A virulent strain of Legionella pneumophila serogroup 1 was established in continuous culture under defined iron-replete conditions at pH 6.9. Iron-limitation and extremes of pH (6.0 and 7.8) influenced the growth and metabolism of L. pneumophila, as manifested by increased metabolic activity, impaired energy coupling, and altered metabolic fluxes. In particular, the physiological versatility of L. pneumophila was demonstrated by a significant decrease in the iron content of biomass (6-fold increase in Yiron), coupled with reduced metabolic efficiency (Y, on), in response to iron-limited growth. Iron limitation promoted the accumulation of significant intracellular reserves of poly- ß-hydroxybutyrate (16 % cell dry wt.), which supported long-term survival of L. pneumophila under starvation conditions. Expression of the important pathogenicity factor, the zinc metalloprotease, was regulated by iron availability. Common iron acquisition mechanisms, such as siderophores and transferrin receptors, were not elaborated by iron-limited cells. However, human transferrin was identified as a potential iron source for L. pneumophila, with the zinc metalloprotease mediating transferrin digestion and possibly iron acquisition. Iron-limitation and extremes of pH also influenced cellular morphology and the surface properties of L. pneumophila, promoting the formation of uniform cultures of short rod-shaped bacteria, with altered fatty acid, phospholipid and protein composition. In addition to morphological and physiological adaptation, iron limitation had a significant effect on the virulence of L. pneumophila. Iron-replete cells grown at pH 6.9 and 6.0 were highly virulent in a guinea pig model. However, the virulence of L. pneumophila was significantly attenuated (P < 0.05) in response to iron-limited growth. This phenomenon was reversible, and correlated with reduced phagocytosis and / or reduced intracellular survival following infection. Decreasing the pH of iron-limited cultures to 6.0 did not stimulate recovery of culture virulence. Therefore, this study clearly demonstrates that environmental stresses, including iron limitation and extremes of pH, play an important role in modulating the physiology and virulence of L. pneumophila, inducing the expression of distinct phenotypes differing in their ability to persist in nature and cause infection.

The biological process of the carbon cycle in the Antarctic Ocean is controlled by the photosynthetic activity of the primary producers. The amount of fixed carbon does not only depend on the photosynthetic activity but also on the carbon losses due to respiration. Thus, the ratio photosynthesis to respiration (rP/R) is an important parameter to predict the effect of climate change on the Antarctic ecosystem. Indeed, the ongoing changes in climate change are influencing the dynamics of environmental conditions, which has tremendous effects on the phytoplankton community. Therefore, two ecologically relevant species from the Southern Ocean were here investigated: the diatom Chaetoceros sp. and the prymnesiophyte Phaeocystis antarctica, studying the changes in the rP/R under global climate change conditions. Three main parameters were examined i.e temperature, salinity and iron limitation. The P/R ratio was significantly affected by temperature, while salinity had only a secondary importance, although with species-specific differences. More specifically, the values were ranging from 12.3 to 7.5 for Chaetoceros sp. and from 12.4 to 2.5 for P. antarctica. The changes in this ratio were principally due to variations in respiration, rather than in photosynthesis. Chaetoceros sp. appears to be less flexible in the regulation of the extent of photoprotective mechanisms (non-photochemical quenching and alternative electrons), but its photoprotective level was generally higher than in P. antarctica. Regarding iron limitation, data were successfully collected only for Chaetoceros sp.. The P/R ratio, equal to 2.8, did not change under iron limitation, with iron limited cells showing a very efficient acclimation to the lowered assimilatory metabolism by decreasing their respiratory losses.

Phytoplankton growth is iron limited in at least 20% of the world’s oceans. Iron is an important nutrient required to synthesise enzymes necessary for photosynthesis, respiration, and nitrogen assimilation. Due to its low solubility in seawater, iron limitation of phytoplankton production has been the focus of much recent research. These organisms secrete ligands in order to solubilise the available iron, but not all of the iron dissolved in seawater is biologically available. In this study a molecular based approach was employed to investigate the acclimation of the marine haptophyte Coccolithus pelagicus to iron limitation. Using two dimensional electrophoresis, subtractive cDNA hybridisation, and RT real time PCR, changes in the proteome and in gene expression were examined. Iron limited cells were characterised by slower specific growth rates, lower chlorophyll a concentrations per unit biomass and less extensive calcification relative to iron replete cells. Addition of iron to iron limited cultures resulted in increased specific growth rates and increased chlorophyll a concentration per unit biomass. A subtracted cDNA library revealed seventeen identifiable sequences of which photosystem I protein E (PsaE), a fucoxanthin binding protein transcript, two chlorophyll binding proteins and a predicted membrane protein were shown to be up-regulated in iron-limited cells to varying extents. Two dimensional SDS PAGE revealed 11 differentially expressed proteins in iron limited cells and 1 highly expressed protein exclusive to iron replete cells. The potential utility of each of these as biomarkers of iron-limitation/iron sufficiency for natural populations of coccolithophorids like Coccolithus pelagicus is discussed.

La fixation de N2, ou diazotrophie, est un processus biogéochimique majeur en raison de son apport en azote nouveau dans la couche de surface de l’océan. Cependant ses facteurs de contrôle restent mal connus. Le fer, dont les concentrations de surface sont faibles, est un élément potentiellement limitant de la fixation de N2 du fait du contenu en fer important de la nitrogénase. En raison de leur découverte récente, peu d’études ont été menées sur les cyanobactéries diazotrophes unicellulaires (UCYN) pouvant être responsables de ~50% de la fixation de N2 à l’échelle globale. Des expériences en culture ont permis de caractériser et quantifier pour la première fois la réponse d’une UCYN, Crocosphaera watsonii, face à la limitation en fer. En condition de limitation en fer, il a été observé une réduction de la croissance et des taux de fixation de N2 ainsi qu’une stratégie d’adaptation des cellules avec une diminution de leur volume. La stimulation de la croissance et de l’activité de C. watsonii cultivées en condition de limitation en fer suite à l’ajout d’une pluie saharienne artificielle a permis de mettre en évidence qu’une partie au moins du fer issu de poussières désertiques est biodisponible. En Atlantique subtropical Nord, où nous avons déterminé une forte contribution de la fixation de N2 à la production nouvelle, nous avons observé une limitation de la fixation de N2 et de la production primaire principalement par les phosphates et mis en évidence le rôle des métaux traces dans le contrôle de la fixation de N2. Un ajout de pluie saharienne a permis de stimuler systématiquement la fixation de N2 et la production primaire. / Despite the biogeochemical importance of N2 fixation, which represents the largest source of newly-Fixed nitrogen to the open ocean, some uncertainties remain about its controlling factors. Iron (Fe) is widely suspected as a key controlling factor due to the high Fe content of the nitrogenase complex and to its low concentration in oceanic surface seawaters. N2 fixation rates associated with unicellular N2 fixing cyanobacteria (UCYN) were estimated to be ~50% of the total N2 fixation at global scale, but as they have been recently discovered few studies have been conducted on these organisms. We performed culture experiments in order to quantify for the first time the response of an UCYN, Crocosphaera watsonii, to Fe limitation. Reduction of ambient Fe concentration led to significant decreases in growth rate and N2 fixation rates per cell and we observed an adaptive strategy to Fe limitation with a cell volume reduction. Then, the enhancement of growth and activity of C. watsonii under Fe limitation condition after artificial Saharan rain addition highlighted that at least a part of the Fe released by the dust is bioavailable. In subtropical North Atlantic, an important contribution of N2 fixation to new production was observed and we showed that primary production and N2 fixation were globally P-Limited. We revealed that trace metals play a key role in controlling N2 fixation in this area. Saharan rain addition stimulated N2 fixation, presumably by supplying these nutrients. All these results contribute to our knowledge of the control of oceanic N2 fixation and provide new insight about interactions between Fe, nitrogen and carbon biogeochemical cycles.