**6. Concluding remarks**

of methane and sulfur [56]. The physiological adaptation to aquarium environment is likely to be aggravated by the expelling of endosymbionts into the aquarium environment, progres‐ sively emptying the gill tissue of its autotrophic bacteria, essential for the host vent mussel nutritional sustenance. Long‐term aquarium acclimatization represents thus a model study to investigate the presence and maintenance of symbiotic associations between chemosynthetic bacteria and vent animals, which depend on controlled cell‐cell communication between host

Presumably, the loss of endosymbiont induces a dramatic change in host gene expression profiles especially if endosymbiont genes exert some transcriptional control over host gene expression. For this reason, acclimatization studies have been instrumental to further our understanding of *B. azoricus* immune system. These studies have provided insights into physiological principles underlying mechanisms of adaptation to aquarium conditions at sea level pressure while taking advantage of the remarkable capacity of vent mussels to survive well decompression once brought to surface [21, 22, 56]. Furthermore, these studies have allowed analyses using immune challenged mussels comparatively to acclimatized control mussels, maintained under aquarium conditions. In view of our previous experiments per‐ formed with live gill tissues and postcapture immune gene expression studies in *B. azoricus* acclimatized to atmospheric pressure, the presence of endosymbiont bacteria is now being under investigation as a driving factor under which host‐immune genes may transcription‐ ally be modulated and reciprocally endosymbiont genes may transcriptionally be modulated by the host [53–56]. Moreover, the impact of aquarium acclimatization on *B. azoricus* immune responses and its capacity to react to *V. diabolicus* challenges was recently evaluated dur‐ ing recurrent incubations with *V. diabolicus* during short periods of time, followed by clean sea water incubations allowing animals to depurate and subsequently be reexposed to the same load of *V. diabolicus* over a period of 3 weeks acclimatization experiment [74]. As pre‐ viously described, we found a time‐dependent immune gene response in *B. azoricus* tied to the endosymbiont presence inside the vent mussel gills. The vent mussel's immune defense capabilities were affected by the gradual loss of symbiont bacteria suggesting a symbiont‐ mediated defense mechanism under which the transcriptional regulation of host immune genes is directly affected by symbiont density and/or activity. The host‐immune system‐ endosymbiont interactions were actively higher during the first week of acclimatization as a result of *Vibrio* exposures, demonstrating the ability of *B. azoricus* to increase the transcrip‐ tion of immune genes while endosymbiont gene expression also correlated with an increased symbiotic metabolism and prevalence. A synergistic response was proposed to counteract the presence and potential infection by *V. diabolicus* bacterium while modulating *B. azoricus* immune defenses‐endosymbiont interactions to an extant, which host‐immune and endosym‐ biont genes are mutually reliant during the first weeks of acclimatization [74]. The evidence presented suggests successful *V. diabolicus* recognition prompting immune genes to increase their levels of transcriptional activity particularly for genes involved in the Toll‐like receptor signaling [75, 76] and apoptosis‐related pathways [77] during first day of acclimatization in aquarium environments. In agreement with this, *B. azoricus* is presented as a suitable model to study molecular interactions involving host‐mediated immune recognition events and adap‐ tation mechanisms, to mitigate apoptosis harmful effects induced by *Vibrio* exposure against which, endosymbionts were prompted to increase their transcriptional activity, evocative of a possible protection role to the host [74]. This work brings to light other questions relating to

and endosymbionts and the role of the host immune system [56, 74].

176 Organismal and Molecular Malacology

In an attempt to understand physiological reactions of animals normally set to endure extreme conditions, in deep‐sea environments, our laboratory has undertaken, for the last 6 years, a series of investigations aimed at characterizing molecular indicators of adaptation processes of which components of the immune and antioxidative stress response systems have received most of our attention. As a research goal, long‐term maintenance of vent mussels to atmo‐ spheric pressure was instrumental to further our understanding on molecular relationships under which the vent mussel‐endosymbiont interactions are affected by aquaria conditions and by the gradual disappearance of endosymbiont bacteria from gill epithelia. Hence, the maintenance of live mussels in our aquarium laboratory system has been a key factor in gain‐ ing knowledge into the physiology of vent animals including the study of evolutionary con‐ served immune, inflammatory, and stress‐related factors commonly found in other marine bivalves. *In vivo* and *ex vivo* experiments conducted with live mussels and their excised gill tissues as primary tissue cultures, allowed the specific host‐endosymbiont interactions to be revealed, and further characterized in the deep‐sea vent model *B. azoricus*, establishing dis‐ tinct genetic signatures for the expression of endosymbiont genes and host‐immune genes in relation to different environmental conditions. Increasing evidence now support the role of gills as a *bone fide* immune‐responsive tissue in *B. azoricus*, consistent with a suitable study model for exploring molecular interactions involving host‐endosymbiont‐mediated immune recognition events and adaptation mechanisms to deep‐sea hydrothermal vent environments. Such adaptation mechanisms are likely to be influenced by the microbial community com‐ position surrounding the mussel beds at hydrothermal vents and therefore it is important to continue metatranscriptomic and metagenomic studies [79] from the gill‐associated microbial diversity and surrounding hydrothermal vent sediments [80, 81] in view of the broader eco‐ logical organization and evolutionary importance of animal‐bacterial microbiomes in chemo‐ synthetic‐based ecosystems in the deep sea [82, 83].

In recent years, researchers have turned to the human microbiome for its functional role in human health [84] and both composition and alterations in the microbiome have been found associated with diabetes, inflammatory bowel disease, obesity, asthma, rheuma‐ toid arthritis, and susceptibility to infections [85]. Other microbiomes from nonmamma‐ lian and nonvertebrate species have also been characterized, for instance in insects where it was found to be highly dependent on the environment, species, and populations and affecting the fitness of species. These fitness effects may have important implications for the conservation and management of species and populations [82, 83]. Given the temporal instability of deep‐sea hydrothermal vents and their constant fluctuations of physical and chemical environmental conditions, vent animal‐microbiome associations have become critical for our understanding of invasion of nonnative species, responses to pathogens, and responses to chemicals and global climate change in the present and future [82] par‐ ticularly when deep‐sea mining activities are projected to have a major impact on deep‐sea vent ecosystems [86].
