Pseudomonas aeruginosa displays an epidemic population structure.

peer reviewedBacteria can have population structures ranging from the fully sexual to the highly clonal. Despite numerous studies, the population structure of Pseudomonas aeruginosa is still somewhat contentious. We used a polyphasic approach in order to shed new light on this issue. A data set consisting of three outer membrane (lipo)protein gene sequences (oprI, oprL and oprD), a DNA-based fingerprint (amplified fragment length polymorphism), serotype and pyoverdine type of 73 P. aeruginosa clinical and environmental isolates, collected across the world, was analysed using biological data analysis software. We observed a clear mosaicism in the results, non-congruence between results of different typing methods and a microscale mosaic structure in the oprD gene. Hence, in this network, we also observed some clonal complexes characterized by an almost identical data set. The most recent clones exhibited serotypes O1, 6, 11 and 12. No obvious correlation was observed between these dominant clones and habitat or, with the exception of some recent clones, geographical origin. Our results are consistent with, and even clarify, some seemingly contradictory results in earlier epidemiological studies. Therefore, we suggest an epidemic population structure for P. aeruginosa, comparable with that of Neisseria meningitidis, a superficially clonal structure with frequent recombinations, in which occasionally highly successful epidemic clones arise

The Neglected Intrinsic Resistome of Bacterial Pathogens

Bacteria with intrinsic resistance to antibiotics are a worrisome health problem. It is widely believed that intrinsic antibiotic resistance of bacterial pathogens is mainly the consequence of cellular impermeability and activity of efflux pumps. However, the analysis of transposon-tagged Pseudomonas aeruginosa mutants presented in this article shows that this phenotype emerges from the action of numerous proteins from all functional categories. Mutations in some genes make P. aeruginosa more susceptible to antibiotics and thereby represent new targets. Mutations in other genes make P. aeruginosa more resistant and therefore define novel mechanisms for mutation-driven acquisition of antibiotic resistance, opening a new research field based in the prediction of resistance before it emerges in clinical environments. Antibiotics are not just weapons against bacterial competitors, but also natural signalling molecules. Our results demonstrate that antibiotic resistance genes are not merely protective shields and offer a more comprehensive view of the role of antibiotic resistance genes in the clinic and in nature

Hydrogen photo-evolution upon S deprivation stepwise: An illustration of microalgal photosynthetic and metabolic flexibility and a step stone for future biotechnological methods of renewable H2 production

The metabolic flexibility of some photosynthetic microalgae enables them to survive periods of anaerobiosis in the light by developing a particular photofermentative metabolism. The latter entails compounds of the photosynthetic electron transfer chain and an oxygen-sensitive hydrogenase in order to reoxidise reducing equivalents and to generate ATP for maintaining basal metabolic function. This pathway results in the photo-evolution of hydrogen gas by the algae. A decade ago Melis and coworkers managed to reproduce such a condition in a laboratory context by depletion of sulfur in the algal culture media, making the photo-evolution by the algae sustainable for several days (Melis et al. 2000). This observation boosted research in algal H2 evolution. A feature, which due to its transient nature was long time considered as a curiosity of algal photosynthesis suddenly became a phenomenon with biotechnological potential. Although the Melis procedure has not been developed into a biotechnological process of renewable H2 generation so far, it has been a useful tool for studying microalgal metabolic and photosynthetic flexibility and a possible step stone for future H2 production procedures. Ten years later most of the critical steps and limitations of H2 production by this protocol have been studied from different angles particularly with the model organism C. reinhardtii, by introducing various changes in culture conditions and making use of mutants issued from different screens or by reverse genomic approaches. A synthesis of these observations with the most important conclusions driven from recent studies will be presented in this review.Micro-H2 (Microbiological production of hydrogen) ARC07/12-0

Hydrogen photoproduction by oxygenic photosynthetic microorganisms

peer reviewe

La réponse photosynthétique d'une algue verte à la carence en soufre

Chlamydomonas reinhardtii possède la capacité de produire de l'hydrogène à la lumière en absence d'oxygène. Cette condition peut être obtenue en cultivant les algues dans un milieu carencé en soufre. La carence en soufre entraîne une forte diminution de l'activité du photosystème II tout en maintenant une respiration élevée, ce qui provoque un passage de cultures fermées en anoxie et induit la production d'hydrogène. Dans cette étude, nous avons caractérisé la réponse photosynthétique à la carence en soufre chez la souche sauvage et la souche déficiente en oxydase alternative mictochondriale (AOX) dans des milieux contenant de l'ammonium ou du nitrate comme source d'azote. L'AOX, inductible par le nitrate, fait partie de la chaîne de transport d'électrons mitochondriale et catalyse l'oxydation de l'ubiquinol en transférant directement ses électrons à l'oxygène. Ainsi l'AOX entre en compétition avec le complexe III et est impliquée dans une voie de dissipation du pouvoir réducteur en excès

Study of photosynthesis of Chlamydomonas reinhardtii under High and Low CO2 conditions.

In photoautotophically air-grown microalgae, CO2 availability is usually limited. The unicellular green algae Chlamydomonas reinhardtii can adapt to low CO2 concentration with the inorganic carbon concentration mechanism (CCM). This has been extensively studied in the past but functional adaptation of the photosynthetic apparatus has been less studied. Photosynthetic organisms can cope with CO2 limitation by dissipating excess absorbed energy with the help of different energy dissipating mechanisms like energy-dependent non-photochemical quenching (NPQ). In Chlamydomonas reinhardtii, this process only seems to develop to high levels in extreme conditions combining high light and strong CO2 limitation. Under moderate CO2 limitation, absence of important energy-dependent NPQ suggests the development of another energy dissipating mechanism. We compared the growth and functional adaptations of the photosynthetic apparatus of the wild-type strain 1690 grown in photobioreactor under low and high CO2 bubbling, 0.039% and 10%, respectively. Under low CO2, where growth was 2 to 4 times slower than under high CO2, the non-linear relationship between electron transport rate (derived from PAM fluorescence measurements) and gross oxygen evolution rate suggested that a significant portion of the electron flux is directed to oxygen at light intensities approaching photosynthetic saturation (either at PSI or at PTOX). The use of the mutant strain PTOX2 indicated that O2 reduction occurs mainly at PSI and not at PTOX. Low temperature fluorescence emission spectra indicated no significant difference in excitation energy distribution between PSI and PSII. Western blot analysis showed no major differences in abundance of Rubisco or of photosystem subunits between the two conditions. In contrast, cytochrome f abundance was lower in high CO2 condition. Although energy-dependent NPQ remained weak, low CO2 cells were characterized by a higher xanthophyll deepoxydation index which usually indicates more dissipation as heat, as also suggested by increased Lhcsr3 expression. Despite a higher ATP requirement of the CCM mechanism in low CO2 condition, only minor difference in cyclic electron transport could be found if compared to high CO2 condition (as determined by P700 spectroscopic measurements). In Chlamydomonas, conflicting views were expressed in earlier studies on the amplitude and role of Mehler-type O2-uptake at steady state. Our analysis of oxygen evolution, electron transport and NPQ after growth under different combinations of light intensities and CO2 supply rates allows us to define Mehler-type alternative electron transport as an important and flexible response to photosynthetic electron transport saturation in Chlamydomonas

A Chlamydomonas mutant locked in anaerobiosis

The soil dwelling microalga Chlamydomonas reinhardtii most likely encounters transient periods of anaerobiosis in its natural environment, for instance at night time or when photosynthesis is turned down in response to macronutrient limitation. Anoxic conditions trigger state I to state II transition in C.r. and the induction of a chloroplast hydrogenase., which ability to accept electrons from reduced Fd results in a transient light driven H2 evolution. We present evidence that hydrogenase induction and state transitions are required for the induction of photosynthesis in anaerobiosis and therefore critical for this alga in order to survive transient anaerobic periods in the dark. In an anaerobic metabolic context the induction of photosynthesis is severely slowed down. The highly reduced state of the NAD(P) pools and the absence of O2 as electron sink hamper light driven reoxydation of the intersystem electron carriers while CO2 assimilation by the Calvin cycle is inhibited by ATP deficiency. We have seen that gradual increase of hydrogenase activity during anaerobiosis restores a PSI acceptor pool and leads to a reduction of the induction lag of oxygenic photosynthesis. A mutant HydEF devoid of hydrogenase maturation genes typically shows 3 to 4 times longer lag phases that the WT. State transitions provide another mechanism by which photosynthetic electron transport can be unlocked in anaerobic conditions. A state II conformation is known to stimulate photo-phosphorylation, and may therefore restore Calvin cycle activity in an ATP depleted metabolic context. We observed that an anaerobically adapted stt7 mutant locked in state I is only able to induce oxygenic photosynthesis upon hydrogenase expression. We therefore constructed a double mutant Stt7HydEF impaired of state transition ability and hydrogenase activity and found it to have lost the capacity of inducing photosynthesis in anaerobic conditions.SUNBIOPATH - ARC MICRO-H

Analysis of PSII antenna size heterogeneity of Chlamydomonas reinhardtii during state transitions - Colloque annuel de la Société Française de Photosynthèse

PSII antenna size heterogeneity has been extensively studied in the past. Based on in vivo DCMU fluorescence rise kinetics, at least two types of photosystems were described. They differ by their apparent antenna size and connectivity (this last term refers to the transfer of absorbed energy from a closed PSII unit to an open neighboring unit). In this study, we analysed PSII heterogeneity in Chlamydomonas reinhardtii using non-linear linear regression fitting on in vivo DCMU fluorescence rise kinetics, with a focus on changes in PSII heterogeneity associated with state transitions. We found that PSIIα possesses a high degree of connectivity and an antenna about 3 times larger than PSIIβ, as described previously. In contrast with most earlier studies, we found some connectivity for PSIIβ (although it was highly variable). This is in agreement with recent models based on biochemical and structural analysis of PSII after gel filtration separation which describe PSII mega-, super- and core- complexes in Chlamydomonas. According to these studies, the smallest unit of PSII in vivo would be a dimer of two core complexes hence still allowing connectivity. We also showed that strain and medium dependent variations in the half-time of the fluorescence rise, generally taken as an indicator of the average cross-section of PSII, can be explained by variations in the proportions of PSIIα and PSIIβ. When analyzing the state transition process, we showed for the first time in vivo that it induces an inter-conversion of PSIIα and PSIIβ. These findings are discussed with respect to the latest insights on the remodeling of the pigment-protein PSII architecture during this process

Interplay between non-photochemical plastoquinone reduction and re-oxidation in pre-illuminated Chlamydomonas reinhardtii: a chlorophyll fluorescence study

In photosynthetic eukaryotes, the redox state of the plastoquinone (PQ) pool is an important sensor for mechanisms that regulate the photosynthetic electron transport. In higher plants, a multimeric nicotinamide adenine dinucleotide (phosphate) (NAD(P))H dehydroge- nase (NDH) complex and a plastid terminal oxidase (PTOX) are involved in PQ redox homeostasis in the dark. We recently demonstrated that in the microalgae Chla- mydomonas reinhardtii, which lacks the multimeric NDH complex of higher plants, non-photochemical PQ reduction is mediated by a monomeric type-II NDH (Nda2). In this study, we further explore the nature and the importance of non-photochemical PQ reduction and oxidation in relation to redox homeostasis in this alga by recording the ‘dark’ chlorophyll fluorescence transients of pre-illuminated algal samples. From the observation that this fluorescence tran- sient is modified by addition of propyl gallate, a known inhibitor of PTOX, and in a Nda2-deficient strain we conclude that it reflects post-illumination changes in the redox state of PQ resulting from simultaneous PTOX and Nda2 activity. We show that the post-illumination fluo- rescence transient can be used to monitor changes in the relative rates of the non-photochemical PQ reduction and reoxidation in response to different physiological situa- tions. We study this fluorescence transient in algae acclimated to high light and in a mutant deficient in mitochondrial respiration. Some of our observations indi- cate that the chlororespiratory pathway participates in redox homeostasis in C. reinhardtii