another frequently examined

figure 1figure 1

TAKAI ET AL. 187

functional gene is the mcrA gene, which encodes an alpha subunit of methyl coenzyme M reductase, a key enzyme of methanogenesis; this gene has also been explored at various deep-sea vents [Dhillon et al., 2005; Nercessian et al., 2005]. More detailed functional diversity can be assessed by using metagenomic approaches [DeLong, 2002, for review], which will ultimately shed light on the physiology and ecology of the as yet largely uncultured diversity at vents.

The next challenge for microbial ecologists is to link the functional and phylogenetic diversity in situ to biogeochemical processes. Several methods that may be applied at hydrothermal vents include stable isotope probing [Radajewski et al., 2000] and fISH-SIMS [Orphan et al., 2001]. Other potential applica-tions of high-resolution physical and chemical measurements to molecular biological approaches and microbial ecology will further enhance our understanding of these systems.

3. EMERGING PATTERNS Of MIcROBIAL DIVERSITY IN GLOBAL

HYDROTHERMAL SYSTEMS

3.1. Molecular Phylogenetic Diversity

Initial microbial ecological studies at deep-sea vents were limited to enrichment culturing or activity measurements [Jannasch and Wirsen, 1979; Baross et al., 1982; Baross and Deming, 1983; Jannasch and Wirsen, 1985; Wirsen et al., 1986]. However, with the development of molecular phylo-

genetic tools for microbial ecology, our view of the microbial diversity at vents has greatly expanded [for review and exam-ples, see Reysenbach and Shock, 2002; Wilcock et al., 2004]. In most cases, diversity assessments reveal numerous lineages that still have no known representatives in culture, although they may provide hints on how to grow novel microbes from vents. These molecular phylogenetic inventories form the framework for more in-depth, hypothesis-driven ecological studies at vents. One can now begin to explore the constraints (chemical or physical) on the patterns of microbial diversity at vents and to compare the microbial ecology of BAB hydro-thermal systems with that of MOR systems.

3.1.1. Archaea from deep-sea vents. The most compre-hensive diversity assessments of deep-sea vents to date have been limited to the archaeal diversity of communities associ-ated with active chimney structures [e.g., Takai and Horiko-shi, 1999; Reysenbach et al., 2000; Corre et al., 2001; Takai et al., 2001, 2004b, 2004c; Schrenk et al., 2003]. Although Archaea appear to represent a smaller component (1.8–40%) of the total cells associated with some sulfide chimneys [Harmsen et al., 1997; Takai and Horikoshi, 1999; Takai et al., 2001; Hoek et al., 2003; Schrenk et al., 2003], they do appear to be important and major components of the diver-sity in certain portions of chimney structures. for example, using quantitative fluorogenic PcR and fISH, Takai et al. [2004c] demonstrated the predominance (nearly 100%) of Archaea in “in situ colonization systems” (IScS) and interior

Figure 1. Schematic illustration of cultivation-independent, molecular ecological research.


parts of sulfide chimneys in the Kairei hydrothermal field of the central Indian Ridge. Schrenk et al. [2003] showed that 40% of the microorganisms colonizing the exterior wall of an active sulfide chimney in the Juan de fuca Ridge were Archaea, the archaeal proportions increasing steadily toward the center of the chimney.

These and other studies have led to emerging patterns of archaeal diversity. In particular, one lineage appears to be distributed throughout the global deep-sea vent system. Takai and Horikoshi [1999] referred to this lineage as the “Deep-sea Hydrothermal Vent Euryarchaeoic group” (DHVEG). The lineage is split into DHVE-1 and DHVE-2 [Reysenbach and Shock, 2002], DHVE-1 being more commonly identi-fied from vents [Takai and Horikoshi, 1999; Reysenbach et al., 2000; Takai et al., 2001; Longnecker and Reysenbach, 2002; Hoek et al., 2003; Schrenk et al., 2003]. Members of the DHVEG lineage have formed the majority of clones (up to 93% of the clones for the DHVE-1) in several archaeal 16S rDNA chimney clone libraries (Table 1).

Some reports have documented numerous other uncul-tured archaeal lineages [e.g., Takai and Horikoshi, 1999; Takai et al., 2001] that included some representatives of the “Korarchaeota” [Barns et al., 1994] or such unusual lineages as “Nanoarchaeum” [Hohn et al., 2002; Huber et al., 2002] (Table 1). Not surprisingly, few members of cultured representatives are present in the environmental clone libraries. Among these cultured members, the hyper-thermophilic methanogens in the order Methanococcales are frequently detected [Reysenbach et al., 2000; Teske et al., 2002; Nercessian et al., 2003; Schrenk et al., 2003; Takai et al., 2004b, 2004c; Nakagawa et al., 2005a], including clone libraries obtained from a 2-day deployment of an in situ enrichment device called the “vent cap” [Reysenbach et al., 2000] and a 7-day deployment of an IScS [Takai et al., 2004c] (Table 1). However, other methanogens belong-ing to the order Methanosarcinales are often detected in clone libraries, but they have never been isolated from vents [Longnecker, 2001; Teske et al., 2002; Schrenk et al., 2003] (Table 1). The order Thermococcales, which is perhaps the most frequently cultured group from deep-sea vents [e.g., Lepage et al., 2004; Takai et al., 2004c; Nakagawa et al., 2005a], is not commonly detected in clone libraries, with a few exceptions [Reysenbach et al., 2000; Teske et al., 2002; Hoek et al., 2003; Schrenk et al., 2003] (Table 1). Perhaps in the shallow subsurface environments at vents, complex community biofilms develop that provide complex organ-ics for the growth of Thermococcales. If this is the case, then the hyperthermophilic Thermococcales might also be good indicators of subsurface heterotrophic activity at vents [Reysenbach et al., 2000; Summit and Baross, 2001; Holland et al., 2004].

3.1.2. Bacteria from deep-sea vents. Bacteria appear to dominate most niches at deep-sea vents, the ε-Proteobacteria being particularly abundant, widespread, and detected in a variety of habitats, including sulfide structures [Polz and Cavanaugh, 1995; Hoek et al., 2003; Nakagawa et al., 2004b], hydrothermal fluid/seawater mixing zones [Reysenbach et al., 2000; Corre et al., 2001; Huber et al., 2003; Sunamura et al., 2004; Takai et al., 2004a; Nakagawa et al., 2005d], hydro-thermal sediments [Teske et al., 2002], and microbial mats [Moyer et al., 1995; Longnecker, 2001]. Some ε-Proteobacte-ria are found in episymbiotic association with deep-sea vent metazoans [Haddad et al., 1995; Polz and Cavanaugh, 1995; López-García et al., 2002; Goffredi et al., 2004] (Table 2) and even as endosymbionts in snails [Suzuki et al., 2005; Urakawa et al., 2005]. Despite their ubiquitous and cosmopolitan dis-tribution, the physiological aspects of these microorganisms are poorly understood because of their strong resistance to cultivation. Previous studies had characterized the ε-Proteo-bacteria as mostly microaerobic sulfur-oxidizers [Taylor et al., 1999; López-García et al., 2002]. However, recent culturing studies have revealed a much greater metabolic diversity of the ε-Proteobacteria. Recently, numerous representatives of the deep-sea hydrothermal vent ε-Proteobacteria have been isolated in pure cultures and characterized [Campbell and Cary, 2001; Campbell et al., 2001; Alain et al., 2002; Mirosh-nichenko et al., 2002, 2004; Inagaki et al., 2003, 2004; Takai et al., 2003a, 2004d, 2005a; Nakagawa et al., 2005a, 2005b, 2005c, 2005d]. These studies indicate that the deep-sea hydro-thermal vent ε-Proteobacteria are comprised of mesophilic to moderately thermophilic chemolithoautotrophs capable of oxidizing hydrogen and sulfur compounds with nitrate, oxygen, and sulfur compounds as terminal electron acceptors [Campbell and Cary, 2001; Alain et al., 2002; Miroshnichenko et al., 2002, 2004; Inagaki et al., 2003, 2004; Takai et al., 2003a, 2004d, 2005a; Nakagawa et al., 2005a, 2005b, 2005c, 2005d]. It is now well accepted that the ε-Proteobacteria play a significant role not only in the cycling of sulfur, but also in the cycling of hydrogen, nitrogen, and carbon in deep-sea hydrothermal environments [Takai et al., 2003a; Nakagawa et al., 2005b, 2005c].

Another group of chemolithoautotrophs is represented by members of the order Aquificales, which utilize similar elec-tron donor/acceptor pairs as the ε-Proteobacteria [Götz et al., 2002; Nakagawa et al., 2003]. Like the ε-Proteobacteria, the Aquificales are found in globally widespread deep-sea hydro-thermal fields [Reysenbach et al., 2000; Nakagawa et al., 2003, 2004b; Takai et al., 2003b] (Table 2). As predicted from the high growth temperatures of the Aquificales, these microorganisms are found only in high-temperature habitats such as active sul-fide structures and vent fluids [Reysenbach et al., 2002; Takai et al., 2003b; Nakagawa et al., 2004b, 2005a].

Table 1Table 1

Table 2Table 2

TAKAI ET AL. 189

Table 1. Summary of phylogenetic diversity of archaeal rRNA gene clones in deep-sea hydrothermal environments.

Group Location Habitat ReferencesEuryarchaeota Archaeoglobales

MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Mothra chimney Schrenk et al., 2003NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei In situ colonization device and

chimneyTakai et al., 2004c

Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Higashi et al., 2004Okinawa Trough (BAB) Iheya North chimney Nakagawa et al., 2005a

ThermococcalesMAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Mothra chimney Schrenk et al., 2003NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei In situ colonization device and


Hydrothermal plume Takai et al., 2004b Edmond Sulfide spire Hoek et al., 2003Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Higashi et al., 2004Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001Okinawa Trough (BAB) Iheya North In situ colonization device and

chimneyNakagawa et al., 2005a

MethanococcalesJuan de fuca (MOR) Mothra chimney Schrenk et al., 2003NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei In situ colonization device and


Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001Okinawa Trough (BAB) Iheya North In situ colonization device and


MethanobacterialesGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002


Group Location Habitat References ANME-1

Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Okinawa Trough (BAB) Iheya Ridge Hydrothermal sediments Takai and Horikoshi, 1999

MethanomicrobialesGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

Methanosarcinales ANME-2

Juan de fuca (MOR) Mothra chimney Schrenk et al., 2003Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

MethanosarcinacIR (MOR) Kairei In situ colonization device and


MethanopyralesNEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei In situ colonization device and


Hydrothermal plume Takai et al., 2004b Halobacteriales

Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001

DHVEG DHVEG-1 (Marine Benthic Group D)

MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Mothra chimney Schrenk et al., 2003NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei chimney Takai et al., 2004cGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001Izu-Bonin Arc (VA) Myojin Knoll chimney Takai and Horikoshi, 1999

DHVEG-2MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Edmond Sulphide spire (diffuse flow) Hoek et al., 2003Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Higashi et al., 2004

chimney Takai and Horikoshi, 1999 Myojin Knoll chimney Takai and Horikoshi, 1999Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006

Table 1. Cont.

TAKAI ET AL. 191

Group Location Habitat ReferencesOkinawa Trough (BAB) Iheya Ridge Hydrothermal sediments Takai and Horikoshi, 1999 Iheya North In situ colonization device Nakagawa et al., 2005a

DHVEG II DHVEG-3, 4, 5, 6, 8 NEPR (MOR)

13°N In situ growth chamber Nercessian et al., 2003Juan de fuca (MOR) Mothra chimney Schrenk et al., 2003Izu-Bonin Arc (VA) Myojin Knoll chimney Takai and Horikoshi, 1999Okinawa Trough (BAB) Iheya Ridge Hydrothermal sediments Takai and Horikoshi, 1999 Iheya North In situ colonization device Nakagawa et al., 2005a

Marine Group IIJuan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003cIR (MOR) Kairei In situ colonization device and


Hydrothermal plume Takai et al., 2004bLoihi Seamount (hot spot) PeleÅfs vent Microbial mat Moyer et al., 1998Okinawa Trough (BAB) Iheya North In situ colonization device and


Marine Benthic Group EcIR (MOR) Kairei In situ colonization device and


crenarchaeota Desulfurococcales

NEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Karai In situ colonization device and


Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006Okinawa Trough (BAB) Iheya North In situ colonization device and


Marine Group INEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003 Mothra chimney Schrenk et al., 2003cIR (MOR) Kairei In situ colonization device and


Hydrothermal plume Takai et al., 2004bIzu-Bonin Arc (VA) Myojin Knoll chimney Takai and Horikoshi, 1999

Table 1. Cont.


Bacterial diversity at deep-sea vents spans most of the known, nonphotosynthetic lineages, including the firmicutes, the Verrucomicrobia, the Thermales, and the Cytophaga–Flexibacter–Bacteroides (cfB) group (Table 2). The Verrucomicrobia have been detected at numerous hydro-thermal sites [Alain et al., 2002; Lopez-Garcia et al., 2002; Teske et al., 2002] on the Juan de fuca Ridge, Northern East Pacific Rise, and Guaymas Basin (Table 2). This group of organisms has been difficult to grow from most environ-ments; enrichment cultures growing on complex organic media at 70°c were gradually selected for a new verruco-microbial strain, but the strain could not be retrieved later from the culture collection storage [A.-L. Reysenbach and D. Götz, unpublished data, 2000].

Another bacterial group also detected in several clone libraries from deep-sea hydrothermal vent environments is the green nonsulfur bacteria, presently designated as the phylum chloroflexi [Alain et al., 2002; Teske et al., 2002; Dhillon et al., 2003; Goffredi et al., 2004] (Table 2). Based on the phylogenetic tree inferred from 16S rRNA gene sequences [Hugenholtz et al., 1998], the phylum chloroflexi is divided into four subphyla. Of these, Subphylum III com-prises the cultured representatives of a wide range of phe-notypes, including thermophilic phototrophic bacteria from terrestrial hot springs, such as Chloroflexus aurantiacus and Herpetosiphon geysericola. Other than Subphylum III, there are very limited cultured representatives of the nonphoto-synthetic members. The anaerobic dechlorinating bacterium

Group Location Habitat ReferencesOkinawa Trough (BAB) Iheya North Vent fluid Takai and Horikoshi, 1999

In situ colonization device and chimney

Nakagawa et al., 2005a

Loihi Seamount (hot spot) PeleÅfs vent Microbial mat Moyer et al., 1998

Marine Benthic Group B (DHVcG-I, SAG, MHVG1-3)

MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Manus Basin (BAB) PAcMANuS chimney Takai et al., 2001Izu-Bonin Arc (VA) Suiyo seamount chimney Takai and Horikoshi, 1999 Myojin Knoll chimney Takai and Horikoshi, 1999Okinawa Trough (BAB) Iheya North In situ colonization device and


Marine Pelagic Arch Group IGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

Hot Water crenarchaeota Group (HWcG) Juan de fuca (MOR)

Mothra chimney Schrenk et al., 2003Okinawa Trough (BAB) Iheya Ridge Hydrothermal sediments Takai and Horikoshi, 1999 Iheya North chimney Nakagawa et al., 2005a

KorarchaeotaNEPR (MOR) 13°N In situ growth chamber Nercessian et al., 2003cIR (MOR) Kairei In situ colonization device and


Okinawa Trough (BAB) Iheya North chimney Nakagawa et al., 2005a

NanoarchaeotaIzu-Bonin Arc (VA) Suiyo Seamount chimney Nakagawa et al., 2004b

Table 1. Cont.

TAKAI ET AL. 193

Dehalococcoides dehalogenes is the only cultivated repre-sentative within the Subphylum II, growing by the oxidation of hydrogen and dechlorination of tetrachlorethene to ethane [Maymo-Gatell et al., 1997]. Anaerolinea thermophila and Caldilinea aerophila are thermophilic chemorogantrophs of Subphylum I, isolated from thermophilic granular sludge and microbial mats in a hot spring, respectively [Sekiguchi et al., 2003]. The habitat preferences of nonphotosynthetic green nonsulfur bacteria are compatible with the geochemi-cal features of sediment-hosted hydrothermal systems, such as the Guaymas Basin, where organic biomass undergoes pyrolysis and thermal alteration to a wide variety of petro-leum hydrocarbons, including unbranched alkanes, cycloal-kanes, triterpanes, steranes and diasteranes, and aromatic hydrocarbons [Teske et al. 2002].

Recently, several studies have explored the diversity of dis-similatory sulfate-reducing bacteria (SRB) at deep-sea vents by using a functional gene approach [Dhillon et al., 2003; Hoek et al., 2003; Nakagawa et al., 2004a, 2004b] (Table 3). These studies, which used the dsrAB gene, have expanded our view of the distribution and diversity of SRB at vents and show a diverse assemblage of sulfate reducers not only in hydrothermal sediments of the Guaymas Basin but also in the walls of actively venting sulfide chimneys. In addi-tion, the molecular phylogenetic surveys of the dsrAB gene have demonstrated the existence of previously uncultivated and unidentified groups of potential SRB, designated as “functional gene-only” groups of SRB [Dhillon et al., 2003; Nakagawa et al., 2004a, 2004b]. It has been suggested that these sulfate-reducers can have a significant impact on the chemistry and mineralogy of local microenvironments and may play a role in the precipitation and alteration of sulfide minerals in deep-sea vent ecosystems [Shanks, 2001]. To begin to address the potential geobiological role of sulfate-reducers in deep-sea vents, J. Hoek and A.-L. Reysenbach [unpublished data, 2003] explored how a thermophilic, H2-oxidizing, chemolithoautotrophic sulfate-reducing bacterium fractionated sulfur isotopes under the steep geochemical gradients of sulfide chimneys. The extent of fractionation was strongly dependent on the concentration of H2 in the environment; where H2 limited growth, sulfide was depleted in 34S by up to ~40‰ from the original sulfate. This reduc-tion suggests that sulfide minerals derived from the sulfide of chemolithoautotrophic SRB can be detected from the isotopic signature of sulfur, which is potentially an important tool for studying the impact of sulfate-reducing prokaryotes on biogeochemical cycling in deep-sea vents.

Metal oxidizers such as iron-oxidizing Bacteria are impor-tant contributors to microbial weathering of extinct sulfides and can have a significant impact on the biogeochemical cycling of metals and sulfur at deep-sea vents [Edwards et

al., 2003a, 2003b]. McCollum and Shock [1997] estimated that the amount of energy available from the oxidation of metal sulfide minerals precipitated from a seafloor hydro-thermal plume exceeded the energy available from dissolved compounds (H2S, cH4, Mn2+, H2) by nearly an order of magnitude per kilogram of vent fluid. Edwards et al. [2003a] examined the role of iron-oxidizing Bacteria in the oxidation of ferrous iron (fe2+) from sulfide minerals as an energy source in seafloor environments. Polished slabs of chimney sulfides, when incubated on the seafloor at ambient tempera-tures (~4°c), were heavily colonized, and colony density showed a positive correlation with the solubility of the min-eral substrate. The observation of concomitant formation of fe-oxide on the chimney slabs suggests that the fe-oxides formed in situ during bacterial growth. further support for the presence and activity of iron-oxidizing Bacteria associ-ated with seafloor hydrothermal vent sulfides comes from the isolation of iron-oxidizing Bacteria from the in situ incu-bation experiments of Edwards et al. [2003a]. Phylogenetic characterization of these isolates revealed a diverse collection of α- and γ-Proteobacteria, which were not closely related to any known iron-oxidizing Bacteria [Edwards et al., 2003b]. These results suggest that iron-oxidizing Bacteria are preva-lent and active at seafloor hydrothermal sulfide habitats.

3.2. Microbial Physiological Diversity at Deep-Sea Vents

Some of the diversity described above represents the range of physiological diversity at vents—from mesophiles to ther-mophiles, from obligate chemolithoautotrophs to facultative heterotrophs—and includes anaerobes, microaerophiles, and aerobes. This diversity is supported by the steep chemical and physical gradients typical of deep-sea hydrothermal environments. These steep gradients generate a wide range of niches and energy sources for microorganisms. for exam-ple, chemolithoautotrophs can generate energy by exploiting the chemical disequilibria resulting from sluggish reaction kinetics for redox reactions at the interface between oxidized seawater (e.g., O2, NO3

–, and SO42–) and reduced hydro-

thermal vent fluids (e.g., H2, H2S, cH4, cO2, and formate). Although a range of novel chemolithoautotrophs have been isolated from deep-sea vents [Jannasch, 1995; Karl, 1995; Prieur et al., 1995; Huber et al., 2000; Jeanthon, 2000; Takai and Fujiwara, 2002; Miroshnichenko et al., 2004], many more remain uncultured, perhaps including some that use rare or unusual redox couples (Table 4) [Shock and Holland, 2004]. Although not all of these processes are microbio-logically mediated at high temperatures, microniches likely exist where thermophiles capitalize on the mixing between cold oxygenated seawater and reduced hydrothermal vent fluid. Microbes that are able to use multiple electron donors

Table 3Table 3

Table 4Table 4


Group Location Habitat Referencesε-Proteobacteria Subgroup A

MAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001Juan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002cIR (MOR) Kairei In situ colonization device and


Okinawa Trough (BAB) Iheya North In situ colonization device and


Subgroup BMAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001

In situ growth chamber Reysenbach et al., 2000 Rainbow In situ growth chamber Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial Volcano Hydrothermal Plume Huber et al., 2003

Paralvinella tube Alain et al., 2002NEPR (MOR) 13°N In situ growth chamber Alain et al., 2004Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002;

Dhillon et al., 2003cIR (MOR) Kairei In situ colonization device and


Loihi Seamount (Hot Spot) Pele’s vent Microbial mat Moyer et al., 1995Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Higashi et al., 2004Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006Okinawa Trough (BAB) Iheya North In situ colonization device and


Subgroup cMAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003cIR (MOR) Kairei In situ colonization device and


Subgroup DMAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001

In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003NEPR (MOR) 13°N In situ growth chamber Alain et al., 2004SEPR (MOR) 17°S chimney Longnecker and Reysenbach,

2001

Table 2. Summary of phylogenetic diversity of bacterial rRNA gene clones in deep-sea hydrothermal environments.

TAKAI ET AL. 195

Table 2. Cont.

Group Location Habitat ReferencescIR (MOR) Edmond Sulphide spire Hoek et al., 2003 Kairei In situ colonization device and


Izu-Bonin Arc (VA) Suiyo Sea-Mount In situ growth chamber Higashi et al., 2004Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006Okinawa Trough (BAB) Iheya North In situ colonization device Nakagawa et al., 2005a

Subgroup E (Sulfurospirillum Group)MAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001Juan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002NEPR (MOR) 13°N In situ growth chambers Alain et al., 2004cIR (MOR) Edmond Sulphide spire diffuse flow Hoek et al., 2003 Kairei In situ colonization device and


Scaly snail (Crysomallon squamiferum)

Goffredi et al., 2004



Subgroup fMAR (MOR) Snake Pit In situ growth chamber Corre et al., 2001

In situ growth chamber Reysenbach et al., 2000 Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003

In situ growth chamber Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003

Paralvinella tube Alain et al., 2002NEPR (MOR) 13°N A. pompejana epibiont Haddad et al., 1995

In situ growth chambers Alain et al., 2004 9°N R. pachyptila tube Lopez-Garcia et al., 2002cIR (MOR) Edmond Sulphide spire diffuse flow Hoek et al., 2003 Kairei In situ colonization device and




Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002; Dhillon et al., 2003

Izu-Bonin Arc (VA) Suiyo Seamount Hydrothermal plume Sunamura et al., 2004Okinawa Trough (BAB) Iheya North In situ colonization device and


Subgroup GMAR (MOR) Snake Pit In situ growth chamber Corre et al., 2000


Table 2. Cont.

Group Location Habitat ReferencesJuan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003SEPR (MOR) 17°S chimney Longnecker and Reysenbach,

2001cIR (MOR) Kairei In situ colonization device and


Mariana Arc (VA) TOTO caldera Sulfur chimney

Okinawa Trough (BAB) Nakagawa et al., 2006 Iheya North In situ colonization device and


Arcobacter GroupMAR (MOR) Rainbow In situ growth chamber Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003NEPR (MOR) 13°N In situ growth chambers Alain et al., 2004cIR (MOR) Kairei In situ colonization device and


α-Proteobacteria MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003NEPR (MOR) 9°N R. pachyptila tube Lopez-Garcia et al., 2002cIR (MOR) Kairei In situ colonization device and


Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Higashi et al., 2004Okinawa Trough (BAB) Iheya North In situ colonization device Nakagawa et al., 2005a

β-Proteobacteria MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003 Snake Pit In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003cIR (MOR) Kairei In situ colonization device and


γ-Proteobacteria MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003

Paralvinella tube Alain et al., 2002NEPR (MOR)

TAKAI ET AL. 197

Table 2. Cont.

Group Location Habitat References 9°N R. pachyptila tube Lopez-Garcia et al., 2002 13°N In situ growth chambers Alain et al., 2004cIR (MOR) Kairei In situ colonization device and




Loihi Seamount (hot spot) Microbial mat Moyer et al., 1995Izu-Bonin Arc (VA) Suiyo Seamount Hydrothermal plume Sunamura et al., 2004Mariana Arc (VA) TOTO caldera Sulfur chimney Nakagawa et al., 2006Okinawa Trough (BAB) Iheya North In situ colonization device and


δ-Proteobacteria Juan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002

cIR (MOR) Kairei In situ colonization device and






DesulfobacteriumGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002; Dhillon et

al., 2003Desulfobulbus

Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Myxobacteria

MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Loihi Seamount (hot spot) Microbial mat Moyer et al., 1995

GeobacterGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

cfB Group MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000 Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003

In situ growth chamber Lopez-Garcia et al., 2003Juan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002; Dhillon et

al., 2003NEPR (MOR) 9°N R. pachyptila tube Lopez-Garcia et al., 2002 13°N In situ growth chamber Alain et al., 2004cIR (MOR) Kairei In situ colonization device and



Table 2. Cont.

Group Location Habitat ReferencesScaly snail (Crysomallon squamiferum)




Green Nonsulfur Group (Chloroflexi)

Juan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002Guaymas Basin (MOR) Hydrothermal sediment Teske et al., 2002; Dhillon et

al., 2003cIR (MOR) Kairei Scaly snail (Crysomallon

squamiferum)Goffredi et al., 2004



Desulfurobacterium GroupMAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003cIR (MOR) Kairei In situ colonization device and


Thermodesulfobacterium GroupJuan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003cIR (MOR) Kairei In situ colonization device and



Aquificales MAR (MOR) Snake Pit In situ growth chamber Reysenbach et al., 2000cIR (MOR) Edmond Sulphide spire diffuse flow Hoek et al., 2003 Kairei In situ colonization device and



VerucomicrobiaJuan de fuca (MOR) Axial volcano Paralvinella tube Alain et al., 2002NEPR (MOR) 9°N R. pachyptila tube Lopez-Garcia et al., 2002Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

Planctomycetales MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Okinawa Trough (BAB) Iheya North chimney Nakagawa et al., 2005a

Cyanobacteria and chloroplasts Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

TAKAI ET AL. 199

and acceptors are best suited for these fluctuating environ-ments and steep gradients. Thus, these Bacteria can also occupy suboxic zones where NO3

–, S2O32–, fe3+, and Mn4+ are

the oxidizing agents instead of O2. Many vent thermophiles show a metabolic versatility that may reflect their ability to thrive in environments where the geochemical gradients are fluctuating, including the facultative aerobic obligate chemo-lithotroph Pyrolobus fumarii [Blöchl et al., 1997], members of Aquificales [Götz et al., 2002; Nakagawa et al., 2003], the Desulfurobacteria group [L’Haridon et al., 1998; Alain et al., 2003; Takai et al., 2003b], and ε-Proteobacteria [Campbell and Cary, 2001; Alain et al., 2002; Miroshnichenko et al., 2002, 2004; Inagaki et al., 2003, 2004; Takai et al., 2003a, 2004d, 2005a; Nakagawa et al., 2005a, 2005b, 2005c, 2005d].

Although calculations of thermodynamic free energy from redox reactions can be used as proxies for metabolic poten-tial, they can be misleading if kinetic inhibition of certain chemical reactions under different conditions as well as com-petition and bioavailability of electron donors and acceptors in the natural environment are not considered. Thus, mod-eling efforts that take into account the different constraints on biological activity provide a framework for exploring novel physiologies and metabolisms. Tivey [2004] estimated environmental conditions (temperature, chemistry, and pH) within chimneys and flanges of seafloor vent deposits by developing models that account for rates of fluid flow, dif-fusive and advective transport of seawater and end-member fluid across chimney walls, the composition and temperature

Table 2. Cont.

Group Location Habitat ReferencesFirmicutes

Juan de fuca (MOR) Axial volcano Hydrothermal plume Huber et al., 2003

Paralvinella tube Alain et al., 2002Guaymas Basin (MOR) Hydrothermal sediment Teske et al., 2002; Dhillon et

al., 2003 13°N In situ growth chamber Alain et al., 2004MAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003

Spirochetes NEPR (MOR) A. pompejana epibionts campbell and cary, 2001Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

Nitrospira GroupMAR (MOR) Rainbow Hydrothermal sediments Lopez-Garcia et al., 2003

Thermus/Deinococcus Group Okinawa Trough (BAB) Iheya North chimney Nakagawa et al., 2005a

OP8 candidate division NEPR (MOR) 9°N R. pachyptila tube Lopez-Garcia et al., 2002Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002; Dhillon et

al., 2003OP1 candidate division

Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Okinawa Trough (BAB) Iheya North chimney Nakagawa et al., 2005a

OP3 candidate divisionGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002

OP5 candidate divisionGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002, Dhillon et

al., 2003OP9 candidate division

Guaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002, Dhillon et al., 2003

OP11 candidate divisionGuaymas Basin (MOR) Hydrothermal sediments Teske et al., 2002Okinawa Trough (BAB)

　 Iheya North In situ colonization device Nakagawa et al., 2005a


Group Location Habitat ReferencesdsrAB Iheya North group (group I)


Guaymas group (group IV)Guaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2003

ArchaeoglobaceaecIR (MOR) Kairei chimney Nakagawa et al., 2004a

NEPR (MOR) 13 ˚N chimney and in situ sampler Nercessian et al., 2004

SyntrophobacteralesGuaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2003

Gram-positive SRB relativesGuaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2003

Desulfoarculus-relatives


Desulfobulbaceae-relatives

NEPR (MOR) 13 ˚N chimney and in situ sampler Nercessian et al., 2004

MAR (MOR) Rainbow Hydrothermal sediments Nercessian et al., 2004


Izu-Bonin Arc (VA) Suiyo Seamount In situ growth chamber Nakagawa et al., 2004b

Thermodesulfobacteria-relatives

cIR (MOR) Kairei chimney Nakagawa et al., 2004aNEPR (MOR) 13 ˚N chimney and in situ sampler Nercessian et al., 2004



DesulfobacteraceaeGuaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2003


mcrA

MethanopyralesNEPR (MOR) 13˚N In-situ sampler Nercessian et al., 2004

MAR (MOR) Rainbow Hydrothermal sediments Nercessian et al., 2004

Methanococcales

MAR (MOR) Rainbow chimney Nercessian et al., 2004

Guaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2005 Methanosarcinales-relatives

MAR (MOR) Rainbow Hydrothermal sediments Nercessian et al., 2004Guaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2005

Methanomicrobiales-relativesGuaymas Basin (MOR) Hydrothermal sediments Dhillon et al., 2005

Table 3. Diversity of dsrAB and mcrA gene clones in deep-sea hydrothermal environments.

TAKAI ET AL. 201

of ambient seawater and end-member vent f luid, and the physical configuration (pore size and mineral distribution) of single-walled vent structures. Results from these modeling efforts indicate that the pH and oxidation state of pore fluids across chimney walls are highly sensitive to the composition of the end-member fluid and to different styles of mixing. The temperature of the transition from oxidized to reduced conditions varied from <3°c to ~90°c. According to the model, metabolic energy available to microorganisms within the vent structures is largely a function of the oxidation state of the pore fluid. furthermore, the pH of pore fluids at tem-peratures of 80°c to 120°c is generally low (pH < 3–4.5), suggesting that actively venting chimneys may harbor as yet uncultivated thermophilic, anaerobic acidophiles.

Patterns of inorganic carbon assimilation are quite diverse in vent environments and might be strongly associated with local physical and chemical conditions coupled with the inorganic carbon flow. Hyperthermophilic Euryarchaeota, such as methanogens and Archaeoglobales members, frequently colonize in the highest temperature ranges of habitats and utilize the acetyl-coA pathway for their auto-trophic cO2 fixation [Fuchs, 1990, 1994; Vorholt et al., 1995, 1997]. In contrast, the carbon fixation pathways of mem-bers of Desulfurococcales such as Pyrolobus, Pyrodictium, Desulfurococcus, and Ignicoccus, which are hyperthermo-philic crenarchaeota dwelling in deep-sea hydrothermal vent environments, are still uncertain, but the modified calvin cycle and a potentially new carbon fixation pathway have been suggested after enzymatic analyses for Pyrodictiaceae

and Desulfurococcaceae, respectively [Hügler et al., 2003]. In the lower temperatures of mixing zones, it has been dem-onstrated that marine crenarchaeota group I (MGI) repre-sents the most abundant archaeal components [Moyer et al., 1998; Takai et al., 2004b]. Stable and radiocarbon isotopic analyses of archaeal membrane lipids (glycerol dibiphytanyl glycerol tetraethers) have suggested that these previously uncultivated MGI members can grow autotrophically by bicarbonate fixation via a 3-hydroxypropionate pathway [Pearson et al., 2001; Wuchter et al., 2003]. In the bacte-rial components, the reductive tricarboxylic acid (rTcA) cycle is probably one of the most predominant cO2 fixation pathways operated by Aquifex and Persephonella, members of Aquificales [Zhang et al., 2002]. In addition, several molecular phylogenetic analyses of key functional genes for autotrophic carbon fixation pathways and enzymatic analy-ses using deep-sea ε-Proteobacteria and γ-Proteobacteria isolates have demonstrated that the rTcA cycle can serve as a cO2 fixation pathway of deep-sea ε-Proteobacteria and contribute to the primary production of the microbial com-munity together with the calvin cycle by γ-Proteobacteria in the mixing zones [Campbell et al., 2003; Campbell and Cary, 2004; Hügler et al., 2005; Takai et al., 2005b].

Tolerance of steep physical and chemical gradients is an important physiological characteristic of microorganisms in deep-sea vent environments. The effects of temperature and hydrostatic pressure at in situ habitats on the survivial of heterotrophic hyperthermophiles have been studied exten-sively [Trent et al., 1990; Holden and Baross, 1993, 1995;

Type of metabolismElectron donor

Electron acceptor

Redox reaction

Methanotrophy cH4 O2 cH4 + 2O2 = cO2 + 2H2OMethanotrophy cH4 SO4

2– cH4 + SO42– = HcO3

– + HS– + H2OMethanogenesis H2 cO2 H2 + 1/4cO2 = 1/4cH4 + 1/2H2OS reduction (sulfate reduction) H2 SO4

2– H2 + 1/4SO42– + 1/2H+ = 1/4H2S + H2O

S reduction (sulfur reduction) H2 S0 H2 + S0 = H2SS oxidation H2S O2 H2S + 2O2 = SO4

2– + 2H+

S oxidation S0 O2 S0 + H2O + 31/5O2 = SO42– + 2H+

S oxidation S2O32– O2 S2O3

– + 10OH– + O2 + 4H+ = 2SO42– + 7H2O

S oxidation/dentrification S2O32– NO3

– S2O3– + 6OH– + 4/5NO3

– + 4/5H+ = 2SO42– + 17/5H2O + 2/5N2

S oxidation/denitrification S0 NO3– S0 + 32/5H2O + 6/5NO3

– = SO42– + 34/5H+ + 3/5N2 + 6OH–

S oxidation/denitrification H2S NO3– H2S + 36/5H2O + 16/5NO3

– = 2SO42– + 84/5H+ + 8/5N2 + 16OH–

H2 oxidation H2 O2 H2 + 1/2O2 = H2Ofe reduction H2 fe(III) H2 + 2fe3+ = 2fe2+ + 2H+

fe oxidation fe(II) O2 fe2+ + 1/4O2 + H+ = fe3+ + 1/2H2Ofe oxidation/denitrification fe(II) NO3

– fe2+ + 1/5NO3– + 2/5H2O + 1/5H+ = 1/10N2 + fe3+ + OH–

Mn reduction H2 MnO2 H2 + MnO2 + 2H+ = Mn2+ + 2H2ONitrification NO2

– O2 NO2– + 1/2O2 + 2OH– + 2H+ = NO3

– + 2H2ONitrification NH3 O2 NH3 + 3OH– + 4O2 = 3NO3

– + 2H2ODenitrification H2 NO3

– H2 + 2/5NO3– + 2/5H2O = 1/5N2 + 8/5H+ + 2OH–

Table 4. Range of energetically favorable redox reactions available to chemolithotrophic microorganisms in deep-sea hydrothermal vents.


Marteinsson et al., 1997; Mitsuzawa et al., 2005]. All the investigations clearly demonstrated significantly increased thermotolerance patterns at elevated hydrostatic pressure, although the tolerance levels demonstrated are all within several minutes to several tens of minutes at or below 120°c. The gap between these laboratory experiments and observa-tion of hyperthermophiles in situ in superheated hydrother-mal fluids far above 120°c is a piece of the puzzle still to be clarified. In addition, tolerance of and susceptibility to metal toxicity are important physiological properties for deep-sea vent microorganisms because gradients of poten-tially toxic metals are formed concomitantly with mixing between hydrothermal fluids and ambient seawater. Vetriani et al. [2005] hypothesize that metal toxicity for microbial communities might be increased inversely with decreasing concentrations of heavy metals because of the increasing ratio of dilution by oxygenated ambient seawater, which is associated with increased bioavailability and solubility. This hypothesis is generally supported by experimental data showing that thermophilic Archaea and Bacteria from higher termperatures and more reductive habitats are more suscep-tible to toxic metal species, whereas mesophilic microbial components from lower temperatures and more oxidative habitats show higher tolerances for toxic metals [Jeanthon and Prieur, 1990; Llanos et al., 2000; Rathgeber et al., 2002; Edgcomb et al., 2004; Vertriani et al., 2005]. It is becoming evident that each of these physical and chemical parameters has a considerable impact on the growth and survival of individual microbial components, but the potential interac-tive effects at the community level of physical and chemical parameters are not well understood.

4. BAcK-ARc BASINS: WHY THEY ARE INTERESTING MIcROBIOLOGIcALLY

4.1. Overview

Several years after the discovery of deep-sea hydrothermal vents on MOR systems [Francheteau et al., 1979; Spiess et al., 1980], areas of deep-sea hydrothermal venting were dis-covered in BABs of the western Pacific. These hydrothermal systems, identified in Manus Basin [Both et al., 1986], Lau Basin and North fiji Basin [Hawkins, 1986; Hawkins and Helu, 1986], Mariana Trough [Craig et al., 1987a, 1987b], and Okinawa Trough [Kimura et al., 1988; Halbach et al., 1989] (figure 2), are geologically and tectonically distinct from hydrothermal systems found on MORs.

from a microbiological perspective, research in BAB hydrothermal systems has been limited. until recently, our understanding of microbial diversity in these environments had been restricted to the isolation and characterization of

some novel hyperthermophilic and heterotrophic microor-ganisms, including Pyrococcus abysii [Erauso et al., 1993], P. horikoshii [Gonzalez et al., 1998], Thermococcus profun-dus [Kobayashi et al., 1994], T. peptonophilus [Gonzalez et al., 1995] and Thermosipho melanesiensis [Antoine et al., 1997]. However, since the turn of this century, numerous microbiological investigations of western Pacific BAB hydro-thermal vent sites [Takai and Horikoshi, 1999, 2000; Takai et al., 2000, 2001, 2002, 2003a, 2004a, 2004d, 2005a; Inagaki et al., 2003, 2004; Nakagawa et al., 2004a, 2005a, 2005b, 2005c; Suzuki et al., 2004] have provided baseline culture-based and culture-independent diversity data. These data provide a framework for comparative studies with microbial diversity from MOR deep-sea hydrothermal systems.

In this section, we introduce the microbial communi-ties in two different well-characterized BAB hydrothermal systems, the PAcMANuS field in the Manus Basin and the Iheya North field in the Mid-Okinawa Trough. In addition

figure 2figure 2

Figure 2. Location and tectonic setting of deep-sea hydrothermal fields identified in arc–back-arc systems of the western Pacific margin. Numbers indicate the hydrothermal field as follows: 1, Iheya North; 2, Izena Hole; 3, Minami Ensei Knoll; 4, Hatoma Knoll; 5, Yonaguni Knoll IV; 6, Myojin Knoll; 7, Suiyo Seamount; 8, Alice Spring; 9, central Mariana Trough; 10, Off axial volcano; 11, TOTO caldera; 12, Vienna Woods; 13, PAcMANuS; 14, DES-MOS; 15, Station 4 (White Lady); 16, Station 14; 17, Vai Lili; 18, Hine Hina; 19, White church.

TAKAI ET AL. 203

to these two BAB hydrothermal fields, systematic and inten-sive microbiological explorations are under way or planned in other BAB systems of the Southern Mariana Trough and the Lau Basin. We discuss the significance of these BAB hydrothermal systems and patterns of diversity revealed by the comparison of microbial ecosystems in MOR and BAB systems.

4.2. Case Study: PACMANUS Site in the Manus Basin

The Manus Basin, located in the Bismarck Sea, is a back-arc spreading basin situated to the north of the New Britain Island Arc-Trench system (figure 2). The Manus Basin is characterized by three major spreading centers (western, central, and eastern spreading centers) linked by transform faults. The central basin has an exceptionally high spreading rate (>100 mm/yr [Baker et al., 1995]) and the eastern basin is currently in a “stretching” phase [Martinez and Taylor, 1996]. Three distinct hydrothermal fields have been identi-fied in the Manus Basin: the Vienna Woods field (Location 12 in figure 2) in the rift valley of the central spreading basin [Lisitsyn et al., 1993], the DESMOS caldera (Location 14 in figure 2) [Gamo et al., 1993, 1997], and the PAcMANuS field (Location 13 in figure 2) in the eastern spreading center. Based on seafloor observations and the geochemi-cal characterization of superheated hydrothermal fluid in the three hydrothermal fields during the DSRV Shinkai 2000 expedition in 1995, Gamo et al. [1996] described the hydrothermal systems in the Manus Basin as follows: (1) the Vienna Woods hydrothermal system, hosted by a basal-tic lava seafloor, is comparable to MOR systems; (2) the PAcMANuS field, hosted by an andesitic lava seafloor, has hydrothermal fluids with lower pH, lower concentrations of ca, and enriched with K, total inorganic carbon (TIc), and heavy metals in comparison with MOR and Vienna Woods hydrothermal systems; (3) the DESMOS caldera represents a novel type of hydrothermal system, fueled by superheated volcanic vapor and characterized by highly acidic hydro-thermal fluids resulting from oxidation of volatile volcanic sulfide gas (H2S) to sulfate.

The distribution of Archaea in a typical black smoker chimney from the PAcMANuS field was investigated [Takai et al. 2001]. An actively venting chimney structure was obtained from a black smoker at a depth of 1644 m (Plate 1, A and B). The in situ temperature of the effluent hydrother-mal fluid was approximately 250°c. In cross section, the chimney was divided into four different zones (Plate 1c): (1) an inner crystalline interface with a hydrothermal fluid conduit, (2) a grayish porous inner layer, (3) a black solid outer layer, and (4) a surface layer coated with orange, white, and gray mineralized crusts [Takai et al., 2001]. chemical

and mineralogical analyses revealed that the chimney con-sisted primarily of zinc sulfide, whereas the surface layer was enriched with barite [Takai et al., 2001]. Based on these characteristics, the chimney was classified as a “kuroko”-type sulfide chimney, which is distinct from the copper- and iron-rich sulfide chimneys often observed in MOR hydro-thermal systems [Iizasa et al., 1999].

The distribution of Archaea in the different layers of the chimney structure was analyzed by a variety of molecular phylogenetic techniques, including rRNA dot-slot hybrid-ization, quantitative fluorogenic PcR, T-restriction frag-ment length polymorphism, and 16S rDNA cloning and sequencing. These analyses revealed shifts of the archaeal community between the different layers over a distance of several centimeters (Plate 1c) [Takai et al., 2001]. Members of Thermococcus and the as-yet-uncultured DHVEG, described above, were the dominant archaeal phylotypes in the outer surface of the chimney (Plate 1c). The presence of Thermococcales was confirmed by a semiquantitative cultivation test [Takai et al., 2001]. Different archaeal phy-lotypes dominated the interior parts of the chimney [Takai et al., 2001]. Desulfurococcales dominated the superheated hydrothermal f luid conduit, and the major archaeal phy-lotype in the interior layers grouped with the extremely halophilic Haloarcula (Plate 1c) [Takai et al., 2001]. This was the first report of extremely halophilic Archaea in deep-sea hydrothermal environments. Although no halophilic archaeon has been isolated from deep-sea vents to date, several halophilic Bacteria have been isolated from a range of deep-sea hydrothermal environments. These include the moderately halophilic Halomonas neptunia, H. sulfidaeris, H. axialensis, and H. hydrothermalis, isolated from low-tem-perature hydrothermal fluids and sulfides [Kaye and Baross, 2000, 2004]; Idiomarina loihiensis [Donachie et al., 2003] isolated from vents on the Loihi Seamount; and Clostridium caminithermale [Brisbarre et al., 2003], first isolated from a deep-sea vent site on the Mid-Atlantic Ridge. The formation of hypersaline conditions in subseafloor environments of deep-sea hydrothermal systems has been proposed to result from phase separation and segregation of hydrothermal fluids [Kaye and Baross, 2000]. The presence of halophilic archaeal phylotypes and of diverse halophilic Bacteria in deep-sea hydrothermal vent environments suggests that hypersaline environments are present in hydrothermal sub-seafloor habitats.

4.3. Case Study: Okinawa Trough

The Okinawa Trough is a “rifting phase” BAB located between the Ryukyu Arc-Trench system and the Asian conti-nent [Letouzey and Kimura, 1986]. Since the initial discovery

Plate 1Plate 1


Plate 2. Bathymetry map of the Iheya North Knoll, including the Iheya North hydrothermal field.

Plate 1. Photographs of black smoker vents of the PAcMANuS field (A) and of a successfully recovered chimney structure (B). A sketch (c) of the substructures of chimney section and the distribution pattern of Archaea correspond-ing to the substructures.

TAKAI ET AL. 205

of submarine hydrothermal activity in the Iheya Knoll and the Izena Hole of the Mid-Okinawa Trough (MOT) [Halbach et al., 1989; Sakai et al., 1990; Glasby and Notsu, 2003], a total of six active hydrothermal fields have been identified: (1) Minani-Ensei Knoll (Location 3 in figure 2), (2) Iheya North (Location 1 in figure 2), (3) Iheya Ridge, (4) Izena Hole (Location 2 in figure 2), (5) Hatoma Knoll (Location 4 in figure 2), and (6) Yonaguni Knoll IV (Location 5 in figure 2). Several interdisciplinary investigations have been conducted in these hydrothermal fields and have focused on the geochemistry [Glasby and Notsu, 2003, for review] and the microbial ecology [Takai and Horikoshi, 1999, 2000; Takai et al., 2000, 2001, 2002, 2003a, 2004c; Inagaki et al., 2003, 2004; Nakagawa et al., 2004a, 2005a, 2005b, 2005c; Suzuki et al., 2004] of the hydrothermal fluids and sulfide deposits. The vent fields of the Okinawa Trough are hosted on felsic volcanic rocks [Ishibashi and Urabe, 1995] and thick terrigenous sediments from the Yangtze and Yellow Rivers [Narita et al., 1990]. Hydrothermal f luids in the Okinawa Trough vent fields are characterized by high con-centrations of gaseous carbon compounds [Ishibashi et al., 1990; u. Tsunogai et al., unpublished data, 2002], which are highly depleted in 13c-cH4 and moderately depleted in 13c-cO2 [u. Tsunogai et al., unpublished data, 2002], and phase separation appears to be controlling the chemistry [Kataoka et al., 2000]. Liquid cO2 and cO2 hydrates are distributed in the sediments of the vent fields [Sakai et al., 1990]. Thus, the geological, physical, and chemical fea-tures of the Okinawa Trough vent fields are characteristic of sediment-hosted, back-arc rifting systems along continental margins and are distinct from spreading centers in MOR hydrothermal systems.

The Iheya North vent field is the best characterized vent field in the Okinawa Trough. The Iheya North field is located on the northwest edge of the central depression of the Iheya North Knoll (Plate 2). Six large hydrothermal mounds, named the North Edge chimney (NEc), Event 18 (E18), North Big chimney (NBc), central Big chimney (cBc), High Radioactivity Vent (HRV), and South Big chimney (SBc), are currently recognized in this field (Plate 3). The

highest f luid temperatures (311°c) and the highest f low rates have consistently been recorded at the NBc mound, suggesting that the NBc is situated over the main conduit of hydrothermal flow. All of the mounds (except HRV) are aligned north to south with the NBc at the center (Plate 3). fluid temperatures and flow rates systematically decrease away from the NBc. In addition, the hydrothermal fluid chemistry is significantly different between the different hydrothermal mounds (Table 5). The composition of the NBc hydrothermal fluids has been relatively stable during the last 5 years, with chlorinities ranging between 75% and 100% of seawater chlorinitiy (Table 5). This range suggests that the hydrothermal fluids from NBc have experienced moderate phase separation and that the mixing ratio of vapor and brine phases is stable. In contrast, the cBc hydrother-mal fluids appear to be brine-rich, and the SBc and E18 hydrothermal fluids, which exhibit lower chlorinities, may be strongly influenced by vapor-phase input (Table 5). These variations in fluid compositions strongly suggest the occur-rence of phase separation and the remixing of the vapor and brine phases at different ratios. The mixing ratios probably depend on the local subseafloor hydrogeologic structure.

The large physical and chemical differences between the various hydrothermal mounds in the Iheya North vent field could generate numerous discrete microbial habitats that could act as a natural labratory for comparing differences in the microbial diversity and community structure. Research on the microbial diversity of the Iheya North field has been conducted with an emphasis on understanding the distribu-tion of thermophilic microorganisms present in superheated hydrothermal fluid and in sulfide chimneys from different hydrothermal mounds and characterized by physical and chemical heterogeneity [Takai et al. 2003a, 2004b; Nakagawa et al., 2005a]. culture-independent molecular phylogenetic analyses were used to characterize the microbial communi-ties in near-end-member hydrothermal fluids of the NBc (311°c), the chimney conduit walls, and IScS deployed in the chimney conduits [Takai et al., 2003a]. The microbial diversity among the different samples was very similar and consisted of bacterial and archaeal phylotypes that grouped

Plate 2Plate 2

Plate 3Plate 3

Table 5Table 5

Plate 3Plate 3

Hydrothermal fluid from

Maximum temp. (°c)

Year

Mg

mM/kg

ca

mM/kg

Na

mM/kg

K

mM/kg

Sr

μM/kg

cl

mM/kg

NBc 311 1997 0 18.1 405 73 65.9 511SBc 205 2000 0 2 8 11.8 13.3 24cBc 247 1998 0 19.9 745 79 67.4 864E18 70 1997 0 11.9 288 56.2 53.4 338Seawater 4 52.7 10.2 463 9.8 87 540Data from Kataoka et al. [2000] and Nakagawa et al. [2005a].

Table 5. End-member chemical compositions of the hydrothermal f luids obtained from various hydrothermal mounds in the Iheya North field.


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TAKAI ET AL. 207

with mesophilic ε-Proteobacteria and the as-yet-uncultured crenarcheota MGI, respectively [Takai and Horikoshi, 1999; Nakagawa et al., 2005a]. As discussed, ε-Proteobacteria are typically identified in a wide range of deep-sea hydrothemal vent environments, whereas MGI are the most abundant and widely distributed Archaea in the global ocean biosphere. Both phylotypes are characteristically found in the mixing zone between hydrothermal fluid and seawater [Takai et al., 2004b].

cultivation-based analyses of the IScS deployed in hydro-thermal chimney conduits detected hyperthermophilic Thermococcocales, which suggests the presence of an indig-enous microbial community in the superheated hydrothermal fluid of NBc [Nakagawa et al., 2005a]. furthermore, thermo-philic Archaea, such as Thermococcales, Archaeoglobales, and Methanococcales, and thermophilic and mesophilic Bacteria, including the Aquificales and ε-Proteobacteria, were frequently cultured from different layers of the NBc chimney (Plate 3) [Nakagawa et al., 2004b].

Higher density microbial communities were detected in the IScS deployed in the brine-rich cBc vent fluids (Plate 3) [Nakagawa et al., 2005a], suggesting that differences in population density between the NBS and cBc mounds may result from differences in the physical and chemical properties between the two vents. The cBc mound, with its brine-rich hydrothermal fluids, may provide more spa-tially abundant and diverse subseafloor microbial habitats than does the NBc mound. In contrast, the microbial diver-sity of the E18 vapor-rich hydrothermal fluids was distinct from the microbial diversity in both the NBc and cBc mounds (Plate 3) [Nakagawa et al., 2005a]. The microbial community discovered in the IScS deployed in the lower temperature (70°c) hydrothermal fluids of the E18 mound was mainly composed of thermophilic methanogens closely related to Methanothermococcus okinawensis [Takai et al., 2002; Nakagawa et al., 2005a]. E18 hydrothermal fluids are characterized by a vapor phase and are comparable with hydrothermal fluids from the SBc mound (Table 5), which are enriched in cO2 and cH4 [u. Tsunogai et al., unpublished data, 2002]. The SBc chemistry suggests that E18 fluids are similarly enriched. High concentrations of cO2, cH4, and perhaps H2 in the hydrothermal fluids correlate with the presence of thermophilic methanogens in the IScS deployed on the E18 mound.

The results from these analyses of microbial diversity and community structure provide compelling evidence for significant intrafield heterogeneity of the indigenous micro-bial community, which may correlate with the substantial differences in the physical and chemical properties of the high-temperature habitats of the Iheya North deep-sea hydro-thermal field. The question of correlation points to the need

for further research to improve our understanding of factors that control the distribution of microorganisms in BAB deep-sea hydrothermal vent environments.

To understand how the microbial diversity of deep-sea-vent thermophilic environments differs from the microbial diversity of planktonic communities in the surrounding seawater, Takai et al. [2004b] used culture-independent molecular phylogenetic techniques to characterize the micro-bial diversity in planktonic habitats near several of the Iheya North hydrothermal mounds. Notably, the MGI phylotypes were identified in every environment investigated, including near-end-member fluids. further, the abundance of MGI in the microbial community appeared higher in the ambient seawaters around hydrothermal plumes than in ambient deep seawater of the Mid-Okinawa Trough [Takai et al., 2004b]. These results support the hypothesis of Moyer et al. [1998] that deep-sea hydrothermal systems are potential sources or sinks of the uncultivated MGI [DeLong, 1992; Fuhrman et al., 1992].

The distribution of ε-proteobacterial phylotypes was con-siderably different from that of the MGI. ε-Proteobacteria were significantly more abundant in the hydrothermal plumes than in the seawater surrounding the plumes and were only a minor component in the ambient deep seawater planktonic community of the Mid-Okinawa Trough [Takai et al., 2004b]. The dominance of ε-Proteobacteria in the hydrothermal plumes is supported by quantitative cultiva-tion analysis [S. Nakagawa et al., unpublished data, 2005d]. Similar distribution patterns of the MGI and ε-Proteobacteria were also found in the Kairei field of the central Indian Ridge [Takai et al., 2004b], suggesting a typical distribution pattern of the planktonic microbial communities in global deep-sea hydrothermal systems.

Recently, more than 150 strains of thermophiles or chemo-lithoautotrophic mesophiles have been isolated from the Iheya North field [Takai and Horikoshi, 2000; Takai et al., 2002, 2003a; Inagaki et al., 2003, 2004; Nakagawa et al., 2005a, 2005b, 2005c]. Several of the isolates represent new genera or species. Thermosipho japonicus is an extremely thermo-philic, fermentative bacterium [Takai and Horikoshi, 2000] isolated from the NBc chimney. Another member from the Thermotogales, Thermosipho melanesiensis, was isolated from the Lau Basin [Antoine et al., 1997]. from the NBc chimney, Methanothermococcus okinawensis is the first thermophilic methane-producing archaeon isolated from a deep-sea hydro-thermal system in the western Pacific margin [Takai et al., 2002]. This archaeon is physiologically similar to M. thermoli-thotrophicus [Huber et al., 1982] but is phylogenetically asso-ciated with the mesophilic Methanococcales: Methanococcus aeolicus [Schmid et al., 1984]. In addition, many of the previ-ously uncultivated ε-Proteobacteria have been isolated from the


Okinawa Trough deep-sea hydrothermal systems, including the Iheya North field [Inagaki et al., 2003, 2004; Takai et al., 2003a; Nakagawa et al., 2005a, 2005b, 2005c]. These isolates include phylotypes related to all of the family-level phylogenetic subgroups, many of which had no representatives in culture [Takai et al., 2003a]. for example, Sulfurimonas autotrophica [Inagaki et al., 2003] within Group B; Sulfurovum lithotro-phicum [Inagaki et al., 2004] and Nitratifractor salsuginis [Nakagawa et al., 2005c] within Group f; and Thioreductor micantisoli [Nakagawa et al., 2005b] within Group G have been described as new genera. Many thermophilic strains phyloge-netically associated with the genera Hydrogenimonas [Takai et al., 2004d] or Nitratifractor [Nakagawa et al., 2005c] within Group A or the genus Lebetimonas [Takai et al., 2005a] within the order Nautiliales (Group D) [Miroshnichenko et al., 2004] have also been isolated from the Okinawa Trough hydrothermal systems [Nakagawa et al., 2005a].

4.4. Other BAB Systems

The microbial ecosystems of the deep-sea hydrother-mal fields in the Okinawa Trough are exceptionally well characterized with respect to other BAB and MOR hydro-thermal systems. However, the Okinawa Trough is not a typical example of BABs along the western Pacific margin. The Mariana Trough and the Lau Basin are the dominant, active intra-oceanic BABs and exhibit various phases of seg-ments from early rifting to mature spreading [Fryer, 1995; Hawkins, 1995].

Several microbiological research projects in the Mariana Trough are now under way. The TOTO caldera is a subma-rine volcano located in the southernmost Mariana Trough (figure 2, Location 11). Active hydrothermal venting in the depression of the TOTO caldera was discovered in 2000 with the ROV Kaiko [Gamo et al., 2004], and in 2003 the geomicrobiology of the TOTO caldera hydrothermal field was investigated by using the DSV Shinkai 6500. As a result of this latter expedition, several novel chemolithoautotrophs were isolated and characterized [Takai et al., 2004e, 2005a], and the microbial communities of the hydrothermal fluids and chimney structures were described using both culture-based and culture-independant techniques [Nakagawa et al., 2006]. Approximately 20 km northeast of the TOTO caldera, a new deep-sea hydrothermal field hosting vigor-ous black-smoker activity was discovered in 2003 (figure 2, Location 10). This deep-sea hydrothermal system is located on the off-axis volcanoes in the Mariana Trough and is comparable to hydrothermal systems recently discovered on a number of other submarine volcanoes along the volcanic front of the Mariana Arc [Embley et al., 2004] and to two previously reported axial hydrothermal fields in the central

Marina Trough (figure 2, Locations 8 and 9) [Craig et al., 1987b; Hawkins et al., 1990]. In the southern Lau Basin, three active hydrothermal fields, named Vai Lili (figure 2, Location 17), Hine Hina (figure 2, Location 18), and White church (figure 2, Location 19), were identified along the Valu fa Ridge [Fouquet et al., 1990, 1991a, 1991b, 1993]. The Vai Lili field is well-known for the unique chemistry of its hydrothermal fluids, which are characterized by very low pH and highly enriched in dissolved base metals (Zn, Pb, cu, and cd) [Herzig et al., 1993]. Most likely, the unique chem-istry of the hydrothermal fluids will support novel microbial communities and geobiological interactions. Nevertheless, except for numerical taxonomy of heterotrophs by Durand et al. [1994], the isolation of T. melanesiensis [Antoine et al., 1997], and the research on heavy metal toxicity on Lau microbes, very little work has been done to understand the community structure and dynamics in the hydrothermal fields of the Lau Basin. Ongoing and future microbiological investigations in the Mariana Trough and the Lau Basin will, no doubt, reveal microbial diversity patterns different from those observed in the Manus Basin and Okinawa Trough and will provide a more thorough understanding of the microbial ecology of the dominant, active intra-oceanic BABs of the western Pacific. further, these studies may provide impor-tant information about the biogeography of microorganisms in deep-sea hydrothermal systems globally.

5. cONcLuDING REMARKS

Much of our understanding of the microbial ecology of deep-sea hydrothermal vents stems from research focused on MOR systems. However, recent research on the microbiology of BAB) and VA hydrothermal vents reveals a greater hetero-geneity of microbial communities between vent systems than previously thought. furthermore, this heterogeneity can be directly correlated to differences in the physical and chemi-cal properties between different vent fields. By comparing MOR, BAB, and VA systems, microbiologists can begin to develop an overview of the patterns of microbial diversity in deep-sea hydrothermal systems globally. As described in this review, culture-independent methods are now rou-tinely being used in microbial ecology together with more traditional culture-dependent methods. These techniques will enable much more reliable interfield comparisons of microbial diversity among the MOR, BAB, and VA hydro-thermal fields. concurrently, microbiological investigations combined with developing technology for in situ physical and chemical measurements could link the emerging diversity and ecophysiological functions of microbial communities to the physical and geochemical variations in deep-sea hydro-thermal vent environments.

TAKAI ET AL. 209

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Joost Hoek and Anna-Louise Reysenbach, Department of Biology, Port-land State university, 1719 SW 10th Ave., Portland, Oregon 97201, uSA.

Satoshi Nakagawa, Subground Animalcule Retrieval (SuGAR) Project, Japan Agency for Marine-Earth Science and Technology (JAMSTEc), Natsushima-cho, Yokosuka 237-0061, Japan, and Department of Biology, Portland State university, 1719 SW 10th Ave., Portland, Oregon 97201, uSA.

Ken Takai, Subground Animalcule Retrieval (SuGAR) Project, Japan Agency for Marine-Earth Science and Technology (JAMSTEc), Natsushima-cho, Yokosuka 237-0061, Japan. ([email protected])

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