The aim of this study was to design and optimize new PCR primers for detection of potential cyanobacterial producers of cytotoxic lipopeptides puwainaphycins and minutissamides in environmental samples. Samples from two distinct localities were tested, as suggested based on preliminary data. The first set of samples consisted of cyanobacterial soil biofilms from sheep pastures affected by Alveld illness in Norway. The other one contained samples of planktic cyanobacaterial blooms from Protected Landscape Area Třeboň and its vicinity. Three different approaches were used for evaluation of the presence of cyanobacterial lipopeptide producers: microscopy, PCR with the designed primeres, and liquid chromatography-mass spectrometry analysis. Results of this study confirmed the specificity of the newly designed PCR primers. The presence of producers of puwainaphycins/minutissamides was proven at both tested localities.

The waters of the open ocean constitute the largest living space on Earth but despite its obvious significance to the biosphere, the open ocean remains an unexplored frontier. With a regional focus on the Indian Ocean, this thesis investigates (i) the distribution of pelagic biota on basin scales, (ii) the effect of abrupt topography on pelagic biota and their predator-prey relationships, and (iii) the use of genetic techniques to elucidate population connectivity and dispersal of pelagic taxa. (i) Pelagic scattering layers (SLs) were surveyed with scientific echosounders across the southwest (SWIO) and central Indian Ocean to investigate their vertical and geographical distribution. Structurally distinct SL regimes were found across the Subantarctic Front, and may explain recently observed foraging behaviours of southern elephant seals. Regression models indicated a close relationship between sea surface temperature and mean volume backscatter, with significantly elevated backscatter in the subtropical convergence zone. The heterogeneous distribution of scattering layer biota may have implications for predator foraging and carbon cycling in the Indian Ocean. (ii) Acoustic surveys revealed diverse interactions between SLs, aggregations and topography around islands as well as shallow ( < 200m) and intermediate (200-800m) seamounts at spatial scales from 1 to 100 km. Epi-and mesopelagic backscatter was increased around reefs and banks of the Chagos archipelago, indicating connectivity between oceanic and neritic systems. SWIO seamounts harboured summit-associated aggregations, but the distributions of surrounding SLs did not follow a general pattern. Downstream SL depletion was observed in one location and combined with stomach content analyses, provides an insight into the mechanics of prey flux between open-ocean and seamount ecosystems. (iii) A mitochondrial marker was used to assess the population structure and demography of the hatchetfish Argyropelecus aculeatus in the SWIO. The results are suggestive of a single, well-connected population and indicate a recent population expansion around 0.14 million years ago. This highlights that even highly abundant mesopelagic populations are vulnerable to global climatic changes. Dispersal and recruitment are key ecological processes structuring seamount communities and are directly relevant for the management of exploited populations. Genetic barcoding was evaluated as a means to identify cryptic larval specimens of eels (leptocephali) and spiny lobsters (phyllosomata). Identification success was limited, but indicated the presence of 3-4 phyllosoma clades and 5-6 leptocephalus clades along the SWIR.

Plankton microorganisms constitute the base of the marine food web and play a great role in global atmospheric carbon dioxide drawdown. Moreover, being very sensitive to any environmental changes they allow noticing (and potentially counteracting) them faster than with any other means. As such they not only influence the fishery industry but are also frequently used to analyze changes in exploited coastal areas and the influence of these interferences on local environment and climate. As a consequence, there is a strong need for highly efficient systems allowing long time and large volume observation of plankton communities. This would provide us with better understanding of plankton role on global climate as well as help maintain the fragile environmental equilibrium. The adopted sensors typically provide huge amounts of data that must be processed efficiently without the need for intensive manual work of specialists. A new system for general purpose particle analysis in large volumes is presented. It has been designed and optimized for the continuous plankton monitoring problem; however, it can be easily applied as a versatile moving fluids analysis tool or in any other application in which targets to be detected and identified move in a unidirectional flux. The proposed system is composed of three stages: data acquisition, targets detection and their identification. Dedicated optical hardware is used to record images of small particles immersed in the water flux. Targets detection is performed using a Visual Rhythm-based method which greatly accelerates the processing time and allows higher volume throughput. The proposed method detects, counts and measures organisms present in water flux passing in front of the camera. Moreover, the developed software allows saving cropped plankton images which not only greatly reduces required storage space but also constitutes the input for their automatic identification. In order to assure maximal performance (up to 720 MB/s) the algorithm was implemented using CUDA for GPGPU. The method was tested on a large dataset and compared with alternative frame-by-frame approach. The obtained plankton images were used to build a classifier that is applied to automatically identify organisms in plankton analysis experiments. For this purpose a dedicated feature extracting software was developed. Various subsets of the 55 shape characteristics were tested with different off-the-shelf learning models. The best accuracy of approximately 92% was obtained with Support Vector Machines. This result is comparable to the average expert manual identification performance. This work was developed under joint supervision with Professor Rubens Lopes (IO-USP). / Microorganismos planctônicos constituem a base da cadeia alimentar marinha e desempenham um grande papel na redução do dióxido de carbono na atmosfera. Além disso, são muito sensíveis a alterações ambientais e permitem perceber (e potencialmente neutralizar) as mesmas mais rapidamente do que em qualquer outro meio. Como tal, não só influenciam a indústria da pesca, mas também são frequentemente utilizados para analisar as mudanças nas zonas costeiras exploradas e a influência destas interferências no ambiente e clima locais. Como consequência, existe uma forte necessidade de desenvolver sistemas altamente eficientes, que permitam observar comunidades planctônicas em grandes escalas de tempo e volume. Isso nos fornece uma melhor compreensão do papel do plâncton no clima global, bem como ajuda a manter o equilíbrio do frágil meio ambiente. Os sensores utilizados normalmente fornecem grandes quantidades de dados que devem ser processados de forma eficiente sem a necessidade do trabalho manual intensivo de especialistas. Um novo sistema de monitoramento de plâncton em grandes volumes é apresentado. Foi desenvolvido e otimizado para o monitoramento contínuo de plâncton; no entanto, pode ser aplicado como uma ferramenta versátil para a análise de fluídos em movimento ou em qualquer aplicação que visa detectar e identificar movimento em fluxo unidirecional. O sistema proposto é composto de três estágios: aquisição de dados, detecção de alvos e suas identificações. O equipamento óptico é utilizado para gravar imagens de pequenas particulas imersas no fluxo de água. A detecção de alvos é realizada pelo método baseado no Ritmo Visual, que acelera significativamente o tempo de processamento e permite um maior fluxo de volume. O método proposto detecta, conta e mede organismos presentes na passagem do fluxo de água em frente ao sensor da câmera. Além disso, o software desenvolvido permite salvar imagens segmentadas de plâncton, que não só reduz consideravelmente o espaço de armazenamento necessário, mas também constitui a entrada para a sua identificação automática. Para garantir o desempenho máximo de até 720 MB/s, o algoritmo foi implementado utilizando CUDA para GPGPU. O método foi testado em um grande conjunto de dados e comparado com a abordagem alternativa de quadro-a-quadro. As imagens obtidas foram utilizadas para construir um classificador que é aplicado na identificação automática de organismos em experimentos de análise de plâncton. Por este motivo desenvolveu-se um software para extração de características. Diversos subconjuntos das 55 características foram testados através de modelos de aprendizagem disponíveis. A melhor exatidão de aproximadamente 92% foi obtida através da máquina de vetores de suporte. Este resultado é comparável à identificação manual média realizada por especialistas. Este trabalho foi desenvolvido sob a co-orientacao do Professor Rubens Lopes (IO-USP).

Several lakes located in the blast zone of the 1980 eruption of Mount St. Helens were studied to determine if the plankton in the lakes were limited in their growth by nitrogen or phosphorus availability. Long term nutrient enrichment experiments were performed on lake water from five lakes and measures of chlorophyll-a, carbon fixation, and nutrient uptake were used to evaluate the extent of limitation. Nutrient concentrations, ratios, and uptake from 14 lakes provided additional evidence for limitation by nitrogen and/or phosphorus. The physical, chemical, and biological characteristics of the lakes were also examined to monitor the return of these lakes to pre-eruption conditions. Lakes heavily impacted by the 1980 eruption (Ryan, Fawn, and Hanaford Lakes) and newly formed Castle Lake produced positive responses to nitrogen additions, reflecting the continuing importance of nitrogen in these lakes. Evidence for colimitation by phosphorous was apparent in some of the lakes. Venus Lake, located near the fringe of the blast zone received less organic debris and responded only with phosphorus additions. Evaluations of nutrient depletion from the photic zones of these lakes during the growing season supported the results from the bioassays. Two measures of growth (chlorophyll-a and carbon fixation) often responded differently to enrichment, suggesting active algal and bacterial communities in some of the lakes. Qualitative measures of alkaline phosphatase activity normalized to chlorophyll-a indicated that those lakes which contained the highest dissolved organic carbon and bacterial counts following the eruption had higher alkaline phosphatase activities than less impacted lakes, suggesting bacterial enzyme production. Alkaline phosphatase activity confirmed that phosphorus limitation was more severe in lakes having less soluble reactive phosphorus. Most of the lakes appear to be similar to other Cascade Mountain lakes, although many have reduced dissolved oxygen concentrations in the bottom waters during stratification. Incomplete mixing of these lakes during periods of circulation may regulate the decomposition of the organic material and hence, full recovery of these lakes.

Dilution and ash weathering are the most important processes controlling the ion chemistry of lakes in the Mount St. Helens blast zone. Gibbs' models indicated total dissolved solids were decreasing as a result of dilution from high precipitation and runoff and the lakes plot in the rock weathering dominated region. Plots of theoretical dilution curves indicated a decline in ion concentrations as a result of dilution. Ion concentrations followed the exponential decline predicted by the dilution curve, although concentrations were higher than predicted by the curve. Increased concentrations were a result of the rapid weathering of ash in basins and on lake bottoms. Rapid weathering of ash in lake watersheds and on lake bottoms continues to influence the ionic concentrations of the lakes. In general, sodium and potassium have declined at a much faster rate than calcium or magnesium. Slower relative declines in concentrations of calcium and magnesium were a result of more rapid rate of leaching of calcium and magnesium from the ash. Ash in the watersheds will continue to be a major contributor to the overall ion chemistry of the lakes until such time as the watersheds are stabilized by vegetation and a permanent soil layer. Ash on lake bottoms will be unavailable as sources of ionic constituents when it becomes buried within deep sediment layers. Ion concentrations observed in study lakes affected by the eruption were similar to those observed in control lakes with few exceptions. Although ion concentrations in affected lakes have declined to values observed in control lakes, most were at higher concentrations than the regional means. Several functions of the ion chemistry were used to correlate planktonic community structure to lake ion chemistry. The data suggested ion chemistry was not influencing biological community structures as no patterns emerged. Analysis of diatom populations with respect to monovalent:divalent cation ratios showed no correlation.

The Deepwater Horizon oil spill that started on April 20, 2010, in the Gulf of Mexico was the largest marine oil spill in the history of the petroleum industry. There was an unexpected and prolonged sedimentation event of oil-associated marine snow to the seafloor due to the oil spill. The sedimentation event occurred because of the coagulation process among oil associated marine particles. Marine scientists are developing models for the coagulation process of marine particles and oil, in order to estimate the amount of oil that may reach the seafloor along with marine particles. These models, used certain assumptions regarding the shape and the texture parameters of marine particles. Such assumptions may not be based on accurate information or may vary during and after the oil spill. The work performed here provided a quantitative analysis of the assumptions used in modeling the coagulation process of marine particles. It also investigated the changes in model parameters (shape and texture) during and after the Deepwater Horizon oil spill in different seasons (spring and summer). An Interactive Visualization Application was developed for data exploration and visual analysis of the trends in these parameters. An Interactive Statistical Analysis Application was developed to create a statistical summary of these parameter values.

Thesis: Thesis (Ph. D.)--Joint Program in Applied Ocean Science and Engineering (Massachusetts Institute of Technology, Department of Civil and Environmental Engineering; and the Woods Hole Oceanographic Institution), 2019 / Cataloged from PDF version of thesis. / Includes bibliographical references (pages 126-137). / Although seawater appears uniform at scales that humans often interact with and sample, the world that marine microbes inhabit can be highly heterogeneous, with numerous biological and physical processes giving rise to resource hotspots where nutrient concentrations exceed background levels by orders of magnitude. While the impact of this microscale heterogeneity has been investigated in the laboratory with microbial isolates and theoretical models, microbial ecologists have lacked adequate tools to interrogate microscale processes directly in the natural environment. Within this thesis I introduce three new technologies that enable interrogation of microbial processes at the microscale in natural marine communities. The IFCB-Sorter acquires images and sorts individual phytoplankton cells, directly from seawater, allowing studies exploring connections between the diversity of forms present in the plankton and genetic variability at the single-cell level. / The In Situ Chemotaxis Assay (ISCA) is a field-going microfluidic device designed to probe the distribution and role of motility behavior among microbes in aquatic environments. By creating microscale hotspots that simulate naturally occurring ones, the ISCA makes it possible to examine the role of microbial chemotaxis in resource acquisition, phytoplankton-bacteria interactions, and host-symbiont systems. Finally, the Millifluidic In Situ Enrichment (MISE) is an instrument that enables the study of rapid shifts in gene expression that permit microbial communities to exploit chemical hotspots in the ocean. The MISE subjects natural microbial communities to a chemical amendment and preserves their RNA in a minute-scale time series. / Leveraging an array of milliliter-volume wells, the MISE allows comparison of community gene expression in response to a chemical stimulus to that of a control, enabling elucidation of the strategies employed by marine microbes to survive and thrive in fluctuating environments. Together, this suite of instruments enables culture-independent examination of microbial life at the microscale and will empower microbial ecologists to develop a more holistic understanding of how interactions at the scale of individual microbes impact processes in marine ecosystems at a global scale. / by Bennett S. Lambert. / Thesis (Ph. D.)--Joint Program in Applied Ocean Science and Engineering (Massachusetts Institute of Technology, Department of Civil and Environmental Engineering; and the Woods Hole Oceanographic Institution) / Thesis(Ph.D.)--JointPrograminAppliedOceanScienceandEngineering(MassachusettsInstituteofTechnology,DepartmentofCivilandEnvironmentalEngineering;andtheWoodsHoleOceanographicInstitution)

Marine microorganisms play a significant role in the cycling of nutrients in the open ocean through production, consumption, and degradation of organic matter (OM). Carbon (C), nitrogen (N), and phosphorus (P) are essential ingredients in every known recipe for life. However, the cycling of each of these elements proceeds at different rates such that the ratio of C:N:P can vary widely between particulate, dissolved, organic, and inorganic pools. To better understand the mechanisms controlling these transformations, this study investigated the bacterial remineralization of photosynthetically-derived organic matter derived from cultures of Trichodesmium IMS101, Thalassiosira weissflogii, Prochlorococcus MED4, and particulate material collected from the surface waters of an upwelling regime. Experiments were conducted at sea for a short duration (<6d) and in the laboratory for longer periods (<150 days). In all treatments, across experiments, we observed rapid and selective P remineralization independent of the type of organic material added. Full solubilization and remineralization of P typically occurred within a week. Conversely, N remineralization was slower, with only 39-45% of particulate N (PN) remineralized in shorter (6d) experiments and 55-75% of PN remineralized in <150d experiments. Nitrification was observed after 70-98 days depending on the remineralizing bacteria (isolated from either the Oregon coastal upwelling regime or the North Pacific Subtropical Gyre (NPSG). Notably, these events did not transform the full complement of ammonium to nitrate. This differential lability between N and P led to rapid changes in the N:P ratio of inorganic pools as organic matter was depolymerized by varying bacterial populations. The variable input of potentially limiting elements could have consequences for primary productivity and particle export. Finally, we observed that in short-term experiments with heterotrophic bacteria collected from the NPSG, the N:P ratio of remineralization (11 ± 2.2) was independent of the N:P of added organic material (5-23). This uniformity of inorganic ratios implies differential lability and N:P composition of residual semi-labile and refractory organic matter. Formation of refractory C and N rich organic matter, often termed the microbial pump, is a significant pathway for the transport and sequestration of elements in the aphotic zone of the ocean interior. The experimental results reported here suggest that differential supply of POM leads to rapid and preferential P remineralization, N:P remineralization independent of the N:P of added substrates, and variable N:P of residual organic matter. These findings help constrain our knowledge of elemental cycling in the marine environment. / Graduation date: 2013

Ph.d.-afhandling, Roskilde Universitet, 2005. / Haves kun i elektronisk udg. Thesis (Ph.D.). 6 ill., 1tabel; approx. 50 ref. Summaries (Da, En).

Planktonic ecosystems provide a key mechanism for the transfer of CO2 from the atmosphere to the deep ocean via the so-called "biological pump". Mathematical models of these ecosystems have been used to predict CO2 uptake in surface waters, and more recently have been embedded in global climate models. While the equilibrium properties of these models are well studied, less attention has been paid to their response to external perturbations, despite the fact that as a result of the variability of environmental forcing such ecosystems are rarely, if ever, in equilibrium. Human induced perturbations to these ecosystems, namely the addition of limiting nutrients (e.g. iron) to areas where nitrate is plentiful to accelerate the biological pump, have been proposed as a solution to reduce atmospheric CO2. Linear theory is used to determine the structure of "unit-norm" perturbations (size in mmol N m^-3) to state variables of an ecosystem model in steady state, describing Ocean Station P (50N 145W) in summer, that optimize either instantaneous export flux of organic matter at fixed times or integrated export as the ecosystem relaxes towards equilibrium. For all perturbations, the flux to higher trophic levels is the primary contributor to export flux, the contribution of aggregation is negligible, and (sinking) detritus increases significantly in the transient dynamics. Two perturbations considered optimize instantaneous export flux; both perturbations synchronize P1 and Z1 relative to their predator prey cycle, resulting in a maximum instantaneous export flux of 4.4 mmol N m^-2 d^-1, and also increased integrated export above that at steady state (6 g C m^-2 over 150 days). An increase in larger phytoplankton (P2), representing diatoms, results in the highest integrated export (7 g C m^-2). The perturbations in which P2 persist the longest give the highest integrated export, and these perturbations are primarily increases in P2. The additional integrated export in response to a proportional increase to steady state concentrations of both large and small phytoplankton is positive, but much lower than the optimal perturbations. However, the additional integrated export in response to an increase in only P1 is negligible. The linear and nonlinear ecosystem and export responses to two perturbations are compared; for perturbations of magnitude 0.5 mmol N m^-3, the linearization of the ecosystem dynamics, rather than of the export flux, is the primary cause for differences between the fully linear and fully nonlinear cases.