The focus of my thesis research is on the interplay between individual behavior, population dynamics and community-level processes within zooplankton communities in coastal Maine. The target organisms of my thesis work are marine copepods. Copepods are small (1-10 mm) crustaceans that perform the essential ecosystem function of consuming and assimilating primary production (phytoplankton) making it available to higher trophic levels such as commercially important fishes. Therefore, copepod population growth is of critical importance to marine food webs. Fertilization limitation has been suggested as a constraint on copepod population growth but field surveys describing the prevalence of fertilization limitation are lacking. During my doctoral research, I explored the in situ fertilization success of two marine copepod species, Temora longicornis and Eurytemora herdmani in coastal Maine. I collected monthly zooplankton samples and analyzed clutches from field-caught females using an egg-staining technique. My results indicate that both species exhibit fertilization limitation in nature and the factors correlated with their fertilization span population, community and ecosystem level factors. To determine a causal relationship between predator density and copepod mating success, I conducted laboratory experiments to assess the effects of a common mysid shrimp predator, Neomysis americana on Eurytemora herdmani mating success. I subjected males and females to predators or predator cues. I found that the presence of a mysid predator, or only a predator cue, reduced copulation frequency and spermatophore transfer leading to a 38-61% decrease in E. herdmani nauplii production. These results suggest that mysid predators can constrain copepod population growth through non-consumptive processes. To determine the effects that resources can impose on copepod behavior, I explored the behavioral and fitness consequences of Temora longicornis ingesting Alexandrium fundyense, a phytoplankton species that forms harmful algal blooms in coastal Maine. My results suggest that ingesting A. fundyense causes copepods to swim faster and with more directional persistence compared to control algae. Temora longicornis increased their average swimming velocity by 24%, which leads to a 24-54% increase in their theoretical encounter rate with predators. Therefore, these findings suggest behaviorally mediated copepod-algal interactions may have significant impacts on harmful algal bloom dynamics and the fate of toxins in marine food webs.

An innovative wastewater treatment technology was developed to recover renewable resources, such as water, energy and nutrients, from sewage. First, a novel synthetic sewage was evaluated for its suitability to serve as an alternative substrate for lab-scale wastewater treatment (WWT) research. Based on granular dried cat food, Complex Organic Particulate Artificial Sewage (COPAS) is a commercially-available, flexible, and easy to preserve feed. Characteristics of COPAS, namely chemical composition, disintegration/dissolution kinetics, and anaerobic biodegradability, were determined. Anaerobic bioassays indicate that COPAS is highly biodegradable at the concentration used to simulate household sewage (1000 mg/L), with more than 72% of the theoretical methane content reached after 30 d of incubation. Results indicate that COPAS is a suitable substrate as a surrogate of domestic sewage. In the second stage of the research, a lab-scale, 10L gas-lift anaerobic membrane bioreactor (Gl-AnMBR) was designed, fabricated and tested. The AnMBR is a hybrid treatment technology that combines anaerobic biological treatment with low-pressure membrane filtration. Although AnMBR has been used in many instances for the treatment of high strength industrial or agricultural wastewater, relatively little has been reported about its application for the treatment of domestic sewage and further conversion and recovery of resources embedded in sewage, such as energy and nutrient enriched water. The 10L column reactor uses a tubular PVDF ultrafiltration membrane (with biogas as sparge gas) for sludge/water separation. COPAS was used as synthetic feed (at 1000 mg/L) to represent household wastewater. The configuration showed excellent removal efficiencies of organic matter (up to 98% and 95% in COD and TOC removal, respectively) while producing energy in the form of methane at quantities suitable for maintaining membrane scrubbing (4.5 L/d of biogas). Soluble nutrients were recovered in the effluent in the forms of NH4, (9.1±4.2 mg/L), NO3 (2.2±0.9 mg/L) and PO4 (20±7.13 mg/L). The energy footprint (net energy) of this reactor was evaluated and the energy requirements per volume of permeate produced was found to be in the range of -1.2 to 0.7 kWh/m3, depending on final conversion of methane to electric or thermal energy respectively. These values could potentially be improved towards energy surplus (-2.3 to -0.5 kWh/m3) if applied to plant scale operation, which would employ more efficient pumps than those used in the lab. Results from this study suggest that the Gl-AnMBR can be applied as a sustainable treatment tool for resource recovery from sewage, which can further be optimized for large scale operation. In the final stage of this research, further resource recovery from sewage was investigated by coupling the Gl-AnMBR with an innovative gas-lift algal photo MBR (APMBR). To our knowledge, this is the first reported application of membranes (in particular gas-lift tubular) for separation of algal cells from effluent in a continuous-flow photobioreactor. Nutrient rich effluent (9 mg/L NH4-N and 20 mg/L PO4-P) from the Gl-AnMBR treating domestic wastewater was used as substrate to grow the biofuel producing microalgae Chlorella sorokiniana (Cs). The initial set of operational conditions tested in this study (HRT of 24 hours, operational flux of 4.5 LMH, air-lift flow rate (Qa) of 0.1 L/min and 0.1 bars of membrane inlet pressure), achieved 100% removal efficiencies for NH4 and PO4. Flux remained constant during the experimental period which demonstrated the efficacy of gas lift as a membrane fouling control strategy for an algae bioreactor. Because the algae is photoautotrophic, little removal of organic carbon was expected nor observed. Further studies are required to better understand the fate and cycling of carbon in the APMBR. Limited information is available in the literature regarding biofuel-producing, algal photo MBRs utilizing anaerobic effluents as feedstock, which makes this study an important step in understanding the design and performance of combined anaerobic/algal biotechnology for large scale application of wastewater resource recovery.

ABSTRACT Benthic foraminifers that host algal symbionts are similar to corals in that they rely on their algal endosymbionts for their energy needs, calcify prolifically, and are sensitive to changes in environmental conditions. They are abundant in the benthos of coastal coral-reef areas and are found throughout the tropical and subtropical regions. Pulse Amplitude Modulated (PAM) chlorophyll fluorometry and chlorophyll a extraction techniques were used to quantify and compare the photosynthetic responses of the benthic foraminiferal, Archaias angulatus and their isolated endosymbionts, Chlamydomonas hedleyi, to short-term changes in temperature. Maximum quantum efficiency (Fv/Fm) and rapid light curves (RLCs), from which relative electron transport rates (rETR) of photosystem II (PSII) were derived, were investigated over a thermal range from 4.4° to 33.9 °C in three experiments that were 7 to 31 days in duration. Typical mean yields (Fv/Fm for healthy holobionts (symbionts in hospite) were 0.6 - 0.7, and for isolated symbionts 0.5 - 0.6. Chronic photoinhibition, indicated by significant decreases in Fv/Fm, occurred at temperatures above 31.0°C; there was minimal reduction in efficiency in cooler treatments. The trends between holobiont and symbionts were very similar in all of the photophysiological parameters measured [yield, photoefficiency (<α>), ETRmax and minimum saturating irradiance (Ek)] and supported the temperature range findings in terms of the tolerance of the specimens in the low temperatures up to 31.0 °C. For all photochemical measurements assessed, the holobiont values tended to be somewhat higher than those for the symbionts, with the exception of Ek, possibly indicating a tight coupling in the host-symbiont response during photosynthesis. Chlorophyll a (<μ>g/foram) was negatively correlated with temperature (r = -0.37, p < 0.001) in Experiments 1 and 2. However, in all 3 experiments, chlorophyll a was variable, suggesting a high degree of individual variability in A. angulatus and the ability to acclimate. Some differences observed among treatments may be related to differences in seasons when the specimens were collected and in length of time in culture prior to experiments. Holobiont median rETR light curve trends and photophysiological derived parameters recorded median Ek ranges of ~100-150 <μ>mol photons m-2 s-1, observed ETRmax light intensities of ~200 <μ>mol photons m-2 s-1 and photoinhibition, induced by increasing irradiance intensities, which occurred > 500 <μ>mol photons m-2 s-1. These light curve trends and derived parameters generally supported previous photosynthesis O2 and CO2 gas production studies of A. angulatus. The differences in responses associated with acclimation should be considered in design of future experimental studies. This was the first known physiological study of the viable temperature range and photobiology of A. angulatus using chlorophyll fluorometry methods. Though commonly found in Caribbean and Atlantic waters ranging from 14.0 - 31.0 °C, these results indicate a wider thermal-tolerance range for A. angulatus than was previously known. Keywords: Foraminifera, Chlamydomonas sp., PAM fluorometry, photosynthesis, algal symbiosis

published_or_final_version / Applied Geosciences / Master / Master of Science

published_or_final_version / Civil Engineering / Doctoral / Doctor of Philosophy

Unicellular microalgae are a renewable bioresource that can meet the challenge forfood and energy in a growing world population. Using sunlight, CO2, nutrients,and water, algal cells produce biomass in the form of sugars, proteins and oils, allof which carry commercial value as food, feed and bioenergy. Flue gas CO2 andwastewater nutrients are inexpensive sources of carbon and fertilizers. Microalgaecan mitigate CO2 emissions and reduce nutrients from waste streams whileproducing valuable biomass.My focus was on some of the challenging aspects of cultivating microalgae ascrop: the response of biomass production and quality to seasonality, nutrients andbiological interactions. Approach spans from laboratory experiments to large-scaleoutdoor cultivation, using single microalgal strains and natural communities insouthern (Portugal) and northern (Sweden) Europe.Half of the seasonal variation in algal oil content was due to changes in light andtemperature in outdoor large-scale cultures of a commercial strain (Nannochloropsisoculata). Seasonal changes also influence algal oil composition with more neutrallipids stored in cells during high light and temperature. Nitrogen (N) stress usuallyenhances lipid storage but suppresses biomass production. Our manipulationshowed that N stress produced more lipids while retaining biomass. Thus,projecting annual biomass and oil yields requires accounting for both seasonalchanges and N stress to optimize lipid production in commercial applications.Baltic Sea microalgae proved to be a potential biological solution to reduce CO2emissions from cement flue gas with valuable biomass production. A multi-speciescultivation approach rather than single-species revealed that natural or constructedcommunities of microalgae can produce equivalent biomass quality. Diversecommunities of microalgae can offer resilience and stability due to more efficientresource utilization with less risk of contamination, less work and cost for culturemaintenance.Stable algal biomass production (annual basis) was achieved in outdoor pilot-scale(1600 L) cultivation of Baltic Sea natural communities using cement flue gas as aCO2 source. Results indicate favorable algal oil content at northern Europeanlatitudes compared to southern European latitudes.My thesis establishes the potential of cultivating microalgae as a bioresource inScandinavia, and using a community approach may be one step towardssustainable algal technology.

This work explores the relationship between nutrient loading and changes in water quality in a sub-tropical, above-ground, off-stream municipal water supply reservoir, the C.W. Bill Young Regional Reservoir. The three source waters for the reservoir have varied but high levels of nutrients such as phosphorus and nitrogen. In other reservoirs, these nutrients have been linked to deterioration of water quality and increased expense in water treatment. The need to minimize excess nutrients results led to the primary research question: what allocation of withdrawals from the three sources will minimize the deterioration of water quality? To answer this question, the relationship between nutrient and other water quality data, such as temperature,phosphorus, chlorophyll a, Secchi depth, and trophic state indices were explored. Results indicate that temperature had a correlation with observed water quality. 27.9% of the variability in trophic state index as a function of Chlorophyll a was correlated with average temperature at one foot below water level. Correlation and regression models were developed using available time-series of linear and log-transformed water quality data to predict Chlorophyll a response. The parameters used in the model were selected from correlation matrices and from the P value in the multiple regression. The models developed were significant at P < 0.05. In the developed models, temperature was found to have greater predictive strength than nutrients indicating that this reservoir may be more strongly influenced by season and light than by nutrient limitation. Lastly, the US Army Corps of Engineers' eutrophication model, BATHTUB, was used to simulate different loading conditions and trophic response. The model results indicate that use of water the middle pool or lower pool of the Tampa Bypass Canal yield similar trophic states with the middle pool slightly lower. Use of water from the Alafia River yielded the highest trophic state and would be expected to have negative impacts on water quality.

This dissertation includes molecular and phytochemical investigations of the harmful, bloom-forming alga, Prymnesium parvum, including analysis of known polyketide metabolites as a function of salinity and growth. Initially, the development of molecular and phytochemical tools was necessary for the detection and quantification of P. parvum and its associated toxins. Suites of oligonucleotides and molecular beacons were designed for conventional and quantitative multiplex PCR to amplify four species- and gene-specific products simultaneously that were used for the detection and quantitation of P. parvum. This built-in redundancy provided increased confidence in reactions with the positive confirmation of four discrete products. Techniques were also developed for the chemical enrichment of toxins produced by P. parvum. Until now, isolation of “prymnesins” has never been reproduced. Polyketide prymnesins possess unique spectral properties that were used to generate an LC-MS fingerprint that comprised 13 ion species. Preliminary investigations using chemifluorimetric methods were also capable of detecting prymnesins in the pico- and nano-molar range. Environmental samples were tested as an independent assessment of these methods. Lastly, the roles of polyketide prymnesins were analyzed with respect to total hemolytic activity (HA) as a function of culture age and salinity. Variation in HA of supernatants was statistically significant relative to both variables (p << 0.05). Salinity was inversely related to HA wherein cultures growing in 5-25 psu were 150-200% more hemolytic. Total HA was inversely related to culture age during the first three weeks, but positively related to it during the next three weeks. Interestingly, no hemolysis was detected in fractions containing prymnesins from culture supernatants and the majority of hemolysins remained in the aqueous phase. Prymnesins extracted from cells varied significantly over the 6-week observation period (p << 0.05); HA was positively correlated during the first half and inversely related during the last half of the study. Salinity was directly related to HA from cell extracts, but these effects were not significantly different until the last three weeks. These investigations suggest that polyketide prymnesins are present at much lower quantities than previously believed, and they may not be the key compounds associated with hemolysis due to P. parvum. / text

This thesis reports the rheological properties of algae slurries as a function of cell concentration for three microalgae species: Nannochloris sp.,Chlorella vulgaris, and Phaeodactylum tricornutum. Rheological properties ofalgae slurries have a direct impact on the agitation and pumping power requirements as well as process design for producing algal biofuels. This study measures the rheological properties of eight diff erent concentrations of each species ranging from 0.5 to 80 kg dry biomass/m³. Strain-controlled steady rate sweep tests were performed for each sample with an ARES-TA rheometer using a double wall couette cup and bob attachment. Shear rates ranged from 5 - 270 s⁻¹, corresponding to typical expected conditions. The results showed that Nannochloris sp. slurry behaved as a Newtonian fluid for concentrations up to 20 kg/m³. Samples with concentrations above 40 kg/m³ behaved as a shear thinning non-Newtonian fluid. The effective viscosity increased with increased biomass concentration for a maximum value of 3.3x10⁻³ Pa-s. Similarly, C. vulgaris slurry behaved as a Newtonian fluid with concentrations of up to 40 kg/m³, above which it displayed a shear thinning non-Newtonianf behavior and a maximum eff ective viscosity of 3.5x10⁻² Pa-s. On the other hand, P. tricornutum slurry demonstrated solely Newtonian fluid behavior, with the dynamic viscosity increasing with increasing biomass concentration for a maximum value of 3.2x10⁻³ Pa-s. The maximum observed e ffective viscosity occurred at a concentration of 80 kg/m³ for all three species. Moreover, an energy analysis was performed where a non-dimensional bioenergy transport e ffectiveness was de termined as the ratio of the energy content of the transported algae biomass to the sum of the required pumping power and the harvesting power. The results show that the increase in major losses due to increase in viscosity was overcompensated by the increase in the transported biomass energy. Also, cultivating a more concentrated slurry requires less dewatering power and is the preferred option. The largest bioenergy transport eff ectiveness was observed for the slurries with the largest initial dry biomass concentrations. Finally, the relative viscosity of algae slurries was modeled using a Kelvin-Voit based model for dilute and concentrated viscoelastic par- ticle suspensions. The model, which depends primarily on the packing factor of the algae species, agrees with the measured viscosity with an average error of 18%, while the concentrated particle suspension model was slightly more accurate than the dilute suspension model. / text

Limited studies focused on reactions occurring during disinfection and oxidation processes of seawater. The aim of this work was to investigate disinfection by-products (DBPs) formation from the chlorination and chloramination of seawater algal organic matter and related model compounds. Simulated algal blooms directly growing in Red Sea, red tide samples collected during an algal bloom event and Hymenomonas sp. monoculture were studied as algal organic matter sources. Experiments were conducted in synthetic seawater containing bromide ion. A variety of DBPs was formed from the chlorination and chloramination of algal organic matter. Brominated DBPs (bromoform, DBAA, DBAN and DBAcAm) were the dominant species. Iodinated DBPs (CIAcAm and iodinated THMs) were detected, which are known to be highly toxic compared to their chlorinated or brominated analogues. Algal organic matter was found to incorporate important precursors of nitrogenous DBPs (N-DBPs), which have been reported to be more toxic than regulated THMs and HAAs. Isotopically-labeled monochloramine (15N- NH2Cl) was used in order to investigate the nitrogen source in N-DBPs. High formation of N-DBPs was found from Hymenomonas sp. sample in exponential growth phase, which was enriched in nitrogen-containing organic compounds. High inorganic nitrogen incorporation was found from the algal samples enriched in humic-like compounds. HAcAms formation was studied from chlorination and chloramination of amino acids. Asparagine, aspartic acid and other amino acids with an aromatic structure were found to be important precursors of HAcAms and DCAN. Factors affecting HAcAms formation (Cl2/ amino acid molar ratio and pH) were evaluated. Studies on the formation kinetics of DCAcAm and DCAN from asparagine suggested a rapid formation of DCAcAm from organic nitrogen (amide group) and a slower incorporation of inorganic nitrogen coming from monochloramine to form DCAN. High amounts of DCAN and DCAcAm were detected from the chloramination of aromatic compounds (i.e., phenol and resorcinol) indicating that N-DBPs can also be formed from organic compounds without any organic nitrogen through the incorporation of inorganic nitrogen from monochloramine. Moreover, results from Hymenomonas sp., aromatic amino acids, and phenolic compounds suggested that aromatic compounds are highly reactive with monochloramine and a major fraction of DBP precursors.