Thesis (M. Phil.)--University of Hong Kong, 1987.

Thesis (Ph. D.)--College of William and Mary. / Vita. Includes bibliographical references. Also available online.

Currently, the US Environmental Protection Agency primarily regulates the discharge of dissolved nitrogen and phosphorous from wastewater treatment plants in the United States. A recent study has shown that the treated effluent of many plants contains concentrations of dissolved carbon dioxide well above the expected theoretical equilibrium concentration of 0.6 mg/L, indicating that carbon dioxide may have been overlooked as a possible pollutant in receiving waters. For this reason, it is necessary to examine the possible presence of a discharge plume containing high levels of dissolved CO2 downstream from the outfall of a major wastewater treatment plant in Tampa, Florida. To examine this possibility, discharge data at the Howard F. Curren Advanced Wastewater Treatment Plant was collected over a two-week period and fed into the UM3 submerged discharge model to simulate discharge conditions at peak ebb tide. In all, five separate runs of the model were performed and compared to examine plume rise, spreading rate, average dissolved CO2 concentration, and plume path. The model predicts that, for this scenario, the plume rises fairly rapidly and is also quickly diluted to near-ambient concentrations of dissolved carbon dioxide within a short distance of being discharged. While this would seem to indicate that the effects of Howard F. Curren on Tampa Bay, in terms of dissolved CO2, are negligible major limitations of the UM3 model make it difficult to say this with a great deal of certainty.

Based on a municipal wastewater treatment plant (WWTP) in Tampa, FL, a dynamic multiple-systems model was developed on the STELLA software platform to explore algae biomass production in wastewater by incorporating two photobioreactors into the WWTP‟s treatment train. Using a mass balance approach, the model examined the synergy through algal growth and substrate removal kinetics, as well as macroeconomic-level analyses of algal biomass conversion to biodiesel, biogas, or fertilizer. A sensitivity analysis showed that biomass production is highly dependent on Monod variables and harvesting regime, and profitability was sensitive to processing costs, market prices of products, and energy environment. The model demonstrated that adequate nutrients and carbon dioxide are available in the plant‟s influent to sustain algal growth. Biogas and fertilizer production were found to be profitable, but biodiesel was not, due to high processing costs under current technologies. Useful in determining the growth potential on a macro-level, the model is a tool for identifying focus areas for bench and pilot scale testing.

The marine diatom genus Pseudo-nitzschia includes species that produce domoic acid, a neurotoxin responsible for illness and mortality in both humans and marine wildlife. Because of the expertise and time required for the microscopic discrimination of species, molecular methods that monitor environmental concentrations of Pseudo-nitzschia provide a rapid alternative for the early detection of blooms and prediction of toxin accumulation. We have developed a nucleic acid sequence-based amplification with internal control RNA (IC-NASBA) assay and a quantitative reverse transcription PCR (qRT-PCR) assay for the detection of the toxic species P. multiseries targeting the ribulose- 1,5-biphosphate carboxylase/oxygenase small subunit (rbcS) gene. Both methods use RNA amplification and fluorescence-based real-time detection. Due to a limited rbcS sequence database, primers were designed and used to sequence this gene from 14 strains of Pseudo-nitzschia (including four P. multiseries) and 19 other marine diatoms. The IC-NASBA and qRT-PCR assays had a limit of detection of one cultured cell of P. multiseries and were linear over four and five orders of magnitude, respectively (r2 ! 0.98). Neither of the assays detected closely related organisms outside the Pseudo-nitzschia genus, and the qRT-PCR assay was specific to P. multiseries. While cross-reactivity of primers with unknown species prevented reliable detection of P. multiseries in spiked environmental samples using IC-NASBA, the qRT-PCR assay had positive detection from 107 cells/L to 103 cells/L. Nearly a 1:1 relationship was observed between predicted and calculated cell concentrations using qRT-PCR. Based on a diel expression study, the rbcS transcript copy number per cell ranged from 2.16 x 104 to 5.35 x 104, with the highest expression during early to mid photoperiod. The rbcS qRT-PCR assay is useful for the detection and enumeration of low concentrations of P. multiseries in the environment.

No description available.

Algae are photosynthetic microorganisms that convert carbon dioxide and sunlight into biomass that can be used for biofuel production. Although they are usually cultivated in suspension, these microorganisms are capable of forming productive biofilms over substrata given the right conditions. This dissertation focuses on algal biofilms and their application in biofuel feedstock production. In particular it reports the construction and performance of an algae biofilm photobioreactor, the physico-chemical surface properties of different algal species and adhesion substrata, and cell-surface interactions based on experimental results and theoretical models. A novel algae biofilm photobioreactor was constructed and operated (i) to demonstrate the proof of concept, (ii) to analyze the performance of the system, and (iii) to determine the key advantages and short comings for further research. The results indicated that significant reductions in water and energy requirements were possible with the biofilm photobioreactor. Although the system achieved net energy ratio of about 6, the overall productivity was low as Botryococcus branunii is notoriously slow growing algae. Thus, further studies were focused on identification of algal species capable of biofilm growth with larger biomass and lipid productivities. Adhesion of cells to substrata precedes the formation of all biofilms. A comprehensive study has been conducted to determine the interactions of a planktonic and a benthic algal species with hydrophilic and hydrophobic substrata. The physico-chemical surface properties of the algal cells and substrata were determined and using these data, cell-substrata interactions were modeled with the thermodynamic, Derjaguin, Landau Verwey, Overbeek (DLVO) and Extended Derjaguin, Landau, Verwey, Overbeek (XDLVO) approaches and critical parameters for algal adhesion were identified. Finally, the adhesion rate and strength of algal species were quantified with parallel plate flow chamber experiments. The results indicated that both cell and substrata surface hydrophobicity played a critical role for the adhesion rate and strength of the cells and XDLVO approach was the most accurate model. Finally, based on these findings the physico-chemical surface properties of ten algal species and six substrata were quantified and a screening was done to determine algae species substratum couples favoring adhesion and biofilm formation. / text

There is limited information currently available regarding the underlying physiological responses and molecular mechanisms of osmoregulation, acetate metabolism [in relation to the synthesis of glycerolipids, polyunsaturated fatty acids (PUFA), and ichthyotoxins], and transport in Prymnesium parvum N. Carter, a microalga that causes devastating harmful algal blooms (HAB) worldwide. This dissertation examines gene expression under environmental conditions that are associated with HAB formation, including phosphate limitation and low salinity, using microarrays and RNA sequencing (RNA-Seq). A comparative fatty acid methyl ester (FAME) analysis at 30 vs. 5 practical salinity units (psu) was performed to gain additional insight into acetate metabolism. The RNA-Seq analysis included a de novo assembly of the P. parvum transcriptome, generating 47,289 transcripts, of which 35.4% were identifiable. This permitted the evaluation of the expression of many more genes compared with the microarray analysis, which examined ~3,500 genes. Relevant candidate genes identified included those whose products are involved in osmolyte production, salinity stress, and ion transport. With respect to the putative synthesis of polyketide ichthyotoxins, 32 different polyketide synthase (PKS) transcripts were identified in the transcriptome assembly, none of which were differentially expressed. Hemolysin and monogalactosyldiacylglycerol synthase were downregulated at 30 vs. 5 psu, suggesting the increased presence of additional ichthyotoxins at the lower salinity. Evidence for several PUFA synthesis pathways was also revealed. Fatty acid compositions were largely similar at the two salinities, containing relatively prominent quantities of the PUFA stearidonic acid, but compositions varied among strains. The transcription of genes whose products are associated with vesicular transport was elevated, and higher levels of extracellular prymnesins were observed in HAB-forming conditions. Thus, with regard to acetate metabolism, I have revealed evidence for the post-transcriptional regulation of the production of prymnesins and the contributory effects of hemolysin, monogalactosyldiacylglycerol, and PUFA towards ichthyotoxicity. Further, I propose that toxin transport is triggered in HAB-forming conditions, in which the toxins are actively being excreted. Collectively, these data shed light on the transcriptional responses that occur following alterations in phosphate availability and salinity, including those associated with the synthesis and delivery of a number of potential ichthyotoxins from P. parvum. / text

The effects of the toxic dinoflagellate, karenia brevis (Wilson clone), on larval survival and development of the northern quahog (=hard clam, Mercenaria mercenaria), eastern oyster (Crassostrea virginica) and bay scallop (Argopecten irradians) were studied in the laboratory. The effects of K. brevis on feeding activities of juveniles from these species plus the green mussel (Perna viridis) were also examined. Finally, adult bay scallops were exposed to K. brevis for two weeks to investigate possible cytotoxic effects. Survival of 3-day-old larvae was generally > 85% for all shellfish species at Karenia brevis densities of 100 cells . ml-1 or less, and not significantly different between whole and lysed culture. At 1,000 cells . ml-1, survival was significantly less in lysed culture than whole culture for both M. mercenaria and C. virginica. Survival of 7-day-old larvae in all species was not significantly affected at densities up to 1,000 cells . ml-1. At 5,000 cells . ml-1, however, survival was reduced to 37, 26 and 19% for A. irradians, M. mercenaria and C. virginica, respectively. Development of C. virginica and M. mercenaria larvae was protracted at K. brevis densities of 1,000 cells . ml-1. Clearance rates of juveniles were determined under static and flow-through conditions using whole and lysed cultures of K. brevis. The bay scallop was most sensitive, exhibiting a 79% reduction in clearance rate at 1,000 cells . ml-1 of whole culture. The eastern oyster was least responsive, showing a 38% reduction in clearance rate between the same treatments. The green mussel and the northern quahog displayed intermediate responses. Similar results were observed during longer (2 day) exposures to a continuous supply of K. brevis. Bay scallops showed a significant decline in clearance rate at 100 cells . ml-1 after 24 hr exposure; clearance rate of oysters was not affected by K. brevis at this concentration. No mortality was observed for any species during these brief exposures. Adult bay scallops exposed to K. brevis for two weeks showed degenerative and inflammatory changes in the digestive gland, including reduced thickness of the epithelium, increased size of digestive tubule lumens and hemocytic infiltration. The prospect for recovery of bay scallop populations in Florida may be hampered by recurring blooms of K. brevis.

Biofuel is a renewable and sustainable energy source with near-neutral carbon footprint. Algae are an ideal feedstock for biofuel production because they reproduce quickly and have high oil. Algae can be cultivated in non-arable land, and would not impact the food supply. Unfortunately, processing algae into biofuel is more expensive than land crops due to the large volumes of dilute algal suspension that must be harvested and concentrated. In order to improve algae-based biofuel economics, resins were developed that reduce costs associated with water pumping and transport. Hydrophobic resins were developed for binding oil out of an algal suspension so that the residual biomass could be recovered without solvent contamination. Binding behavior displayed lipid species specificity, and binding capacity was improved by ethanol treatment of the biomass. Algae was harvested by binding to anion exchange resin and directly converted into biodiesel. One-step, room temperature in situ transesterification of algae yielded nearly as much biodiesel as two-step, heated transesterification of dried biomass. Elution with transesterification reagent also regenerated the resin for subsequent algal binding. Functionalized resins were developed with high algal binding capacity at neutral pH. Binding was easily reversed, as treatment with buffer with pH higher than pKa of the resin functional group removed the algae and regenerated the resin for subsequent use. The resin bound 10% of its weight in algae and released it as a 100-fold concentrated suspension. The polymers developed can be scaled up for commercial processes and reduce algal harvesting and concentration costs. / text