Étude de l'homéostasie des micronutriments de la fixation d'azote au sein de la symbiose lichénique en forêt boréale

L’azote est un des éléments les plus importants dans la nature. Sa disponibilité limite la productivité d’un grand nombre d’écosystèmes naturels, et influencera sans doute de manière importante leurs réponses aux changements climatiques globaux. La première source d’azote dans les écosystèmes non anthropisés est la fixation biologique de l’azote. Ce processus repose sur un groupe de métallo-enzymes spécifiques, les nitrogénases, dont le cofacteur métallique contient soit du fer et un atome de molybdène, soit du fer et un atome de vanadium, soit uniquement du fer. A ce jour, seule la nitrogénase au molybdène est prise en considération dans la dynamique de l’azote dans les écosystèmes, et ce malgré de nombreux indices indiquant que la nitrogénase au vanadium pourrait avoir un rôle important. Est-ce que la nitrogénase au vanadium est utilisée dans les écosystèmes naturels et quelles sont les conditions favorisant son utilisation ? Nous avons cherché à répondre à ces questions à l’aide d’un modèle symbiotique tripartite, un lichen, association entre une algue, un champignon et une cyanobactérie fixatrice d’azote. Nous avons tout d’abord développé une méthode d’étude des contenus en métaux des différents symbiontes, puis nous avons étudié la répartition et la régulation du vanadium au sein des différents symbiontes dans différentes conditions environnementales. Nous avons pu démontrer que dans ce modèle, le vanadium possède toutes les caractéristiques d’un micronutriment essentiel à la fixation d’azote. Nous avons également démontré que la disponibilité du molybdène ainsi que les températures, telles que rencontrées en milieux boréaux, seraient deux facteurs importants contrôlant l’utilisation de la V-Nase. Les résultats présentés dans cette étude apportent une meilleure compréhension de la gestion des métaux cofacteurs de la nitrogénase au sein de la symbiose lichénique. Mais ils permettent surtout de remettre en question le paradigme de l’hégémonie du molybdène sur la fixation biologique de l’azote. Ainsi, la fixation d’azote en milieu continental repose sur un ensemble hétérogène d’enzymes, ce qui autorise aux organismes fixateurs d’azote une grande flexibilité vis-à-vis des paramètres environnementaux comme les basses températures. Cela leurs permet également une meilleure adaptation au stress métallique résultant de carences en micronutriments, notamment celle en molybdène. Ces résultats invitent également à réévaluer les modèles biogéochimiques liant les cycles des micronutriments aux cycles des macronutriments, particulièrement celui de l’azote

Molybdenum threshold for ecosystem scale alternative vanadium nitrogenase activity in boreal forests

Biological nitrogen fixation (BNF) by microorganisms associated with cryptogamic covers, such as cyanolichens and bryophytes, is a primary source of fixed nitrogen in pristine, high-latitude ecosystems. On land, low molybdenum (Mo) availability has been shown to limit BNF by the most common form of nitrogenase (Nase), which requires Mo in its active site. Vanadium (V) and iron-only Nases have been suggested as viable alternatives to countering Mo limitation of BNF; however, field data supporting this long-standing hypothesis have been lacking. Here, we elucidate the contribution of vanadium nitrogenase (V-Nase) to BNF by cyanolichens across a 600-km latitudinal transect in eastern boreal forests of North America. Widespread V-Nase activity was detected (∼15–50% of total BNF rates), with most of the activity found in the northern part of the transect. We observed a 3-fold increase of V-Nase contribution during the 20-wk growing season. By including the contribution of V-Nase to BNF, estimates of new N input by cyanolichens increase by up to 30%. We find that variability in V-based BNF is strongly related to Mo availability, and we identify a Mo threshold of ∼250 ng·glichen−1 for the onset of V-based BNF. Our results provide compelling ecosystem-scale evidence for the use of the V-Nase as a surrogate enzyme that contributes to BNF when Mo is limiting. Given widespread findings of terrestrial Mo limitation, including the carbon-rich circumboreal belt where global change is most rapid, additional consideration of V-based BNF is required in experimental and modeling studies of terrestrial biogeochemistry

A diazotrophy-ammoniotrophy dual growth model for the sulfate reducing bacterium Desulfovibrio vulgaris var. Hildenborough

Sulfate reducing bacteria (SRB) comprise one of the few prokaryotic groups in which biological nitrogen fixation (BNF) is common. Recent studies have highlighted SRB roles in N cycling, particularly in oligotrophic coastal and benthic environments where they could contribute significantly to N input. Most studies of SRB have focused on sulfur cycling and SRB growth models have primarily aimed at understanding the effects of electron sources, with N usually provided as fixed-N (nitrate, ammonium). Mechanistic links between SRB nitrogen-fixing metabolism and growth are not well understood, particularly in environments where fixed-N fluctuates. Here, we investigate diazotrophic growth of the model sulfate reducer Desulfovibrio vulgaris var. Hildenborough under anaerobic heterotrophic conditions and contrasting N availabilities using a simple cellular model with dual ammoniotrophic and diazotrophic modes. The model was calibrated using batch culture experiments with varying initial ammonium concentrations (0–3000 µM) and acetylene reduction assays of BNF activity. The model confirmed the preferential usage of ammonium over BNF for growth and successfully reproduces experimental data, with notably clear bi-phasic growth curves showing an initial ammoniotrophic phase followed by onset of BNF. Our model enables quantification of the energetic cost of each N acquisition strategy and indicates the existence of a BNF-specific limiting phenomenon, not directly linked to micronutrient (Mo, Fe, Ni) concentration, by-products (hydrogen, hydrogen sulfide), or fundamental model metabolic parameters (death rate, electron acceptor stoichiometry). By providing quantitative predictions of environment and metabolism, this study contributes to a better understanding of anaerobic heterotrophic diazotrophs in environments with fluctuating N conditions

Effects of Tungsten and Titanium Oxide Nanoparticles on the Diazotrophic Growth and Metals Acquisition by <i>Azotobacter vinelandii</i> under Molybdenum Limiting Condition

The acquisition of essential metals, such as the metal cofactors (molybdenum (Mo) and iron (Fe)) of the nitrogenase, the enzyme responsible for the reduction of dinitrogen (N₂) to ammonium, is critical to N₂ fixing bacteria in soil. The release of metal nanoparticles (MNPs) to the environment could be detrimental to N₂ fixing bacteria by introducing a new source of toxic metals and by interfering with the acquisition of essential metals such as Mo. Since Mo has been reported to limit nonsymbiotic N₂ fixation in many ecosystems from tropical to cold temperate, this question is particularly acute in the context of Mo limitation. Using a combination of microbiology and analytical chemistry techniques, we have evaluated the effect of titanium (Ti) and tungsten (W) oxide nanoparticles on the diazotrophic growth and metals acquisition in pure culture of the ubiquitous N₂ fixing bacterium <i>Azotobacter vinelandii</i> under Mo replete and Mo limiting conditions. We report that under our conditions (≤10 mg·L^–1) TiO₂ NPs have no effects on the diazotrophic growth of <i>A. vinelandii</i> while WO₃ NPs are highly detrimental to the growth especially under Mo limiting conditions. Our results show that the toxicity of WO₃ NPs to <i>A. vinelandii</i> is due to an interference with the catechol-metalophores assisted uptake of Mo