Bathymodiolus azoricus est un bivalve endémique des sources hydrothermales de la dorsale Médio Atlantique qui présente une forte capacité à accumuler différents métaux dans ses tissus, la qualifiant comme espèce modèle en écotoxicologie. L’objectif de ce travail a consisté à décrire les mécanismes moléculaires de tolérance et de détoxication des métaux, que ce soit au sein des populations naturelles ou durant des expositions expérimentales, par le biais d’approches chimiques (quantification des métaux) et transcriptomiques (PCR quantitative et puces à ADN), afin de comprendre comment les facteurs environnementaux pouvaient influencer l’expression des gènes de B. azoricus et d’identifier des biomarqueurs potentiels utiles pour des études écotoxicologiques en environnement extrême. Les mesures de bioaccumulation de différents métaux (Cd, Cu, Fe, Pb,…) sur des individus provenant de sites aux caractéristiques contrastées ont révélé des patrons spécifiques aux populations et aux tissus examinés chez cette espèce, suggérant des phénomènes d’organotropisme. Nos résultats suggèrent également que les bactéries symbiotiques des branchies pourraient participer à la tolérance aux métaux et aux processus de détoxication. Des tendances comparables sont observées pour l’expression relative de certains gènes candidats impliqués dans la réponse au stress métallique, par exemple les métallothionéines, les superoxyde dismutases, les ferritines et les phytochélatines. Les réponses distinctes observées pour certaines populations ou certains tissus traduisent des différences d’état physiologique qui ne seraient pas directement reliées avec l’accumulation de métaux. En outre, l’analyse des puces à ADN à une échelle globale nous a permis d’identifier les réseaux métaboliques principaux pour chaque population et chaque tissu. / Bathymodiolus azoricus is an endemic bivalve from hydrothermal vents in the Mid-Atlantic ridge, which is known to accumulate different types of metals in various tissues at high concentrations, and has therefore been granted model species for exotoxicology. The objective of this work is to describe the mechanisms of metal tolerance and detoxification as they occur in natural populations and during experimental exposures, using both chemical (metal quantification) and transcriptomic (qPCR and microarrays) approaches to understand how environmental factors influence gene expression response in B. azoricus and to identify potential biomarkers useful for ecotoxicological studies in extreme environments. Bioaccumulation of different metals (Cd, Cu, Fe, Pb,…) was measured on individuals from contrasted vent sites, revealing specific population patterns and tissue differences for this species that could be related to processes of organotropism. Our results also suggest that the symbiotic bacteria in gills may be involved in metal tolerance and detoxification. Similar variation trends are observed in the relative expression of candidate genes involved in response to metal stress, e.g. metallothioneins, superoxide dismutase, ferritin, and phytochelatin, revealing contrasted responses at population and tissue level, and reflecting differences in physiological status not directly correlated with the accumulation of metals. In addition, global scale microarray analysis allowed us to identify the principal biological pathways representative of each population and tissues.

Symbiosis is one of the most widespread evolutionary strategies on Earth. In the deep-sea, symbioses between chemosynthetic bacteria and invertebrates are abundant at hydrothermal vents and cold seeps. These mutualisms, in which symbiont carbon fixation provides for host nutrition, are analogous to the ancient endosymbioses that resulted in the chloroplast and the eukaryotic mitochondrion. However, the evolutionary processes that led to the widespread dispersal of deep-sea organisms and the mechanisms by which symbioses are initiated and maintained are poorly characterized. This thesis examined the evolution of deep-sea mussels (Bathymodiolinae) and their chemosynthetic symbionts. Bathymodioline mussel taxonomy is in need of a comprehensive systematic revision because the majority of named genera do not constitute monophyletic groups. First, this thesis demonstrated that mussels found on the Northeast Pacific Ridges are members of Adipicola, a paraphyletic genus within Bathymodiolinae, refining the evolutionary history of this poorly characterized group. Second, an updated multi-locus phylogeny of bathymodiolines was presented and used to evaluate the statistical evidence for previously proposed hypotheses describing the directional evolution of bathymodioline traits. The results indicated that patterns of directional evolution in this group are not well supported and instead suggests that trait evolution has proceeded in a non-directional manner. Third, this thesis presented the first evidence of detection and abundance of bathymodioline symbionts in the deep-sea environment, providing direct evidence that these symbionts are environmentally acquired. Fourth, this thesis presented the first multi-locus phylogenies of bathymodioline symbionts and tested the hypothesis of environmental acquisition of symbionts in this group. The results demonstrated that symbiont and host lineages are decoupled, which is consistent with the environmental acquisition hypothesis. Finally, environmental acquisition implies that symbionts have opportunities to exchange genetic information with other bacterial strains and evidence for recombination in bathymodioline symbionts is also presented. This thesis advances our understanding of the evolutionary history of bathymodioline symbioses by clarifying host and symbiont evolutionary history and symbiont transmission strategy. In aggregate, these results suggest that bathymodiolines are more flexible with regard to the habitats they inhabit and the symbionts they harbor than previously understood.

En bientôt quarante ans de recherche, de nombreuses connaissances ont été acquises sur la géologie des champs hydrothermaux, la chimie des fluides qui en réchappent et l’écologie des communautés qui les habitent. Celles-ci s’organisent en assemblages denses, distribués le long de la zone de dilution du fluide hydrothermal dans l’eau de mer, et dominés visuellement par une poignée d’espèces. La forte variabilité spatio-temporelle du fluide hydrothermal a une forte influence sur la distribution des communautés. Cependant, les mécanismes à l’origine de la réponse de la faune à cette variabilité sont peu compris. Pour cela, une approche modélisatrice est présentée. Les données collectées pendant plus de 20 ans sur l’édifice Tour Eiffel, sur le champ hydrothermal Lucky Strike (ride médio-Atlantique) ont été intégrées afin d’en extraire les composantes principales. L’étude intégrative des biomasses sur l’édifice montre que celles-ci sont largement dominées par la modiole Bathymodiolus azoricus. Ce bivalve est susceptible d’avoir une influence importante sur le fonctionnement de l’écosystème, et fait donc l’objet d’un premier modèle. La recherche de données pour le contraindre ont mené à mesurer des taux métaboliques in situ. Une fois le modèle paramétré, le modèle a fourni des estimations de flux encore inconnu. La simulation d’interruption du flux hydrothermal a fourni des indices sur la réponse de la modiole à la variabilité de son environnement. / In nearly forty years of research, significant insights have been gained on vent field geology, on the chemistry of emitted fluid and on the ecology of the communities inhabiting hydrothermal ecosystems. The fauna forms dense assemblages, distributed along the hydrothermal fluid/sea water mixing gradient, and visually dominated only by a few species. The high spatio-temporal variability of the hydrothermal fluid has a strong influence on species distribution. However, the mechanisms determining the species response to this variability is still poorly understood. In order to investigate this issue, a modelling approach is presented. Data collected for more than 20 years on the Eiffel Tower edifice, on the Lucky Strike vent filed (Mid-Atlantic Ridge) were integrated in order to identify meaningful elements for our problem. An integrative study of the faunal biomasses on the edifice showed that these are dominated by the mussel Bathymodiolus azoricus. This bivalve is likely to have a significant influence on the ecosystem functioning and is thus the object of a first model. The search for data to constrain it led to the measurement of in situ metabolic rates. Once parametrized, the model provided quantitative estimates of unknown fluxes. The simulation of hydrothermal flow interruption provided some insights on the mussel biomass response to its environment variability.

xx, 198 p. A print copy of this thesis is available through the UO Libraries. Search the library catalog for the location and call number. / "Bathymodiolus" childressi is a mixotrophic mussel from Gulf of Mexico cold seeps. There is no genetic differentiation of mussels among the seeps, suggesting wide dispersal of their larvae. This dissertation describes larval biology, ecology, and recruitment dynamics for "B." childressi. Cleavage is spiral at a rate of one per 3-9 hours, with blastula larvae hatching by 40 hours at 7-8 à à °C. At 12-14 à à °C, D-shell veligers developed by day 8 without being fed. Egg size and shell morphology indicate planktotrophy, but feeding was not observed. Embryos developed normally from 7-15 à à °C and 35-45 ppt. Although survival of larvae declined with temperature, some survived at 25 à à °C. Larval survivorship was similar at 35 and 45 ppt. Oxygen consumption increased from blastulae to trochophores and was higher for "B." childressi than for shallow-water mussel trochophores. Estimated energy content of "B." childressi eggs was greater than the energy content of shallow-water mussel eggs. An energetic model predicts that the eggs provide sufficient energy for "B." childressi trochophores to migrate into the euphotic zone. In fact, "B." childressi veligers were found in plankton tows of surface waters. The influence of recruitment on fine-scale distributions of adults at the Brine Pool cold seep was examined through manipulative field experiments. The "Bathymodiolus" childressi population at this site has a distinct bimodal size structure that shifts across an environmental gradient. New recruits of "B." childressi are abundant in the inner zone, where methane and oxygen are high and sulfide is low, leading to the inference that larvae settle preferentially there. Experiments were placed in the inner and outer zones and 2-m away from the bed. The number of larvae collected in traps did not differ among the three zones, nor did settlement density. Juveniles survived and grew in all zones, but more caged than uncaged juveniles survived. Mortality of uncaged juveniles was similar in all zones, suggesting that predation does not cause the bimodal distribution. These results suggest that the bi-modal distribution cannot be attributed to settlement preferences or juvenile mortality, but instead to migration or early post-settlement mortality. This dissertation includes my co-authored materials. / Adviser: Craig M. Young

<p>Reactive oxygen species (ROS) are produced endogenously in all aerobes and are induced by environmental stressors. ROS oxidize and disable essential cellular components such as DNA, proteins, and lipid membranes. Exposure to metals, polycyclic aromatic hydrocarbons (PAHs), and some pesticides can induce oxidative stress in marine invertebrates. All aerobic organisms have a network of antioxidants and enzymes to quench ROS and prevent oxidative damage. This dissertation examines antioxidant and oxidative stress biomarkers in endemic molluscs and crabs from two natural extreme environments: deep-sea hydrothermal vents in the Lau and North Fiji Basin, and cold seeps in the Gulf of Mexico. In addition, the acute toxicity and sub-lethal effects of four insecticides and an herbicide are examined in the estuarine blue crab, <italics>Callinectes sapidus</italics>. Blue crabs are North Carolina's most important fishery species and are frequently found in agricultural drainage ditches, an example of an anthropogenic extreme environment. </p> <p>Total glutathione, catalase, superoxide dismutase, and lipid peroxidation levels were of the same respective order of magnitude in the two vent gastropods, <italics>Alviniconcha</italics> sp. and <italics>Ifremeria nautilei</italics>, and vent mussel, <italics>Bathymodiolus brevior</italics>. These biomarkers activities were similar to those from previous reports on Mid-Atlantic Ridge mussels, except for ~100-fold higher lipid peroxidation levels among Lau molluscs. Principal component analysis (PCA) of mollusc tissue-specific biomarker levels grouped individuals by species rather than by site. </p> <p>Biomarker levels in the seep mussels <italics>Bathymodiolus childressi, B. brooksi</italics>, and <italics>B. heckerae</italics> were similar across species except for elevated foot and gill cytosolic SOD in mussels from MC-640 compared to those from AC-645. PCA of seep mussel biomarker levels differentiated by species with <italics>B. childressi</italics> isolated from <italics>B. brooksi</italics> and <italics>B. heckerae</italics>. The addition of <italics>B. brevior</italics> biomarker data to the PCA showed them grouping around <italics>B. brooksi</italics> and <italics>B. heckerae</italics>. <italics>Bathymodiolus childressi</italics> is ancestral to the other species and contains only methanotrophic endosymbionts. Whether symbionts play a role in alleviating possible toxic conditions remains unknown.</p> <p>Pesticides were acutely toxic to blue crabs in the order of Lambda-cyhalothrin > imidacloprid &#8776; aldicarb > acephate &#8776; Roundup® (glyphosate). Megalopae were almost always more sensitive to pesticides than early stage juveniles. Commercial formations of pesticides generally showed similar toxicity to active ingredients alone. Exposure to LC₂₀ levels of acephate, aldicarb, imidacloprid and Roundup significantly increased the frequency of juvenile mortality after molting. There was no significant change in total glutathione or lipid peroxidation of exposed megalopae. Lambda-cyhalothrin-, imidacloprid-, and aldicarb-based products have the potential to cause acute toxicity and molting-related mortality in shallow creeks and ditches.</p> / Dissertation

Oceanic uptake of anthropogenic CO2 emissions is causing wholesale shifts seawater carbonate chemistry towards a state of decreased carbonate ion concentration and reduced ocean pH. This change in water chemistry has potentially dire implications for marine organisms, especially those that build and maintain calcium carbonate structures. Our understanding of how ocean acidification may affect marine organisms is limited, as most studies have been short-term laboratory experiments. The CO2 flux from hydrothermal vent fluids on NW Eifuku submarine volcano (Mariana Volcanic Arc) provides a natural setting to investigate the effects of acidification. Here, the vent mussel, Bathymodiolus septemdierum, lives in water with pH as low as 5.22. This study was designed to examine the consequences of a low pH environment on reproduction, calcification and somatic growth in B. septemdierum, since the presumed elevated cost of acid-base regulation may diminish available energy for these processes. Histological analysis reveals both females and males display synchronous gametogenesis across collection sites with spawning occurring between late winter and early spring. Mussels are functionally dioecious, although evidence of protogynous hermaphroditism was found– a first record for the genus. In comparison with mussels at near normal pH, we find no evidence that the pattern of gametogenesis is affected by low pH conditions. However, calcification is compromised: at a given shell volume, shells from NW Eifuku weigh about half those from sites with near normal pH mussels. The condition index (CI = body ash free dry weight/ shell volume) was assessed in mussels collected from four low pH sites on Northwest Eifuku and two control sites from Lau Basin and Nifonea Ridge; we show that low pH conditions negatively affect CI, especially when energy availability is limited. Bathymodiolus septemdierum acquires energy from chemoautotrophic symbionts in the specialized gill epithelial cells. Using a gill condition index (GCI = gill ash free dry weight/ shell volume) and transmission electron microscopy to determine symbiont abundances in gill tissues, we show that NW Eifuku mussels with healthy gills and abundant symbionts have a higher CI than mussels from NW Eifuku with unhealthy gills. Optimal environmental sulphide concentrations appear to sustain higher symbiont abundances. While the survival of mussels on NW Eifuku is remarkable, it can come at a considerable cost to body and shell condition when during periods of energy limitation. Bathymodiolus septemdierum shows substantial resilience to low pH conditions when energy availability is sufficient due to energy budget adjustments that maximize fitness. / Graduate / 0329, 0472, 0416

L'importance des symbioses dans l'évolution du vivant est désormais admise et les associations symbiotiques sont observées dans une grande diversité d'habitats. Notre étude porte sur une symbiose au sein d'un écosystème réduit, les sources hydrothermales de l'océan profond. Bathymodiolus azoricus est un bivalve hydrothermal vivant le long de la ride Médio-Atlantique, qui héberge dans des cellules branchiales spécialisées, deux types de γ-protéobactéries différentes : des méthanotrophes (MOX) et des sulfo-oxydantes (SOX). Ces dernières sont capables d'oxyder les composés réduits présents dans le fluide hydrothermal fournissant ainsi énergie et/ou source de carbone à leur hôte. Cette double endosymbiose est plastique ainsi, l'abondance relative du type de symbionte hébergé (SOX vs. MOX) varie en fonction des concentrations en composés réduits présent dans le milieu (H2S, CH4). L'objectif de ce travail de thèse est d'identifier les acteurs moléculaires impliqués dans l'acquisition, le maintien et la régulation des bactéries symbiotiques. Pour ce faire, une analyse OMICs globale (protéomique -nano LC-MS/MS- et transcriptomique -micro-array-) a été mise en ¿uvre sur des individus symbiotiques issus de population naturelle (site hydrothermal Lucky Strike, -1700m) et sur des individus ayant expérimentalement perdu ou maintenu leurs symbiotes. Suite à cette approche globale et exploratoire, une approche plus spécifique a été menée sur des familles de protéines impliquées dans des processus immunitaire et/ou d'interactions hôte/symbiotes. Cette thèse apporte un éclaircissement sur les mécanismes régissant les relations et la communication hôte/symbiote. / Hydrothermal vents are located on the mid-ocean ridges, and are characterized by challenging physico-chemical conditions. Despite these conditions dense hydrothermal communities develop down around hydrothermal fluid emissions. The presence of marine invertebrates relies on their capacity to cope with these challenging factors, and, for those forming most of the biomass, on their ability to live in symbiosis with chemoautotrophic bacteria. Bathymodiolus azoricus is one of these symbiotic species that harbors two types of γ-proteobacteria, a sulfide-oxidizing bacterium (SOX) (using the oxidation of H2S as the source of energy and CO2 as source of carbon) and a methane-oxidizing bacterium (MOX) (that uses the oxidation of CH4 as both a source of energy and carbon). These bacteria are located in specific epithelial cells in the gill tissue of the mussel. The proportion and number of these symbiont types (SOX vs. MOX) in B.azoricus can change in response to environmental conditions, and especially on the relative concentration of reduced compounds. The aim of our study is to understand the molecular mechanisms of acquisition, regulation and maintenance of the symbiotic charge in B .azoricus gills. We therefore, performed a global OMICs analysis (proteomics –nano LC-MS/MS and transcriptomics- micro-array) on mussels from natural population (Lucky Strike, -1700m) and on mussels that experimentally loose or maintain their symbiotic rate. This exploratory approach was followed by a more specific approach on family of proteins involved in immunity process and/or in host/symbiont interactions. This PhD provides hypotheses on the mechanisms governing the relationship and communication between host and symbionts.