Lakes are important global ecosystems and many of them are nutrient-poor (unproductive). Especially in northern boreal latitudes, lakes may be heavily subsidized by terrestrial organic material (t-OM) from peat layers in the catchment. Thus, in addition to heterotrophic bacteria and phytoplankton, zooplankton may also use the particulate fraction of peat layer t-OM (t-POM) as a potential food source in those systems. Inputs of t-OM in northern latitudes are anticipated to increase in the future due to increasing precipitation and temperature. As t-OM is a good substrate for bacterial growth and as bacteria can often outcompete phytoplankton for inorganic nutrients, the proportions of heterotrophic bacteria and phytoplankton are expected to change in unproductive lakes. This may have pronounced impacts on zooplankton population dynamics. The aim of my thesis was to investigate how changes in food quality and quantity will affect metazoan zooplankton performance in unproductive lakes. Three laboratory studies assessed the quality of specific food components (phytoplankton, bacteria and peat layer t-POM) and their effects on Daphnia survival, growth and reproduction. Further, a mesocosm study with a full natural plankton community tested the predictions of the Light:Nutrient-Hypothesis in an unproductive clear water lake in situ by adding carbon and inorganic nutrients and changing light availability. I found that pure bacterial (Pseudomonas sp.) or t-POM diets could not sustain Daphnia populations, even though both were readily ingested. Daphnids needed at least 10-20% phytoplankton (Rhodomonas) in the diet to survive and even higher proportions (≥ 50%) were necessary for the production of viable offspring. Further, I showed that the dilution of non-limiting concentrations of Rhodomonas with increasing proportions of Pseudomonas or t-POM led to decreased Daphnia performance. Both Pseudomonas and t-POM lack essential biochemicals (fatty acids and sterols). In contrast, mineral nutrient limitation only occurred on t-POM-dominated diets as evidenced by a labeling experiment that showed Daphnia can incorporate carbon and phosphorus from Rhodomonas and Pseudomonas with similar efficiencies. Thus, peat layer t-POM was a lower quality food than Pseudomonas. This was corroborated by the finding that intermediate additions of Pseudomonas to limiting amounts of Rhodomonas supported increased Daphnia survival, growth and reproduction while t-POM additions had no beneficial effect. My results suggest that high terrestrial stable isotope signals in metazoan zooplankton are most likely derived from t-OM that is channeled tohigher trophic levels via the microbial loop (i.e. heterotrophic bacteria and/or bacterivorous protozoa) but not from direct metazoan feeding on t-POM. Furthermore, bacteria may serve as an important supplement to zooplankton diets when phytoplankton abundance is low. However, a sufficient proportion of high quality phytoplankton is always necessary to fulfil mineral and especially biochemical requirements of zooplankton in unproductive aquatic systems. The results of the mesocosm study showed that the Light:Nutrient-Hypothesis is not applicable to unproductive clear water systems in which the phytoplankton community is dominated by mixotrophs. In the face of the theoretical predictions, low light levels led to decreased zooplankton biomass. This was most likely caused by a shift in the algal community composition towards less edible taxa. Another reason may have been a weakening of the microbial loop. This is in line with the results of the laboratory studies that point out the importance of the microbial food web for zooplankton nutrition in unproductive lakes.

Little is known about the role of microbial biofilms in nutrient cycling and ecosystem processes within desert springs. Furthermore, the difference between nutrient limitation of biofilms in desert springs and other ecosystems is unknown. Biofilms produce micro-scale physicochemical variation important to ecosystem function. We measured the variation in microscale physicochemical heterogeneity in biofilms of Devils Hole, Nevada. Microelectrodes were used to measure micro-scale chemical gradients of temperature, pH, O2, and H2S in addition to water column and pore water nutrient measurements in Spirogyra, cyanobacteria, and Beggiatoa biofilms over one year. Biofilm physicochemical gradients were used to calculate diffusion and metabolic rates. The rate of O2 and H2S diffusion ranged over two orders of magnitude. Biofilm production and respiration were influenced by biofilm type, light exposure, and sample month. Maximum O2 production occurred in spring and summer during direct light exposure. Oxygen production and consumption varied with light exposure and season. The H2S production and consumption varied with biofilm type. Higher concentrations of SO4 in Beggiatoa suggested that H2S production in Beggiatoa was quickly oxidized in the ecosystem. Spirogyra and cyanobacteria followed similar physicochemical trends; however, Spirogyra had more pronounced diurnal and seasonal variation. The differences between cyanobacteria and Spirogyra have implications on the ecosystem function of Devils Hole as well as other ecosystems with diverse biofilm communities. The heterogeneous physicochemistry of microbial biofilms and the differences in biofilm nutrient limitation suggests that a change in microbial biofilms or nutrient concentrations could alter ecosystem biogeochemical dynamics. Additionally, we assessed the nutrient limitation of two desert springs in comparison with a temperate stream. A nutrient diffusing substrata experiment was used to measure chlorophyll a, respiration, and biomass with phosphorus, nitrogen, and sulfide treatments. Autotrophic and heterotrophic biofilms responded differently to treatments and the temperate stream had higher chlorophyll a biofilm accrual but lower respiration relative to the desert springs. / Spatial and temporal variation of microbial biofilm biogeochemistry in a desert spring, Devils Hole, Nevada -- Autotrophic and heterotrophic nutrient limitation across biomes. / Department of Biology

Conselho Nacional de Desenvolvimento CientÃfico e TecnolÃgico / O pescado à um alimento susceptÃvel à deterioraÃÃo devido Ãs suas caracterÃsticas intrÃnsecas, sendo necessÃrio o monitoramento bacteriano no armazenamento pÃs-morte. Dessa forma, objetivou-se quantificar bactÃrias, psicrotrÃficas, psicrÃfilas e mesÃfilas em tilÃpias do Nilo (Oreochromis niloticus) durante 21 dias de estocagem sob aÃÃo do gelo. Foram realizados quatro experimentos, separados em dois grupos, nÃo eviscerados e eviscerados, cada um com oito amostras perfazendo um total de 32 amostras. A cada trÃs dias era retirado um exemplar de tilÃpia para as anÃlises do experimento: sensorialmente, conforme a avaliaÃÃo do MÃtodo do Ãndice de Qualidade (MIQ), adaptado à tilÃpia do Nilo e microbiologicamente. Na anÃlise sensorial do peixe cru foram avaliados os atributos: aspecto geral, olhos, brÃnquias, textura e musculatura dos peixes, segundo os somatÃrios dos caracteres sensoriais (S.C.S). De cada exemplar foram quantificados os grupos bacterianos: psicrotrÃficos (7ÂC), psicrÃfilos (23ÂC), e mesÃfilos (35ÂC), na superfÃcie e no mÃsculo, pelo mÃtodo de Contagem PadrÃo em Placas (CPP). NÃo houve diferenÃa estatÃstica entre os trÃs grupos bacterianos analisados à um nÃvel de significÃncia de 5% dentro dos mesmos tratamentos e entre os grupos. O tempo de estocagem apresentou alta correlaÃÃo com a contagem dos grupos bacterianos e o aumento do MIQ, tanto para os peixes eviscerados, quanto para os nÃo eviscerados. Foi considerado que o processo de evisceraÃÃo no local de venda dos peixes nÃo aumentou sua vida Ãtil. Sugere-se, portanto, uma maior atenÃÃo no processo de evisceraÃÃo em peixarias que comercializam peixes vivos, pois esse processo nÃo foi realizado de forma eficaz para seguranÃa e valorizaÃÃo do produto. / Fish is a food susceptible to deterioration due their intrinsic characteristics, requiring bacterial monitoring in postmortem storage. Thus, the objective was to quantify mesophilic, psychrotrophic, psychrophilic bacteria in Nile tilapia (Oreochromis niloticus) during to 21 days of storage under the action of ice at temperature refrigeration. Four experiments were conducted, separated to two groups, gutted and ungutted, with eight samples totaling 32 samples. Every three days was taken a exemplar of tilapia for analyzes of the trial: sensory evaluation according to the Quality Index Method (QIM) adapted to the Nile tilapia and heterotrophic bacterial count of cultivable. In sensory analysis of raw fish attributes were assessed: general appearance, eyes, gills, muscles and texture of the fish, according to the Sensory Characters Sum (S.C.S). To each specimen were quantified bacterial groups: psychrophilic (23 ÂC), psychrotrophic (7 ÂC) and mesophilic (35 ÂC), the surface and the muscle, using the method of standard plate count (SPC). There was no statistical difference between the three bacterial groups analyzed the significance level of 5% within the same treatments and between groups. The storage time was highly correlated with the counting of bacterial groups and increased QIM for both eviscerated fish, and for those not drawn. It was considered that the process of evisceration on the premises of the fish did not increase its useful life. It is suggested, therefore, greater attention in the process of gutting fish markets that sell live fish, because this process was not carried out effectively to security and recovery product.

En milieu marin, les communautés bactériennes hétérotrophes sont exposées à un mélange hétérogène de composés organiques dissous présentant différents degrés de labilité, qui peuvent contrôler à la fois leurs activités et leur composition. Dans le cadre de cette thèse, nous avons étudié la réponse de communautés bactériennes présentes dans des environnements côtiers contrastés à l’ajout simple ou combiné de différentes sources naturelles de matière organique dissoute (MOD), préalablement photo oxydée ou non et présentant différentes bio réactivités. Dans un premier temps, nous avons étudié l’impact d’ajout de MOD labile provenant de méduses sur l’activité et la diversité bactériennes d’une communauté d’une lagune côtière méditerranéenne. Nous avons observé que la résilience des communautés bactériennes suite à cet ajout était plus importante pour les fonctions métaboliques que pour la diversité. Ceci suggère que les efflorescences de méduses peuvent entrainer des changements durables de la structure des communautés bactériennes en environnement côtier. Dans un deuxième temps, nous nous sommes intéressés à la possibilité d’un priming effect (accélération de la dégradation bactérienne de la MOD réfractaire en présence de MOD labile) en milieu côtier. Des expériences d’ajout simple ou combiné de MOD labile et récalcitrante ont été réalisées sur des communautés bactériennes de Mer Méditerranée et d’un fjord de Patagonie chilienne. Dans les deux cas, nous avons observé des changements plus importants de la composition communautaire bactérienne suite à un ajout combiné. Cependant, nous n’avons pas observé une plus forte consommation de MOD récalcitrante suite à l’ajout de composés labiles, ce qui suggère que le priming effect n’a pas eu lieu au cours de nos expériences. Enfin, nous avons étudié l’impact de la photodégradation de différentes sources de MOD (i.e. MOD récalcitrante de rivière, MOD labile provenant de phytoplancton) sur l’activité et la diversité de communautés bactériennes côtières. Nous avons observé que la photodégradation de la MOD issue d’une culture de phytoplancton entrainait à la fois une croissance bactérienne plus faible et une modification de la diversité bactérienne en faveur des Alphaprotéobactéries. / Heterotrophic bacterial communities in marine environment are exposed to a heterogeneous mixture of dissolved organic compounds with different bioavailability that may control both their activities and their composition. In the frame of this thesis, we studied the response of different coastal bacterial communities to the single or combined addition of various natural dissolved organic matter (DOM) photo-oxidized or not, presenting different bioavailability characteristics. Firstly, we studied the effects of the addition of highly bioreactive DOM derived from jellyfish on bacterial activities and community structure in a Mediterranean coastal lagoon. We observed that resilience of the bacterial community after the addition of DOM from the jellyfish was higher for metabolic functions than diversity, suggesting that jellyfish blooms can induce durable changes in the bacterial community structure in coastal lagoons. Secondly, we investigated the occurrence of priming effect (increase in microbial degradation of refractory DOM upon the addition of labile DOM) on coastal marine environments. Experiments with single or combined additions of recalcitrant and labile DOM sources were performed with a Mediterranean and a Patagonian fjord bacterial communities. In both cases we observed a greater effect of combined addition on bacterial community composition. However we did not observe an increase in recalcitrant DOM degradation of recalcitrant DOM following the addition of labile compounds, suggesting that priming effect did not occur during our experiments. Finally, we studied the impact of contrasted DOM (i.e. recalcitrant riverine DOM, labile phytoplankton-derived DOM) photodegradation on coastal bacterial communities activity and composition. We observed that photodegradation of phytoplankton-derived DOM led to a lower bacterial growth and changes in bacterial community diversity, in favour of Alphaproteobacteria.

This thesis investigates the effects of co-culturing microorganisms including 37 yeast, 38 bacteria, nine diazotrophic cyanobacteria, and three fungi on biomass and lipid production in fresh- and saltwater algae. Algal lipid content was measured using Nile Red method and gravimetric techniques. Among the algal strains tested, freshwater Coelastrum sp. 46-4, and saltwater Cricosphaera sp. 146-2-9, showed enhanced biomass yield and lipid content in response to co-culture with bacteria, cyanobacteria, and fungi. While co-culture with yeast caused inhibition of algal productivity, no difference in algal productivity was observed between nitrogen-free diazotrophic cyanobacterial co-culture and nitrogen-replete monoalgal culture. Results indicated that extracellular compounds from the freshwater bacteria Pseudomonas stutzeri and marine fungus Fusarium sp. significantly account for stimulation of lipid accumulation within algal cells, while co-cultivation with live microorganism cells stimulated biomass production in algae.

Oceans are powered by active, metabolically diverse microorganisms, which are important in regulating biogeochemical cycles on Earth. Most of the ocean surface is often limited by nutrients, influencing bacterial growth and activities. Bacterial adaptation to fluctuating environmental conditions involves extensive reprogramming, and redirection of bacterial metabolism and physiology. In this thesis, I investigated the molecular mechanisms of bacterial adaptation strategies to sustain their growth and survival, focusing on the regulation of gene and protein expression in heterotrophic marine bacteria. Comparative proteomics analyses of the growth and non-growth conditions, uncovered central adaptations that marine bacteria employ to allow them to change their metabolism to support exponential growth in response to nutrients and to readjust to stationary phase under nutrient limitation. Our results highlight that during nutrient rich conditions three distinct bacteria lineages have great similarities in their proteome. On the other hand, we observed pronounced differences in behavior between taxa during stationary phase. Analyses of the proteorhodopsin containing bacterium Vibrio sp. AND4 during starvation showed that significantly improved survival in the light compared to darkness. Notably, proteins involved in promoting cell vitality and survival had higher relative abundance under light. In contrast, cells in the dark need to degrade their endogenous resources to support their basic cellular demands under starvation. Thus, light strongly influences how PR-containing bacteria organize their molecular composition in response to starvation. Study of alternative energy generation metabolisms in the Alphaproteobacteria Phaeobacter sp. MED193 showed that the addition of thiosulfate enhanced the bacterial growth yields. Concomitantly, inorganic sulfur oxidation gene expression increased with thiosulfate compared to controls. Moreover, thiosulfate stimulated protein synthesis and anaplerotic CO2 fixation. These findings imply that this bacterium could use their lithotrophic potential to gain additional energy from sulfur oxidation for both improving their growth and survival. This thesis concludes that analyses in model organisms under defined growth conditions gives invaluable knowledge about the regulatory networks and physiological strategies that ensure the growth and survival of heterotrophic bacteria. This is critically important for interpreting bacterial responses to dynamic environmental changes. Moreover, these analyses are crucial for understanding genetic and proteomic responses in microbial communities or uncultivated organisms in terms of defining ecological niches of planktonic bacteria

Flow cytometry (FCM) is a well established technique used for enumeration and characterization of marine biological particles, which fulfills the scientific demands of rapid cell counting automation. FCM allows the discrimination of pico- and nanoplankton populations regarding its abundance, cell size, and pigment content both by natural or induced fluorescence. The cyanobacterium Prochlorococcus is widespread in the euphotic zone of the tropical and subtropical oceans, and is considered the smallest and most abundant photosynthetic organism in the planet. Synechococcus, other important cyanobacterium genus present in the picoplankton, is highly diverse and is widely distributed in marine ecosystems from cold and mesotrophic, to warm, open ocean oligotrophic waters. Photosynthetic pico- and nanoeukaryotes display a range of physiologies and life strategies. Although its abundance is generally lower, the larger cell size leads to a significant contribution to the epipelagic community biomass. In this thesis I aimed to investigate the abundance and carbon biomass distribution of heterotrophic bacteria, Prochlorococcus, Synechococcus, autotrophic pico- and nanoeukaryotes in the Southwest Atlantic Ocean off Brazil, their relation to the different water masses and the influence of hydrodynamic (South Atlantic Central Water intrusion) and biological processes (Trichodesmium spp. and Mesodinium rubrum blooms) on such distributions. / A citometria de fluxo (FCM, sigla em inglês) é uma técnica bem estabelecida, usada para enumeração e caracterização de partículas biológicas marinhas, a qual supre a demanda científica por automação rápida de contagem de células. A FCM permite a discriminação de populações pico- e nanoplanctônicas no que concerne à sua abundância, tamanho de célula e pigmentação, tanto por fluorescência natural ou induzida. A cianobactéria Prochlorococcus é amplamente disseminada pela zona eufótica dos oceanos tropicais e subtropicais, sendo considerada o menor e mais abundante organismo fotossintético do planeta. Synechococcus, outro importante gênero de cianobactérias presente no picoplâncton, é diverso e amplamente distribuído em ecossistemas marinhos, de águas frias e mesotróficas à regiões oceânicas quentes e de águas oligotróficas. Pico- e nanoeucariotos fotossintéticos apresentam uma grande variedade de fisiologias e estratégias de vida. Embora a abundância destes grupos seja geralmente menor, o maior tamanho de suas células resulta numa contribuição significante dos mesmos para a biomassa da comunidade epipelágica. Na presente tese eu objetivei investigar a abundância e a distribuição da biomassa de carbono de bactérias heterotróficas, Prochlorococcus, Synechococcus, pico- e nanoeucariotos autotróficos no sudoeste do Oceano Atlântico ao largo do Brasil, sua relação com as diferentes massas d\'água e a influência de processos hidrodinâmicos (como a intrusão da Água Central do Atlântico Sul) e biológicos (florações de Trichodesmium spp. e Mesodinium rubrum) em tais distribuições.

Flow cytometry (FCM) is a well established technique used for enumeration and characterization of marine biological particles, which fulfills the scientific demands of rapid cell counting automation. FCM allows the discrimination of pico- and nanoplankton populations regarding its abundance, cell size, and pigment content both by natural or induced fluorescence. The cyanobacterium Prochlorococcus is widespread in the euphotic zone of the tropical and subtropical oceans, and is considered the smallest and most abundant photosynthetic organism in the planet. Synechococcus, other important cyanobacterium genus present in the picoplankton, is highly diverse and is widely distributed in marine ecosystems from cold and mesotrophic, to warm, open ocean oligotrophic waters. Photosynthetic pico- and nanoeukaryotes display a range of physiologies and life strategies. Although its abundance is generally lower, the larger cell size leads to a significant contribution to the epipelagic community biomass. In this thesis I aimed to investigate the abundance and carbon biomass distribution of heterotrophic bacteria, Prochlorococcus, Synechococcus, autotrophic pico- and nanoeukaryotes in the Southwest Atlantic Ocean off Brazil, their relation to the different water masses and the influence of hydrodynamic (South Atlantic Central Water intrusion) and biological processes (Trichodesmium spp. and Mesodinium rubrum blooms) on such distributions. / A citometria de fluxo (FCM, sigla em inglês) é uma técnica bem estabelecida, usada para enumeração e caracterização de partículas biológicas marinhas, a qual supre a demanda científica por automação rápida de contagem de células. A FCM permite a discriminação de populações pico- e nanoplanctônicas no que concerne à sua abundância, tamanho de célula e pigmentação, tanto por fluorescência natural ou induzida. A cianobactéria Prochlorococcus é amplamente disseminada pela zona eufótica dos oceanos tropicais e subtropicais, sendo considerada o menor e mais abundante organismo fotossintético do planeta. Synechococcus, outro importante gênero de cianobactérias presente no picoplâncton, é diverso e amplamente distribuído em ecossistemas marinhos, de águas frias e mesotróficas à regiões oceânicas quentes e de águas oligotróficas. Pico- e nanoeucariotos fotossintéticos apresentam uma grande variedade de fisiologias e estratégias de vida. Embora a abundância destes grupos seja geralmente menor, o maior tamanho de suas células resulta numa contribuição significante dos mesmos para a biomassa da comunidade epipelágica. Na presente tese eu objetivei investigar a abundância e a distribuição da biomassa de carbono de bactérias heterotróficas, Prochlorococcus, Synechococcus, pico- e nanoeucariotos autotróficos no sudoeste do Oceano Atlântico ao largo do Brasil, sua relação com as diferentes massas d\'água e a influência de processos hidrodinâmicos (como a intrusão da Água Central do Atlântico Sul) e biológicos (florações de Trichodesmium spp. e Mesodinium rubrum) em tais distribuições.

<p>Heterotrophic bacteria utilize dissolved organic matter, and the carbon flow through an ecosystem depends on the fractions of the utilized carbon that is either respired or transferred to higher trophic levels. The major aim this thesis is to investigate 1) the relationship between composition and functioning in heterotrophic bacterioplankton communities and 2) the influence of environmental conditions on both parameters. I set up several batch culture experiments, where lake water filtrates containing bacteria but no grazers were inoculated into sterile freshwater medium to investigate the importance of the origin of the source community (the inoculum) versus the environmental conditions (the medium) for the composition and functional performance of bacterial communities. In some experiments the medium was manipulated to simulate changes in salinity, pH and dissolved organic matter quantity and quality. Functional parameters (biomass yield, respiration, growth efficiency and enzyme activities) and the genetic composition of the emerging bacterial communities were determined.</p><p>When bacterial inocula obtained from different habitats were re-grown under identical conditions, differently composed communities emerged. This indicates that the history and distribution of taxa within the inoculum was an important regulating factor of community composition. The coupling between community composition and functioning was not very tight, and there was functional equivalency with respect to aggregated functions important at the ecosystem scale (e.g., biomass production and respiration). The functional performance of bacterial communities could to a large extent be predicted from the medium alone, except when it deviated strongly from the ambient settings. When bacterial communities were exposed to dilution, a strong change in pH or an increase in salinity, growth of structurally and functionally distinct communities occurred. I therefore suggest that it depends on the disturbance regime how bacterial community structure and function are related to each other. </p>

Heterotrophic bacteria utilize dissolved organic matter, and the carbon flow through an ecosystem depends on the fractions of the utilized carbon that is either respired or transferred to higher trophic levels. The major aim this thesis is to investigate 1) the relationship between composition and functioning in heterotrophic bacterioplankton communities and 2) the influence of environmental conditions on both parameters. I set up several batch culture experiments, where lake water filtrates containing bacteria but no grazers were inoculated into sterile freshwater medium to investigate the importance of the origin of the source community (the inoculum) versus the environmental conditions (the medium) for the composition and functional performance of bacterial communities. In some experiments the medium was manipulated to simulate changes in salinity, pH and dissolved organic matter quantity and quality. Functional parameters (biomass yield, respiration, growth efficiency and enzyme activities) and the genetic composition of the emerging bacterial communities were determined. When bacterial inocula obtained from different habitats were re-grown under identical conditions, differently composed communities emerged. This indicates that the history and distribution of taxa within the inoculum was an important regulating factor of community composition. The coupling between community composition and functioning was not very tight, and there was functional equivalency with respect to aggregated functions important at the ecosystem scale (e.g., biomass production and respiration). The functional performance of bacterial communities could to a large extent be predicted from the medium alone, except when it deviated strongly from the ambient settings. When bacterial communities were exposed to dilution, a strong change in pH or an increase in salinity, growth of structurally and functionally distinct communities occurred. I therefore suggest that it depends on the disturbance regime how bacterial community structure and function are related to each other.