Maite Maldonado

Professor

Relevant Degree Programs

 

Graduate Student Supervision

Doctoral Student Supervision (Jan 2008 - Mar 2019)
The physiological and proteomic response of two strains of the diatom Thalassiosira oceanica to copper and/or iron limitation (2018)

Diatoms are responsible for over 20% of the Earth's photosynthetic productivity, thus impact global fisheries, biogeochemical cycles and climate. However, marine primary productivity is limited by the micronutrient iron (Fe) in ~40% of the ocean. Diatoms inhabiting these regions have evolved unique physiological strategies to survive under these extremely low Fe conditions. Several physiological adaptations to Fe-limitation in diatoms require an increased dependency on copper (Cu), suggesting an interaction between Fe and Cu nutrition and the potential for Fe-Cu co-limitation in these regions. Though some published work has illustrated transcriptomic and proteomic adaptations of some diatoms to low Fe, there is limited knowledge on diatoms' acclimation strategies to low Cu or Fe-Cu co-limitation. My thesis research focused on elucidating concomitant physiological and proteomic responses of two strains of the open ocean diatom Thalassiosira oceanica (CCMP1003 and CCMP1005) acclimated to Fe, Cu, and Fe-Cu limiting conditions. I measured over 20 physiological parameters, including carbon assimilation rate, oxygen production, respiration rate, an array of photosynthetic parameters [such as electron transport rate of photosystem II (ETRPSII), non-photochemical quenching (NPQ), and antenna absorption cross section of photosystem II, ϬPSII], and photosynthetic rates as a function of light intensity. Moreover, I investigated the differential expression of proteins in T. oceanica in response to these three metal limitations, using stable isotope dimethyl labelling proteomics.I first describe the physiological and proteomic responses to Cu limitation in T. oceanica, focusing on the changes to and the interplay among proteins and pathways involved in the light reactions of photosynthesis, the carbon and nitrogen metabolisms, and the prevention of oxidative stress. I then compare unique changes to the photosynthetic apparatus induced by each metal limitation (Fe, Cu and Fe-Cu) vs. changes induced by general cellular stress. Furthermore, given that my research investigated two strains of T. oceanica, I uncovered stunning intraspecific differences in their proteomic and physiological responses to trace metal limitation. My research unveiled a comprehensive restructuring of the photosynthetic apparatus, and a sophisticated interaction among metabolic pathways in T. oceanica (CCMP1003) in response to low metal availability (especially Cu), demonstrating exceptional adaptations to low trace metal availabilities.

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Biogeochemical cycling of copper in the Northeast Pacific Ocean : role of marine heterotrophic bacteria (2017)

Copper (Cu) is important in regulating microbial activity in the ocean, as it can act both as a limiting nutrient and a toxic inhibitor depending on its concentration. Yet, our knowledge of its biogeochemical cycle is limited in many oceanic regions including the subarctic Northeast (NE) Pacific, as is our knowledge of Cu nutrition in marine heterotrophic bacteria. To address this, I investigated Cu biogeochemical cycling along a coastal‒oceanic transect, Line P, in the subarctic NE Pacific (Chapter 2). I also explored physiological responses to varying Cu availability (limiting to sufficient) of taxonomically diverse heterotrophic bacteria, which include isolates from surface waters of the Line P transect (Flavobacteriia member: Dokdonia sp. Dokd-P16, and Gammaproteobacteria members Pseudoalteromonas sp. strain PAlt-P2 [coastal] and PAlt-P26 [oceanic]), and a member of the marine Roseobacter clade within class Alphaproteobacteria (Ruegeria pomeroyi DSS-3). Several important processes were identified to moderate dissolved Cu along Line P. These include fluvial and sedimentary inputs (near the coast), upwelling of deep, Cu-rich waters in the Alaskan gyre (offshore), atmospheric inputs (offshore), as well as scavenging within the intermediate waters of the Oxygen Minimum Zone (OMZ) across the transect. Bacterial responses to changing Cu availability were diverse. Flavobacteriia member Dokd-P16 reduced its growth rate, carbon metabolism, and Cu quota (Cu:P) under Cu limitation, but enhanced its Mn quota. In contrast, both Pseudoalteromonas spp. were mostly unaffected by different Cu levels. Ruegeria pomeroyi maintained constant growth rates but moderated quotas of several metals (under low Cu: decreased Cu and Co, but increased Mn and Fe quotas), and some aspects of its C metabolism. These findings illuminate on the role of Cu in shaping bacterial species composition in the ocean, and the bacterially-mediated cycles of carbon and bioactive metals (i.e. Fe, Zn, Mn, Co). Copper quotas of heterotrophic bacteria are similar to those of cultured marine phytoplankton. Estimates of Cu partitioning between these planktonic groups in the euphotic zone of the NE Pacific revealed that up to 50% of biogenic Cu could be associated with bacterial biomass. Therefore, marine heterotrophic bacteria should not be overlooked in studies of Cu biogeochemical cycling.

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On the light and iron dependent coupling of carbon fixation and photosynthetic electron transport in Arctic and Subarctic marine phytoplankton (2016)

Marine phytoplankton primary productivity, the photosynthetic conversion of CO₂ into organic carbon by microscopic photosynthetic algae in the surface ocean, plays a fundamental role in ecosystem dynamics and global biogeochemical cycles. Consequently, the ability to accurately measure, monitor and predict environmental influences on this process over a range of spatial and temporal scales is crucial. The work presented in this thesis evaluates the application of fast repetition rate fluorometry (FRRF) for instantaneous, high resolution estimates of phytoplankton primary productivity. Results from both laboratory experiments and field work in Arctic and Subarctic marine waters show that the conversion factor required to derive carbon-based primary productivity estimates from FRRF-derived rates of electron transport in photosystem II (ETR) varies significantly in response to the interacting effects of iron and light availability (Chapter 2), over diurnal cycles (Chapter 3), and in response to nitrogen and light availability under low temperatures (Chapter 4). At a photo-physiological level, a high conversion factor is observed under conditions of excess excitation energy, where the amount of light energy absorbed in the pigment antenna exceeds the capacity for downstream metabolic processes, i.e. carbon fixation. Phytoplankton employ numerous mechanisms to alleviate excess excitation energy after charge separation, and these processes are postulated to be responsible for the increased de-coupling of ETR and carbon fixation. Consistent with this hypothesis, a strong correlation was observed between the derived conversion factor and the dissipation of excess excitation energy before charge separation, which can be estimated as non-photochemical quenching (NPQ). Because NPQ can be estimated from FRRF measurements, it can be used as a proxy for the magnitude and variability of the conversion factor between carbon fixation and ETR, and this approach holds potential to significantly improve carbon-based primary productivity estimates from FRRF measurements. The work presented in this thesis advances our understanding of the coupling between light absorption, photo-chemistry, and carbon fixation in response to various environmental gradients. The experimental approach taken demonstrates how an appreciation of photo-physiological processes of photosynthesis is critical for improved estimates of phytoplankton primary productivity at regional scales.

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Copper nutrition and transport mechanisms in plankton communities in the northeast Pacific Ocean (2014)

Copper (Cu) is an essential micronutrient for phytoplankton, particularly during iron limitation, but can also be toxic at relatively low concentrations. While Cu stoichiometry and metabolic functions in marine phytoplankton have been studied, little is known about the substrates for Cu transport and the Cu nutritional state of indigenous phytoplankton communities. The aim of my thesis was two-fold: investigate the bioavailability of organically bound Cu to laboratory and indigenous phytoplankton, and evaluate Cu nutrition of phytoplankton along Line P, a coastal- open ocean transect in the northeast subarctic Pacific Ocean. Organically complexed Cu was bioavailable to four laboratory phytoplankton strains and an Fe-limited phytoplankton community. A laboratory investigation of the substrates for the high-affinity Cu transport system in the model diatom Thalassiosira pseudonana confirmed that organically complexed Cu(II) can be acquired, and likely via extracellular reduction and internalization of Cu(I). Cellular uptake rates of the laboratory strains were similar to those estimated for the natural phytoplankton assemblage, and provide additional evidence that some in situ Cu ligand complexes are likely bioavailable. Using bottle incubations, I investigated the potential for Cu limitation and toxicity in open ocean Fe-limited phytoplankton communities. In 2010, I provided physiological evidence for an interaction between Fe and Cu metabolisms in an Fe-limited phytoplankton community. In 2011, Cu availability to an Fe-limited community was reduced, using a strong Cu(II)-specific ligand, resulting in slower Cu uptake rates, faster growth rates, and increased cyanobacteria abundance. Despite large variations in macronutrient, light, and iron along Line P in 2011, net primary productivity was negatively correlated with inorganic Cu concentrations, and positively correlated with the strength of the in situ ligands. The potential roles of Cu ligands in the sea are discussed, highlighting that the bioavailability of in situ organic Cu complexes is a key determinant for marine primary productivity.

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Copper requirements and acquisition mechanisms in marine phytoplankton (2012)

Copper nutrition is essential for the growth of marine diatoms, especially under Fe-limiting conditions. I investigated the physiology of Cu in marine phytoplankton by studying how the Fe and Cu availability interact and control Cu uptake and demand. I used short-lived Cu radioisotopes to determine the Cu requirements and steady-state Cu transport rates (ρssCu) of ten species from three classes of marine phytoplankton, isolated from coastal and oceanic environments. I also determined the Cu uptake kinetics of two marine centric diatoms (Thalassiosira oceanica and T. pseudonana) grown under various Fe and Cu conditions (sufficient and limiting). Furthermore, putative genes encoding potential components of Cu transport and homeostasis were identified in T. pseudonana, and their expression was investigated. Copper had significant effects on growth rates and ρssCu of the oceanic phytoplankton, but not the coastal strains. Although Cu quotas (Cu:C) were not significantly higher in oceanic strains, there were five independent lines of evidence supporting a more important role of Cu in the physiology of oceanic phytoplankton. Distinct biphasic Cu transport rates as a function of Cu concentrations were observed in the centric diatoms, Thalassiosira oceanica and T. pseudonana, suggesting the presence of a high- and a low-affinity Cu transport system. The high-affinity Cu transport system followed Michaelis-Menten saturation kinetics, but was controlled differently by Fe and/or Cu availability. A strong interaction between Fe and Cu nutrition in controlling the expression of genes encoding Cu transport and homeostasis was observed. Most genes, including putative Cu transporters (CTR), Cu transporting P-type ATPases, Cu chaperones and putative Zn transporters in T. pseudonana were up-regulated by low Fe, while low Cu either had no effect or the effect was dependent on Fe availability. These results suggest a complex interaction between Cu and Fe response networks. The function of a putative Cu transporter (CTR) in T. pseudonana was examined using functional complementation of Saccharomyces cerevisiae ctr1Δctr3Δmutant. Though the results were inconclusive, various explanations for these findings were discussed. This thesis highlights a complex interaction between Fe and Cu nutrition in marine phytoplankton at the protein and gene expression level.

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The role of phytoplankton and environmental variables in Pacific oyster (Crassostrea gigas) aquaculture in British Columbia (2012)

Pacific oyster aquaculture in British Columbia faces serious challenges, such as high cadmium (Cd) concentrations, low growth and high mortalities during summer, and inability to directly gauge stress levels. The goal of this dissertation was to address these challenges by investigating the role of biological, physical and chemical oceanographic parameters in controlling them in various oyster farms in the Strait of Georgia. Three studies were undertaken. The first, from August 2004 to July 2005, investigated the role of phytoplankton in controlling Cd levels in the oysters in a Deep Bay farm. Phytoplankton mediated the transfer of dissolved Cd to the particulate phase, accounting for 90% of the summer reduction in dissolved Cd. This suggests that phytoplankton act as a sink for dissolved Cd, reducing the main source to the oysters. Two descriptive models for annual oyster Cd concentrations were developed based on environmental variables. The second study, from June to October of 2008, investigated how environmental factors, culture depth and seed size controlled oyster mortality and growth in four farms. Farms with less stratified, colder waters rich in diatoms fared better than those with highly stratified, higher temperature waters and persistent blooms of flagellates. Larger oyster seed presented low mortalities, while smaller seed were more susceptible to adverse conditions due to their ineffective particle processing capabilities. The best yield was obtained at a culture depth of 3 m, despite higher mortalities. A depth manipulation technique was investigated as a means to reduce summer mortalities without success. The third study, during the summer and fall of 2007 in Deep Bay, investigated a novel proteomic technique to detect and quantify heat-shock proteins (HSP) 70 and 90 in oysters to assess their stress levels. Mortalities were relatively low during that year (8.5% accumulated). The abundance of HSP 70 sequences was positively directly with non-harmful diatom biomass and negatively with high temperature and reproductive state. In contrast, the levels of HSP 90 were correlated negatively to the biomass of non-harmful diatoms, and positively to that of potentially-harmful algae, indicating that HSP 70s and 90s may have different triggers.

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Master's Student Supervision (2010-2017)
Particulate trace metals & iron availability to phytoplankton in a changing Arctic Ocean (2017)

This thesis, focusing on the Canadian Arctic Ocean, investigates the cycling of particulate trace metals, and the bioavailability of iron to phytoplankton in this rapidly changing ocean. Full depth profiles of particulate Al, Cd, Pb, P, V, Mn, Fe, Co, Cu, Zn and Ba were determined. Trace elements displayed various vertical distributions. Firstly, some elements had a strong lithogenic component (Al, Fe and V), and were characterized by a maximum at the surface. Indeed, their concentrations strongly correlated with each other across basins. Secondly, elements with a significant biogenic component (Cd and Cu) were characterized by a decrease in concentration with depth. Furthermore, preferential remineralization of P over Cd at shallow depths in numerous stations is reported. In some stations across basins, the molar ratio of particulate Cu and P approached a plateau at the meso- or bathy-pelagic zone, suggesting the presence of ammonium oxidizing archaea, which require Cu-dependent enzymes. Thirdly, Mn, a trace metal with a predominant redox cycle, showed a spike in concentration between 100-200 m, as well as a bottom enrichment. In the Canada Basin, we suggest interactions between the production of manganese oxide, cobalt oxide and barite by manganese oxidizing bacteria. The Arctic Ocean is experiencing the greatest decrease in seawater pH, as well as rapid ice melting which elevates light intensity in surface waters. To examine changes in Fe bioavailability to Arctic phytoplankton under a varying environment, two incubation experiments were conducted. After natural phytoplankton assemblages were acclimated to different light/CO2 treatments for one week, short-term Fe uptake assays were performed to assess the capability of phytoplankton to access Fe. Generally, Fe uptake capability was positively influenced by CO₂ level, and negatively impacted by light level in the incubations. These observations imply that high CO₂ has a significant negative effect on Fe bioavailability, while high light has a positive effect. Furthermore, when comparing future scenario (higher atmospheric CO₂ and underwater irradiance) with present-day conditions, the bioavailability of Fe to phytoplankton appeared to be similar.

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Spring and summer phytoplankton community dynamics and comparison of FRRF- and ¹³C-derived measurements of primary productivity in Rivers Inlet, British Columbia (2013)

Spring and summer phytoplankton community dynamics were monitored in the temperate coastal fjord, Rivers Inlet, British Columbia, to understand their impact on the growth of juvenile sockeye salmon. Spatial patterns in timing and magnitude of the diatom-dominated spring bloom appeared to be controlled by differences in mixing and stratification. At the sheltered head of the inlet, riverine input stratified the water column, the phytoplankton bloom appeared earlier and more intense. At the well-mixed mouth, where currents diluted the phytoplankton seed population, the bloom appeared delayed and reduced in intensity. The spring bloom was terminated by nitrate depletion except where salinity was low and phosphate became limiting due to oligotrophic freshwater input. While the spring community was diatom-dominated, the summer community was more diverse, with increased abundance of flagellates and ciliates.Primary productivity measurements using fast repetition rate fluorometry (FRRF) correlated well (~1:1) with estimates derived from 2-hour ¹³C-uptake incubations when phytoplankton were healthy in the lab. In contrast, under high light or low nutrient conditions, FRRF underestimates primary productivity. Community composition may also influence FRRF estimates of primary productivity. We calculated annual FRRF-derived primary productivity of 550 – 1100 g C · m⁻² · yr⁻¹. Potential sockeye production was estimated using primary productivity estimates derived from our ¹³C data. These calculations suggest that the sockeye carrying capacity of the fjord and lake are well matched and that current levels of primary production could support the ~10-fold higher historical sockeye returns. This implies that contemporary and historical levels of primary production are similar and are not the cause of the sockeye decline. More likely, a timing mismatch between trophic levels is negatively impacting sockeye smolts. Our results suggest that future sockeye production in Rivers Inlet may be negatively impacted by increased freshwater input and stratification, both of which may be influenced by climate warming. This physical forcing may precipitate a shift to a flagellate-dominated phytoplankton community, a longer food chain, and reduced energy transfer to smolts. Continued monitoring of phytoplankton dynamics is critical for refining predictions of ecosystem change and facilitating improvements in sockeye stock management policies.

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Studies in Fe bioavailability : co-limitation of primary productivity by iron, light, and nitrate in the Beaufort Sea, and direct iron-siderophore uptake mechanisms in Fe deficient phytoplankton (2012)

This study explores two relevant questions in the realm of iron (Fe) bioavailability to phytoplankton. First, does Fe availability limit (or co-limit) growth of indigenous plankton communities in the Arctic Ocean? Second, can phytoplankton internalize ferrated siderophores with a non-reductive uptake mechanism? To address the first question, an 8-day grow out experiment was conducted in the Beaufort Sea in early September 2009, during which light, Fe, and nitrate (NO₃⁻) levels were manipulated. Bottles were sampled on days 0, 2, 4, 6, and 8 for accessory pigments, size-fractionated chlorophyll α, phytoplankton abundance and composition, nutrients, Fe quotas and uptake rates. It was found that NO₃⁻ was limiting plankton growth at the time of sampling. The community also appeared to be light limited. Additionally, co-limitation of primary production by Fe and light at light levels ≤ 10 % surface irradiance (I₀) was observed. These results have interesting implications about how the seasonality of NO₃⁻, light, and Fe availability may control primary productivity in the Beaufort Sea. To address the second question, I investigated the potential of a non-reductive Fe uptake mechanism for siderophore-bound Fe in the model diatom Thalassiosira oceanica and the in situ plankton communities along Line P in the subarctic Pacific Ocean in late summer 2010. To do this, we radiolabeled the siderophore desferrioxamine B (DFB) by methylating its terminal amine group with radioisotope ¹⁴C methyl iodide. Internalization of ⁵⁵Fe¹⁴DFB was observed both in phytoplankton cultures and field communities along Line P, suggesting the presence of a non-reductive Fe uptake mechanism in phytoplankton. However, the results are inconclusive due to the inability to purify and verify the concentration of ¹⁴DFB. The overarching goal of this investigation was to gain a better understanding on the bioavailability and acquisition of Fe by phytoplankton. This is imperative in order to predict the role of Fe in future primary productivity, and subsequently the fate of phytoplankton communities and the biological carbon pump, as our oceans respond to global warming.

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