Molecular cloning of superoxide dismutase (Cu/Zn-SOD) from aquatic molluscs

The potential of the first line of the active oxygen-scavenging system, partial cDNA encoding Cu/Zn superoxide dismutase (SOD) was isolated in three aquatic mollusc species: Ruditapes decussatus (marine clam), Dreissena polymorpha (continental water mussel) and Bathymodiolus azoricus (hydrothermal vent mussel). These SOD cDNA fragments were amplified by PCR with degenerate oligonucleotide primers derived from the amino acid sequence conserved in the Cu/Zn-SOD from several other organisms. A partial cDNA of CuZn-SOD was obtained for R. decussates (510 bp), D. polymorpha (510 bp) and B. azoricus (195 bp). The deduced amino acid sequence showed high similarity among the three mollusc species (57-63%) and among other species (50-65%). The residues involved in coordinating copper (His-47, 49, 64, 121) and zinc (His-64, 72, 81 and Asp-84) were well conserved among the three Cu/Zn-SOD sequences.     

Heat shock protein expression pattern (HSP70) in the hydrothermal vent mussel Bathymodiolus azoricus

We previously reported evidence of increased levels of DNA damage in the hydrothermal mussel Bathymodiolus azoricus, which suggested that the species was not fully resistant to the natural toxicity of its deep-sea vent environment. In the present study, HSP70 was used as a biomarker of sub-cellular stress. Differences in HSP70 expression pattern were observed between vent sites, typified by different depths/toxicity profiles, and between different mussel tissue types. A comparison of specimens collected by remote operated vehicle (ROV) and acoustically-operated cages showed that less stress (as indicated by changes in HSP70 levels) was induced by the faster cage recovery method. Therefore alternatives to ROV collection should be considered when planning experiments involving live deep sea organisms. Significantly, a positive correlation was found between the levels of DNA strand breakage, as measured using the Comet assay, and HSP70 expression pattern; evidence was also obtained for the constitutive expression of at least one HSP isoform which was located within the cell nucleus.     

Spatial variation of metal bioaccumulation in the hydrothermal vent mussel Bathymodiolus azoricus

The variability of the bioaccumulation of metals (Ag, Cd, Cu, Fe, Mn and Zn) was extensively studied in the mussel Bathymodiolus azoricus from five hydrothermal vent sites inside three main vent fields of increasing depth along the Mid-Atlantic Ridge: Menez Gwen, Lucky Strike and Rainbow. Metal bioaccumulation varied greatly between vent fields and even between sites inside a vent field with B. azoricus showing a great capacity to accumulate metals. The bioaccumulation of these metals also varied significantly among tissues. The main target was the gills where metals were mainly associated with soluble compounds whereas in the digestive gland they were mainly associated with insoluble compounds. Storage of metals under insoluble forms in B. azoricus seems to be a major pathway for the detoxification of both essential and non-essential metals. Mussels from the studied fields can be discriminated following their metallic load but the segregation relies partially on the composition of the metal-enriched fluids.     

Effect of cadmium, copper and mercury on antioxidant enzyme activities and lipid peroxidation in the gills of the hydrothermal vent mussel Bathymodiolus azoricus

Metals are known to influence lipid peroxidation and oxidative status of marine organisms. Hydrothermal vent mussels Bathymodiolus azoricus live in deep-sea environments with anomalous conditions, including high metal concentrations. Although B. azoricus are aerobic organisms they possess abundant methano and thioautotrophic symbiotic bacteria in the gills. The enzymatic defences (superoxide dismutase (SOD), catalase (CAT), total glutathione peroxidase (Total GPx) and selenium-dependent glutathione peroxidase (Se-GPx)) and lipid peroxidation were determined in the gills of B. azoricus exposed to Cd (0.9 microM), Cu (0.4 microM) and Hg (0.1 microM) with different times of exposure. The experiments were performed in pressurized containers at 9+/-1 degrees C and 85 bars. Results show that vent mussels possess antioxidant enzymatic protection in the gills. Cd and Cu had an inhibitory effect in the enzymatic defence system, contrarily to Hg. These enzymatic systems are not completely understood in the B. azoricus, since reactive oxygen species might be produced through other processes than natural redox cycling, due to hydrogen sulphide and oxygen content present. Also the symbiotic bacteria may play an important contribution in the antioxidant protection of the gills.     

深海热液口大型底栖生物遗传多样性的研究进展

深海热液口及其生物群落自1977年发现以来,受到海洋生物学界的广泛关注。近年来伴随深海调查技术的进步,大量热液生物群落相继被发现,并在生物地理学、生态学和生物多样性方面取得了显著的进展。目前,受采样不足及分类学研究的限制,针对深海热液口的遗传多样性研究多集中于大型底栖生物。本文拟对该领域的研究成果进行简要综述,以期加深对深海热液口大型底栖生物的物种分布模式、扩散途径及群体连通性的认识,为深海热液口生物多样性的研究提供新视角。     

Peripheral communities of the Eastern Lau Spreading Center and Valu Fa Ridge: community composition,temporal change and comparison to near‐vent communities

Western Pacific hydrothermal vents will soon be subjected to deep‐sea mining and peripheral sites are considered the most practical targets. The limited information on community dynamics and temporal change in these communities makes it difficult to anticipate the impact of mining activities and recovery trajectories. We studied community composition of peripheral communities along a cline in hydrothermal chemistry on the Eastern Lau Spreading Center and Valu Fa Ridge (ELSC‐VFR) and also studied patterns of temporal change. Peripheral communities located in the northern vent fields of the ELSC‐VFR are significantly different from those in the southern vent fields. Higher abundances of zoanthids and anemones were found in northern peripheral sites and the symbiont‐containing mussel Bathymodiolus brevior, brisingid seastars and polynoids were only present in the northern peripheral sites. By contrast, certain faunal groups were seen only in the southern peripheral sites, such as lollipop sponges, pycnogonids and ophiuroids. Taxonomic richness of the peripheral communities was similar to that of active vent communities, due to the presence of non‐vent endemic species that balanced the absence of species found in areas of active venting. The communities present at waning active sites resemble those of peripheral sites, indicating that peripheral species can colonize previously active vent sites in addition to settling in the periphery of areas of venting. Growth and mortality were observed in a number of the normally slow‐growing cladorhizid stick sponges, indicating that these animals may exhibit life history strategies in the vicinity of vents that differ from those previously recorded. A novel facultative association between polynoids and anemones is proposed based on their correlated distributions.     

Adaptation of the antioxidant defence system in hydrothermal-vent mussels (Bathymodiolus azoricus) transplanted between two Mid-Atlantic Ridge sites

The vent mussel Bathymodiolus azoricus is the dominant member of the Northern Mid‐Atlantic Ridge (MAR) hydrothermal megafauna, and lives in an environment characterized by temporal and spatial variations in the levels of heavy metals, methane and hydrogen sulphide, substances which are known to increase reactive oxygen species levels in the tissues of exposed organisms. To evaluate the effects of two contrasting hydrothermal environments on the antioxidant defence system of this vent mussel species, a 2‐week transplant experiment was carried out involving mussels collected from the relatively deep (2300 m), and chemical rich, Rainbow vent field. These were transplanted to the shallower (1700 m), and relatively less toxic, Lucky Strike vent field. To achieve this objective, levels of superoxide dismutase, catalase (CAT), total glutathione peroxidase (GPx), selenium‐dependent glutathione peroxidase and lipid peroxidation (LPO) were measured in the gills and mantle tissues of resident and transplant mussels before and after the transplant experiment. With the exception of CAT, the gills of the transplanted mussels had significantly higher antioxidant enzyme activity compared with the basal levels in the donor (Rainbow) and recipient (Lucky Strike) populations; whereas the antioxidant enzyme levels in the mantle tissues of the transplants reflected the baseline levels of activity in the native Lucky Strike mussels after 2 weeks. In contrast, LPO levels were significantly higher in both tissue types in the transplants than in either the source or the recipient populations, which suggested a response to hydrostatic pressure change (note, the transplant animals were brought to the surface for transportation between the two vent fields). The fact that the Rainbow mussels survived the transplant experience indicates that B. azoricus has a very robust constitution, which enables it to cope behaviourally, physiologically and genetically with the extreme conditions found in its naturally contaminated deep‐sea environment.     

A fungal epizootic in mussels at a deep-sea hydrothermal vent

Mass mortalities due to disease are important determinants of population and community structure in marine ecosystems, but the speed at which an epizootic may sweep through a population, combined with rapid selection for disease‐resistant stocks, can mask the ecological impact of disease in all but the most closely monitored populations. We document an emergent epizootic event in the deep sea that is occurring in mussels (Bathymodiolus brevior) at the Mussel Hill hydrothermal vent in Fiji Basin and we identify the causal agent as a black yeast (order Chaetothyriales) that elicits a pronounced host immune response and is associated with tissue deterioration. The yeast was not observed in other invertebrate taxa (the gastropods Ifremeria nautilei, Alviniconcha aff. hessleri; the limpets Lepetodrilus schrolli, Symmetromphalus aff. hageni; the polychaetes Branchipolynoe pettiboneae, Amphisamytha cf. galapagensis) associated with the mussel bed, nor in mussels (Bathymodiolus brevior) collected from adjacent Lau Basin mussel beds. Massive mussel mortality resulting from the fungal infection is anticipated at the Mussel Hill site in Fiji Basin; we expect that epizootic outbreaks in dense invertebrate communities have the potential to be major determinants of community structure in deep‐sea chemosynthetic ecosystems. The possibility that submersible assets may serve as vectors for transport of the fungus warrants further attention.     

Hydrothermal vent meiobenthos associated with mytilid mussel aggregations from the Mid-Atlantic Ridge and the East Pacific Rise

Deep-sea hydrothermal vents occur along the mid-ocean ridges and back-arc basins around the globe. There are very few community analyses of vent meiobenthos. The central objectives of this study were to identify and quantify for the first time the entire metazoan meiobenthic community associated with mussel aggregations of Bathymodiolus thermophilus Kenk and Wilson, 1985 from the EPR, 11°N and of Bathymodiolus puteoserpentis Cosel et al., 1994 from the Mid-Atlantic Ridge (MAR), 23°N. Using a quantitative sampling method, abundance, biomass, sex ratio, species richness, diversity, evenness, and trophic structure were studied based on three samples from each site. Meiobenthic abundance in each sample was unexpectedly low, but similar between sites. The community was composed of nematodes, copepods, ostracods, and mites, with a total of 24 species at EPR vents, and 15 species at MAR vents. While most copepod species were vent endemics within the family Dirivultidae, nematodes and harpacticoid copepods belonged to generalist genera, which occur at a variety of habitats and are not restricted to hydrothermal vents or the deep sea. The meiobenthos of hydrothermal-vent mussel beds constitutes a unique community unlike those of other sulfidic habitats, including the thiobios of shallow-water sediments and the meiobenthos of deep-sea, cold-seep sediments. The trophic structure was dominated by primary consumers, mainly deposit feeders, followed by parasites. Predatory meiofaunal species were absent.     

Variations in deep-sea hydrothermal vent communities on the Mid-Atlantic Ridge near the Azores plateau

Near the Azores Triple Junction as the Azores Plateau is approached, the ridge axis becomes shallower; its depth decreases from ca. 2400 m in the Rainbow vent field (36°13′N) to ca. 850 m in the Menez Gwen vent field (37°35′N). In this area, extensive mussel beds of the mytilid Bathymodiolus azoricus dominate the hydrothermal vent fauna, along with populations of three shrimps (Rimicaris exoculata, Mirocaris fortunata and Chorocaris chacei). The main physical and chemical characteristics of the vent habitat were studied by discrete sampling, in situ analysis and sediment trap moorings. The vent fauna is distributed along a variable band where the vent fluids and seawater mix, with R. exoculata living in the most concentrated areas and Bathymodiolus azoricus in the most diluted zones. Various non-endemic species live at the border of the vent field. The variations observed in structure and composition of the communities along the depth gradient are most likely due to changes in vent fluid toxicity (metallic and sulphide content) and suspended mineral particles, which render the fluids harsher for species living there. The main faunal differences observed between Lucky Strike and Menez Gwen hydrothermal fields are due to an impoverishment in the hydrothermal endemic species and to the penetration of bathyal species. The comparison of the three studied vent fields suggests the existence of a succession of several biogeographic islands rather than a single province.     

High-resolution dynamics of a deep-sea hydrothermal mussel assemblage monitored by the EMSO-Açores MoMAR observatory

Although the spatial distribution of hydrothermal vent assemblages in relation to environmental conditions has been assessed in several studies, there is little documented data on the temporal variation of the fauna and corresponding abiotic factors in a vent community. Here, we present one of the longest integrated (faunal and environmental data) time series ever obtained in a hydrothermal ecosystem. The data were acquired using the TEMPO ecological module that was deployed between 2006 and 2008 on the Mid-Atlantic Ridge, providing the first insights into the day-to-day variations in a Bathymodiolus azoricus mussel assemblage from the Lucky Strike vent field for the 48 days during which the video camera operated. The time-series yielded additional valuable information on longer-term variation in faunal distribution (comparing ~2 years), temperature (11.7 months) and iron concentrations (3.8 months).Results from daily observations showed that the vent mussel assemblage was quite stable over the 48 days of the study, reflecting the relative stability of environmental conditions during this period. B. azoricus mussels appeared to thrive in areas of very limited hydrothermal fluid input in habitats that are, as in other deep-sea ecosystems, significantly influenced by ocean tidal signals. Variation in species abundance was observed but, with the exception of Mirocaris fortunata shrimp, no links could be established with measured environmental variables. Although we did not observe any clear tidal influence on vent fauna, it is likely that physiological processes and species’ activities are influenced by these periodic variations. Longer time series are currently being acquired by different experiments deployed on the EMSO-Açores MoMAR observatory (2010–2013 and still recording). They should further improve our knowledge of the dynamics of hydrothermal systems and their associated faunal communities.     

Adaptation to metal toxicity: a comparison of hydrothermal vent and coastal shrimps

Rainbow vent field is one of the most metal‐contaminated hydrothermal sites on the Mid‐Atlantic Ridge near the Azores region. Two hydrothermal shrimp species dominate the fauna at the Rainbow site along with the mussel Bathymodiolus azoricus. Although the levels of essential and non‐essential metals in these shrimps have been studied, the biological consequences of a metal‐rich environment are still largely unknown. Therefore, the aim of this study was to determine the levels of metal‐binding proteins – metallothioneins (MT) and the activities of antioxidant enzymes – superoxide dismutase, catalase, total glutathione peroxidase and selenium‐dependent glutathione peroxidase in two hydrothermal vent shrimps (Mirocaris fortunata and Rimicaris exoculata) collected from the Rainbow site and to compare them with two coastal shrimps (Palaemon elegans and Palaemonetes varians) from a south Portugal lagoon (Ria Formosa) to evaluate their different adaptation strategies towards metals in their environment. Results show significant differences in MT levels and antioxidant enzymatic activities between vent and coastal shrimps and also between shrimp species collected from the same site. This suggests that biochemical responses in both vent and coastal shrimps are affected not only by the environmental characteristics but also by inter‐specific differences. Nevertheless, these responses apparently confer successful adaptation for survival in a metal‐extreme environment.     

Metal concentrations in the shell of Bathymodiolus azoricus from contrasting hydrothermal vent fields on the Mid-Atlantic Ridge

Specimens of Bathymodiolus azoricus were sampled along the Mid-Atlantic Ridge at the Menez Gwen, Lucky Strike and Rainbow hydrothermal fields. Individual shells (n = 51), through the weight range 0.62 to 15.70 g, were analyzed for their magnesium, strontium, iron, manganese, copper, zinc and cadmium concentrations. Amongst the marine molluscs the shell of B. azoricus is confirmed as being particularly impoverished in strontium (mean 943 microg g(-1)). Trace metal concentrations in the shells decreased in the order Fe> Mn> Zn> Cu> Cd. Despite originating from trace metal rich environments mean concentrations were low (37.9, 13.2, 10.7, 1.1 and 0.7 microg g(-1), respectively). Irrespective of geographical origin magnesium, strontium and copper concentrations were primarily dictated by shell weight. In contrast cadmium concentrations were elevated in shells from the Rainbow field and ambient seawater chemistry imparted site specific chemical fingerprints to the shells with respect to the iron to manganese ratio.     

Microbial Diversity of Deep-sea Sediments from Three Newly Discovered Hydrothermal Vent Fields in the Central Indian Ridge

Since the discovery of hydrothermal vents in the late 1970s, deep-sea hydrothermal vent fields have attracted great attention as biological hotspots. However, compared with other ocean ridges, the structure and function of microbial communities inhabiting vent fields in the Central Indian ridge (CIR) remain understudied. Here, we provide for the first time 16S rRNA gene-based comparative metagenomic analysis of the sediment-associated microbial communities from three newly discovered vent fields in the CIR. Sediment samples collected in the Invent B, Invent E and Onnuri vent fields varied in geochemical properties, elemental concentrations and associated microbial communities. Proteobacteria (Gammaproteobacteria) was the dominant phylum in Invent B and Onnuri vent fields. In contrast, Invent E mainly consisted of Chloroflexi and Euryarchaeota. Predicted functional profiling revealed that the microbial communities in the three vents are dominated by chemoheterotrophic functions. In addition, microbial communities capable of respiration of sulfur compounds, nitrification, nitrite oxidation, methylotrophy, and methanotropy were found to be the main chemolithoautotrophs. Compared to other vent fields, Invent E showed a predominance of archaeal methanogens suggesting it exhibits slightly different geochemistry. Multivariate analysis indicated that the biogeochemical and trace metal differences are reflected in the sediment microbial compositions of the three vent fields. This study expands our current understanding of the microbial community structure and potential ecological functions of the newly discovered hydrothermal vent fields in the CIR.

     

Biological data extraction from imagery – How far can we go? A case study from the Mid-Atlantic Ridge

In the past few decades, hydrothermal vent research has progressed immensely, resulting in higher-quality samples and long-term studies. With time, scientists are becoming more aware of the impacts of sampling on the faunal communities and are looking for less invasive ways to investigate the vent ecosystems. In this perspective, imagery analysis plays a very important role. With this study, we test which factors can be quantitatively and accurately assessed based on imagery, through comparison with faunal sampling. Twelve instrumented chains were deployed on the Atlantic Eiffel Tower hydrothermal edifice and the corresponding study sites were subsequently sampled. Discrete, quantitative samples were compared to the imagery recorded during the experiment. An observer-effect was tested, by comparing imagery data gathered by different scientists. Most factors based on image analyses concerning Bathymodiolus azoricus mussels were shown to be valid representations of the corresponding samples. Additional ecological assets, based exclusively on imagery, were included.     

Bioaccumulation des métaux chez le mytilidae hydrothermal Bathymodiolus sp. de la ride médio-atlantique

The hydrothermal environment is characterised, among other things, by a high metal concentration, related to the convective sea-water circulation inside the oceanic crust and its interaction with basaltic rocks. The biological communities associated with the hydrothermal system can survive in this toxic environment owing to their ability to regulate their intracellular metal levels by excretion or accumulation of metal ions in non-toxic forms. Various detoxification processes have been previously studied within an hydrothermal organism: immobilisation and precipitation of the metal in lysosomal systems, or binding to specific and soluble ligands such as metallothioneins. The later are heat stable and characterised by a high content of cysteine. The quantification of Fe, Zn, Cu, Mn, Cd, Ag, Ba and Sr in the tissues of the vent mussel Bathymodiolus sp. indicates a high accumulation of these metals in two target organs, the gill and the digestive gland. This accumulation is in relation with high concentrations of metallothioneins, but the subcellular distribution of metals indicates a higher contribution of the insoluble compartment for detoxification processes. These results are compared with the literature data about metal bioaccumlation in Bathymodiolus thermophilus collected at the Galapagos Rift.     

Temporal variation in the antioxidant defence system and lipid peroxidation in the gills and mantle of hydrothermal vent mussel Bathymodiolus azoricus

Hydrothermal vent mussels are exposed continually to toxic compounds, including high metal concentrations and other substances like dissolved sulphide, methane and natural radioactivity. Fluctuations in these parameters appear to be common because of the characteristic instability of the hydrothermal environment. Temporal variation in the antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), total glutathione peroxidases (Total GPx), selenium dependent glutathione peroxidases (Se-GPx)), metallothioneins and lipid peroxidation (LPO) in the gills and mantle of the mussel Bathymodiolus azoricus from Menez–Gwen hydrothermal vent site was evaluated and related to the accumulated metal concentrations (Ag, Cu, Cd, Fe, Mn and Zn) in the tissues. Maximum antioxidant enzyme activities in the gills were detected in the beginning of summer, followed by a gradual decrease throughout the following months. One year after, the levels of antioxidant enzyme activities were similar to those reported one year before. LPO in this tissue exhibited a similar temporal variation trend. A different pattern of temporal variation in antioxidant enzyme activities was observed in the mantle, with a gradual increase from summer to the end of autumn (November). LPO in the mantle exhibited an almost reverse trend of temporal variation to that of antioxidant enzyme activities in this tissue. Antioxidant defences in the gills of B. azoricus were significantly enhanced with increasing concentrations of Ag, Cu and Mn, while negative relationships between antioxidant enzymes and Cd, Cu, Mn and Zn concentrations in the mantle were observed, suggesting different pathways of reactive oxygen species (ROS) production and that these tissues responded differently to the metal accumulation. However, temporal variation in biomarkers of defence and damage were in general similar to coastal bivalve species and can be associated with temporal variations of the physiological status due to reproduction. These variations might also be linked to the highly unstable nature of the hydrothermal environment.     

热液和冷泉10种无脊椎动物基因组大小测定及比较

基因组大小(或称C值)作为生物单倍体细胞中全套染色体的DNA总量,在一定程度上是恒定的,因而C值可以作为生物物种的一个特定参数。深海热液和冷泉为更好地理解C值与不同环境之间的关系提供了一个特征性的模型。本文采用流式细胞术,测定了来自热液和冷泉环境中的10种深海无脊椎动物的C值,其分布范围从0.87 pg到12.28 pg,其中,相比于软体动物和多毛类,甲壳生物基因组大小及变异均较大。对比热液和冷泉两个群落中共有种(深海偏顶蛤Bathymodiolus platifrons、柯氏潜铠虾Shinkaia crosnieri以及长角阿尔文虾Alvinocaris longirostris)的基因组大小,发现C值差异并不显著。同时,综合已有的数据,对深海化能极端环境与其他环境条件下的物种C值进行对比分析,结果显示深海化能极端环境下生物的基因组大小并没有发现明确的变化趋势。     

Biomass and production of bacterioplankton at the hydrothermal vent areas in the rift system of the North Fiji Basin

Mesoscale distribution of the density and productivity of bacterioplankton at representative geological locations in the rift ecosystem of the North Fiji Basin were clarified by sampling using a mannedShinkai 6500 submersible. The bacterioplankton density inside the hot and warm fluids of hydrothermal vents was one order magnitude greater than that in the deep water outside the vent plume. The bacterial production was highest inside the hot and warm vent fluids, and was higher just outside the hot and warm vent fluids than further outside in the deep water surrounding the vent plume, respectively. Productivity from outside to inside the vent fluid varied from 0.01 to 0.71 gC m^–3 day^–1.     

Biogeographic relationships among deep-sea hydrothermal vent faunas at global scale

The discovery of deep-sea hydrothermal vent fauna, kilometres deep in the oceans, is a great achievement of 20th-century marine biology. The deep-sea hydrothermal food web does not directly depend on the sun's energy. Vent communities rely primarily on trophic associations between chemoautotrophic bacteria and consumers. A small number of endemic taxa are adapted to the inhospitable vent environments that are distributed along ridge crests. Where these vent communities originated and how they dispersed are among the important questions ecologists must answer. Here, by statistical analysis of the most comprehensive database ever assembled about deep-sea hydrothermal fauna, we delineate six major hydrothermal provinces in the world ocean and identify seven possible dispersal pathways between adjacent provinces. Our model suggests that the East-Pacific Rise may have played a pivotal role as a centre of dispersal for the hydrothermal fauna. Our data-driven conclusion will have to be tested by phylogenetic studies and completed by surveys of less-explored fields.