Thesis (Ph. D.)--University of California, Los Angeles, 2003. / Vita. Includes bibliographical references.

Many contaminants, such as industrial chemicals, fertilizers, herbicides, pharmaceuticals and heavy metals are released to e environment. 3,4-dichloroaniline（3,4-DCA） originated from degradation of some herbicides such as diuron, propanil and linuron, is toxic to aquatic organisms and affects human being immune system. Triclosan, widely used as antimicrobial agent in pharmaceuticals and personal care products (PPCPs), has been detected as contaminat in various aquatic environments. In this work, green algae were isolated from local environment, then applied for the removal and biodegradation of 3,4-DCA and triclosan. Two axenic pure algae were isolated using the solid agar method. One of the algae was identified morphologically as Desmodesmus sp. based on the experimental results. The other one was identified morphologically as Chlorella pyrenoidosa by accredited authority. At the same time, alga S. obliqnus was obtained commercially. All the three green algae were cultured in tris-acetate-phosphate (TAP) medium. Firstly, the alga C. pyrenoidosa was applied to remove and biodegrade 3,4-DCA with a concentration of 4.6 μg/mL for 7 d. A removal percentage of 78.4% was obtained over a 7-d period. Two major metabolites with less toxicity were identified as 3,4-dichloroformanilide and 3,4-dichloroacetanilide using HPLC-ESI-ion trap-MS. iii Secondly, all the three green microalgae species including C. pyrenoidosa, Desmodesmus sp., and S. obliqnus, were compared in the removal and biodegradation of triclosan in aqueous medium. When triclosan with concentration of 400 ng/mL was cultured with the three algal species separately, triclosan was quickly eliminated from medium in the 1 d cultivation by algae with removal percentages of 62.4%, 92.9% and 99.7% for C. pyrenoidosa, Desmodesmus sp. and S. obliqnus, respectively. The dominant mechanism for the removal of triclosan by C. pyrenoidosa was determined as cellular uptake. Biotransfromation of triclosan involved hydroxylation and methylation, glucose conjugation was determined as the predominant mechanisms for the removal of triclosan by algae Desmodesmus sp. and S. obliqnus. The intermediates from hydroxylation, reductive dechlorination, or ether bond cleavage were immediately subjected to glucosylation and/or methylation via the hydroxyl group of triclosan or introduced, which served as detoxification mechanisms of the chlorinated aromatic chemicals. In order to find the intermediates in the metabolic pathway of triclosan by algae, Desmodesmus sp. was exposed to 400 ng/mL triclosan. 2,4-DCP was detected during the cultivation period 3-12 h using ultra performance liquid performance (UPLC)-ESI-MS/MS. The metabolites from multi metabolic reaction like the glucose conjugate of hydroxylated triclosan were detected in the first 30 min after exposure. The metabolites as products from glucosylation and consecutive hydroxylation and methylation of triclosan or 2,4-DCP were detected after 3 h iv cultivation. To provide more information about the reductive capability of C. pyrenoidosa, the reaction between C. pyrenoidosa and triclosan was investigated. When C. pyrenoidosa was exposed to triclosan with concentration from 100 to 800 ng/mL, more than 50% of triclosan was eliminated by algal uptake from the culture medium during the first 1 h exposure. In the biodegradation experiments, a major metabolite from the reductive dechlorination of triclosan was identified by using liquid chromatography (LC)-ESI-MS. The ability of reductive dechlorination of C. pyrenoidosa might potential application for bioremediation of polychlorinated biphenyls (PCBs) that with similar chemical structure to triclosan, but belonging to the catagory of persistent organic pollutants (POPs). Through the TEM observation, it was found that the triclan treatment resulted in the disruption of the chloroplast of algal cells, which indicated that triclosan may affect membrane metabolism.

Saline deterioration in the South African public water system has been documented and disposal of brine wastes has been identified as part of the problem. The broad aim of this research programme was to undertake an initial technical study to evaluate the feasibility of integrating algal biotechnology into a disposal function for these wastes. A demonstration of utility in the form of products and waste treatment could produce a beneficiation of saline effluents and provide incentives necessary to deal with the disposal issue. The study attempted to demonstrate a synthesis between the two main thrusts in algal biotechnology that have produced large-scale practical applications - stable, predictable algal production in saline media and the cost effective High Rate Oxidation Ponding (HROP) process for incorporating algal production into a waste treatment function. Tannery organic saline effluents and the biotechnology of Dunaliella salina culture producing β- carotene were chosen as paradigms for the study. 1. The alga was shown to grow in certain tannery effluents producing enhanced biomass yields compared to defined inorganic medium cultivation. The potential for amino acid or protein supplementation of defmed culture media was noted. 2. A reduction in organic load simultaneous with the growth of D.salina was recorded in laboratory-scale simulations of the HROP process. Rates similar to the fresh water HROP equivalent were demonstrated. 3. These results suggested the uptake and storage of organic nitrogen by D.salina. The consequent inhibition of β-carotene accumulation by the organism presented a potentially insurmountable obstacle to the feasibility of β-carotene production in this medium. Uptake and release of organic compounds, previously demonstrated in phytoplankton and other micro-algae, was confirmed in this study for D.salina. The evidence acquired indicated the internalization of both glycine and bovine serum albumin. An ultrastructural study demonstrated mechanisms by which this process might occur. 4. The release of substantial quantities of glycerol was shown. A mechanism whereby D. salina may use this to regulate ammonia availability via control of its associated bacterial population was observed. Glycerol release was identified as presenting an application in treating refractory organic wastes, such as secondary sewage sludges, by elevating C:N ratios. This could demonstrate a significant utility for brine waste impoundments. 5. A multistage production process was proposed to deal with the problem of β-carotene inhibition by separation of the growth and metabolite accumulation functions into separate unit operations. It was shown in this study that the stress of nitrogen deficiency combined with high salinity provides for effectiveβ-carotene accumulation under the conditions of low illumination that pertain in dense cultures. Subjected to these conditions effluent-grown cells show delayed but unimpaired {j-carotene accumulation. 6. A role for the plant hormone abscisic acid in mediating the stress response was demonstrated in D.salina. Fluorescence induction studies suggested the presence of a signalling process forming part of a sensitivity control mechanism. Stress induction of β-carotene accumulation could occur through four clearly defined stages. Potential was identified for using this response as a physiological probe for monitoring and regulating the stress induction process. 7. The multistage processing concept requires effective algal cell separation technology. The use of cross-flow ultrafiltration and diafiltration with a polyethersulfone tubular membrane system was demonstrated as an effective process for the recovery and washing of D. salina. Cell concentrates were produced in a viable form. 8. Process designs incorporating the findings of the research programme are presented demonstrating how effluent and organic waste treatment functions may be combined with the production of D.salina and its products. Application of the multi-stage processing concept to β-carotene production in a defined medium process was identified as offering a potential four-fold yield enhancement. This could have a significant impact on a high cost, marginal algal biotechnology process. Aspects of novelty have been claimed in provisional patents applications. A provisional demonstration of the feasibility of D.salina production in tannery effluent indicates that algal biotechnology may provide a utility for, and hence the beneficiation of saline effluent wastes.

No description available.

Haptophytes are renowned for the most prominent and biogeochemically important group of marine calcifiers: coccolithophores. The unexpected discovery of a unique, silicifying member of this clade - Prymnesium neolepis - prompted questions regarding mechanisms of silicification and their origin in the calcifier-dominated haptophytes. To address these questions I used cell physiology, biochemistry and molecular approaches, investigating cellular and molecular mechanisms involved in silicification in haptophytes. Comparisons of this system with calcification in coccolithophores and other silica-based systems in eukaryotes were also made. Here I report that P. neolepis is an obligate silicifier, producing silica scales in a process fundamentally different to that observed in coccolithophores. Scale deposition and secretion in P. neolepis is localized in the posterior, vacuolar part of the cell rather than in the anterior part near the flagellar roots as in calcifying coccolithophores. The organic matrix underlying silica scales in P. neolepis was found to be non-homologous with organic scales, which in coccolithophores serve as coccolith baseplates. This suggests, that silica scales and coccoliths arise from two distinct, most likely non-homologous processes, which is further supported by the comparative investigation of the role of cytoskeleton in silica scale production in P. neolepis and coccolithogenesis in a representative calcifier, Coccolithus pelagicus. Using cytoskeleton inhibitors I established, that the cytoskeleton components used for morphogenesis and secretion of biomineralised structures are different in these two systems. Analysis of P. neolepis biosilica revealed the presence of an intimately associated organic fraction consisting of a putatively chitin-containing material, potentially serving as an organic matrix underlying silica scales. Further biochemical investigation of the biosilica-associated organics confirmed the presence of long chain polyamines (LCPAs) dissimilar to those previously reported in diatoms and sponges. Additionally, a potentially novel, proline and lysine-rich protein sharing a weak homology with lipocalins was recovered, suggesting that this silicification system is unique to haptophytes. Several theories concerning acquisition of the ability to silicify in haptophytes were proposed. Overall, the findings presented in this study provide a detailed description of Si biomineralisation system in this unique, silicifying haptophyte and supply novel information on biomineralisation systems in marine haptophytes. This study contributes a basis on which the phenomenon of silicification in haptophytes can be further investigated, as well as novel information which can be further used in elucidation of origins of silicification in algae and other Eucarya.

This thesis aims to initially define the present knowledge of pathogens which infect algae, highlighting the potential economic significance of such pathogens, given the recent rise in algaculture. Focus is given to the oomycetes, a group of organisms which already contain several significant genera of plant (Phytophthora) and animal (Saprolegnia) pathogens, as well as Olpidiopsis; a genus reported to significantly impact algaculture in Asia. Subsequent chapters aim to genetically characterise stramenopiles (specifically oomycetes and hyphochytrids) associated with algae in two ways. Firstly known pathogens of algae, Olpidiopsis feldmanni and Anisolpidium rosenvingei are morphologically diagnosed by microscopy and genetically surveyed at phylogenetically significant loci. Further morphological information on these two pathogens is reported here expanding the current knowledge of these rarely reported organisms. Surprisingly, genetic evidence indicates that Anisolpidium, a uniflagellate genus, belongs to the biflagellate class Oomycota and not the closely related uniflagellate class Hyphochytriomycota, as previously suspected. Morphological and genetic features of these classes are contrasted to justify this molecular interpretation. These first two studies, along with the publically available sequences of algal pathogens are then used to design primers, which enable an oomycete-directed metagenomic survey of brown algae, allowing the investigation of what organisms are associated with these. At present the results of this survey are unreported. Finally a Eurychasma dicksonii transcriptome as well as genomic sequences of other stramenopiles are investigated for the presence of cadherin protein models. At present such models are automatically annotated and reported. This study is the first manual curation of the protein and defines a unique protein family which is presented by the oomycete genera Eurychasma, Pythium, Phytophthora and Albugo as well as the labyrinthulomycete Aurantiochytrium limanicum. The protein model is not reported from other sequenced oomycete genera or photosynthetic stramenopiles.

K. mikimotoi has been classified under many names and has been mis-assigned to different species and genera in the North Atlantic and Pacific because of its morphological similarities to other Gymnodinoid species. It is now known to be widely distributed, but there remain unresolved questions about whether K. mikimotoi was introduced into the North Sea from Japanese waters, or whether it has always inhabited this region and been erroneously classified as Gymnodinium spp. or has been a part of the hidden flora prior to be recognised in a bloom off the Norwegian coast in 1966. To address questions about geographical genetic variation within K. mikimotoi and broader issues about its biogeography it was deemed important to develop a suitable diagnostic molecular marker that could then be used to monitor the presence/absence of different K. mikimotoi ecotypes over long time scales in European waters. This study showed that the partial rDNA LSU (D1-D2) was too conserved to separate the different strains of K. mikimotoi, while, the ITS region was better able to discriminate between the different strains. However, the rbcL gene was the most informative gene and contained sufficient substitutions to separate the different strains of K. mikimotoi. Specific PCR-primers were designed to amplify a variable region of the rbcL gene able to distinguish differences between K. mikimotoi isolates from the different regions. The innovative high resolution melting temperature (HRM) technique based on specific primer set allowed rapid discrimination of K. mikimotoi from distinct geographic localities (= sequence variants) that differed by only a single nucleotide. Moreover, this study used archival environmental samples collected from the Celtic Sea shelf-break region. The high resolution melting temperature assay successfully detected the European K. mikimotoi isolate within the south-western English Channel in a 1963 sample, which is prior to thefirst report of a K. mikimotoi bloom in Norwegian waters in 1966 and in the south-western English Channel in 1975 and in western Japan in 1965. HRM observations were further validated using clone libraries and sequencing. In summary, this data provided more information about the genotypes present over the analysed timescales, revealing that K. mikimotoi sub-species 2 (European and New Zealand strains) was present in south-western English Channel and south-west Ireland for over 47 years, with sub-species 1 (the Japanese isolate) being absent from all examined samples. This finding supports the hypothesis that K. mikimotoi isolates within Europe are not of Japanese origin and suggests that they are native species to the region.

published_or_final_version / Biological Sciences / Master / Master of Philosophy

Supplies of nonrenewable fossil fuels are becoming more limited even as they continue to contribute to pollution and economic concerns. Alternative sources of energy must be developed that help minimize these problems. One potential source of energy is the production of biofuels from algae. Here we evaluate algae found in South Texas brackish water ponds used for aquaculture of fish as a possible source of biofuels. In particular, we examine the effects of salinity and pH on the growth and lipid content of the algae. Samples of algae from the ponds exhibited high levels of growth and lipid production at a salinity of 9 ppt and pH 7. These conditions are similar to the natural conditions of the ponds, indicating that they may be a good source of algal biofuels. / text

The intertidal zone is finely partitioned in species distributions and abundances. The demographies of key species over varying spatial and temporal scales are fundamental to understanding the population structure and overall dynamics of habitats and assemblages. In this thesis, settlement, dispersal and early life stage survival and growth were examined in several habitat-forming intertidal fucoid algae in New Zealand and Oregon, U.S.A. Natural settlement patterns of Hormosira banksii, Cystophora torulosa and Cystophora scalaris were quantified for over three years at a semi-protected shore in southern New Zealand. Settlement was monitored in four tidal zones, within bare rock and algal habitats. Settlement was synchronous between tidal zones but the density of settlement varied spatially and temporally. There were significant differences between tidal zones, habitats and times of the year. For H. banksii, small pulses of settlement occurred year-round with greatest densities during spring and early summer. Greatest settlement occurred at low tidal zones and under adult canopies. Both Cystophora species also reproduced year-round, but had much lower settlement densities than H. banksii. Most settlement occurred during spring and summer, while only small pulses occurred in autumn and winter months. Most settlement was in the lowest tidal zone (0.4 m above chart datum), with only a few zygotes settling at higher shore zones. Canopy cover had no significant effect on settlement densities. Dispersal was examined in Durvillaea spp., H. banksii, C. torulosa and Fucus gardneri. For all species, settlement densities declined with distance from the source populations, but densities were variable between species. Durvillaea spp. dispersal was more extensive than expected, with significant settlement occurring 32 m from the source population, the maximum sample range of the study. However, settlement densities were much higher within 8 m from the source. The extensive dispersal of Durvillaea spp. is a result of the combination of small, slowly sinking eggs and the presence of buoyant mucilage. The other species studied showed far more restrictive dispersal, and much lower settlement densities. Settlement occurred 2 m from the source, but most settlement occurred under or near the canopy. The eggs of these species are much larger and sink faster than the eggs of Durvillaea spp. The consequences of settling at different shore heights and seasons were examined in H. banksii and D. antarctica in New Zealand, and F. gardneri and Pelvetiopsis limitata in Oregon. Transplant experiments tested the effects of grazing and heat/desiccation stress on survival and growth of germlings at different shore heights, during different seasons. High germling mortality was a feature of all species, but rate of mortality depended on conditions and species. There is a trade-off for settling at different times of the year; overall, growth was faster in warmer seasons, but survival was better in cooler seasons. During cooler seasons, germlings are exposed to less heat/desiccation stress, but their slow growth exposes them to grazing and competitive interactions for longer periods. For New Zealand species, shore height had large effects, with better survival and growth in the low shore. Grazers were very effective in the low shore, and heat/desiccation stress had strong effects in the mid and high shores. For Oregon species, effects of grazing and heat/desiccation stress were generally weaker than for New Zealand species. Shore height had weak effects, but ultimately low shore germlings had poor survival, primarily because of overgrowth by ephemeral algae. This is in contrast to the generalisation that survival and growth in the low shore should be better due to a more benign environment. In this study, species had specific demographies that related to their life history characteristics and responses to the local environment. Differences in settlement, dispersal abilities, survival and growth over small spatial and temporal scales clearly underpinned large scale differences in recruitment and adult distribution and abundances.