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Ammonia excretion from the freshwater fish gill is thought to occur mainly via passive diffusion of NH3 aided by a favourable plasma-to-water ammonia gradient sustained by a pH gradient formed by an acidified gill boundary layer. Rhesus (Rh) proteins are the newest members of the ammonia transporter superfamily. In this thesis research, ten rainbow trout Rh cDNA sequences were cloned and characterized. Rhcg2 mRNA and H+-ATPase mRNA and activity levels were upregulated in the trout gill pavement cells in response to experimentally elevated plasma ammonia, concurrent with enhanced ammonia excretion. Controversially, Rh proteins are thought to transport C02. However, Rh mRNA levels in most tissues of hypercapnia-exposed trout remained stable suggesting that trout Rh proteins likely do not conduct C02. Xenopus oocytes expressing trout Rh proteins facilitated the bi-directional transport of methylamine, an ammonia analogue. Methylamine transport was inhibited by ammonia and sensitive to a pH gradient and the concentration of the protonated species. Use of the scanning ion electrode technique (SIET) indicated that trout Rh proteins have an ammonia affinity within the physiological range, which is greater than that for methylamine, and they transport ammonia more rapidly than methylamine. A model of ammonia excretion in the trout gill pavement cell is proposed wherein ammonia enters via basolateral Rhbg and exits via apical Rhcg2, binding to these channels as NH4+ but transiting as NH3. In the gill boundary layer, NH3 combines with an H+ ion released from H+-ATPase and/or Na+/H+ exchange, forming NH4+. As low-affinity, high-capacity ammonia transporters, Rh proteins in the trout gill would exploit the favourable pH gradient formed by the acidic boundary layer to facilitate rapid ammonia efflux when plasma ammonia levels are elevated. Basal plasma ammonia levels are likely maintained by simple passive NH3 diffusion with a smaller role for Rh proteins under these conditions. / Thesis / Doctor of Philosophy (PhD)

Blooms of the toxic dinoflagellate Karenia brevis are common on the West Florida Shelf (WFS), yet little is known of the relationships between zooplankton and K. brevis. A comprehensive analysis was undertaken to examine 1) perturbations in zooplankton community composition within K. brevis blooms 2) the contribution of zooplankton ammonium and phosphate excretion to K. brevis bloom nutrient requirements, and 3) the role of zooplankton grazing in K. brevis bloom termination. Prior to undertaking the first portion of the study, an examination of the perturbations in the normal zooplankton assemblage within K. brevis blooms, it was first necessary to define the normal zooplankton assemblage on the WFS. To this end, a seasonal analysis of abundance, biomass and community composition of zooplankton was undertaken at 6 stations on the WFS. Monthly sampling was conducted for one year at the 5, 25 and 50- m isobaths. Two major groups in community composition were observed at the near shore (5-m and 25-m) and offshore (50-m) stations. Considerable overlap was seen in community composition between the 5-m to 25-m and 25-m to 50-m isobaths, but little overlap in community composition was observed between the 5-m and 50-m isobaths. Of the 95 species identified, only 25 proved to be important (>90%) contributors to community composition. Near shore, important contributors were Parvocalanus crassirostris, Penilia avirostris, Paracalanus quasimodo, Oithona colcarva, Oikopleura dioica, Centropages velificatus and Pelecypod larvae. As distance offshore increased, important contributors to community composition were Euchonchoichiea chierchiae, Clausocalanus furcatus, Oithona plumifera, Oithona frigida, Oncaea mediteranea, Calaocalanus pavoninius, Oithona similis, and Gastropod larvae. Variations in abundance and biomass between non-bloom and bloom assemblages were evident, including the reduction in abundance of 3 key species within K. brevis blooms. One potential source of nutrients to support K. brevis blooms may be zooplankton regeneration of nutrients. To test this hypothesis, ammonium and phosphate excretion rates of several West Florida Shelf copepods (Labidocera aestiva, Acartia tonsa, Temora turbinata, and Paracalanus quasimodo) were measured and prorated to a 24-hour day. These excretion rates were then extrapolated to other West Florida Shelf zooplankton, combined with available literature excretion rates for some taxa, and applied to zooplankton abundances found for K. brevis blooms on the West Florida Shelf in 1999 and 2001. Ammonium excretion rates were found to be inadequate to support all but 104 cells l-1 of K. brevis, though phosphate excretion rates were adequate to support even 106 cells l-1 of K. brevis. Grazing assessment was conducted for three common zooplankton species that were found within two K. brevis blooms, A. tonsa, P. quasimodo, and L. aestiva, using 14C labeled K. brevis cells. Grazing rates were then applied to the zooplankton community and grazing assessed. Grazing pressure was occasionally heavy, and was capable of reducing K. brevis to background concentrations at stations in the 1999 bloom and at 1 station in the 2001 bloom. Generally, however, grazing pressure proved to be insufficient to reduce K. brevis to background concentrations during the 1999 and 2001 blooms.

Aims of this thesis were to assesed the impact of feeding, temperature of water and fish size on ammonia excretion and oxygen consumption in Clarias gariepinus in recirculating system. Fish were divided in 8 weight categories from 21 to 2495 g. Experiments were performed under four temperatures: 22°C, 25°C, 28°C and 30°C. Fish were acclimatized, light conditions were: 14 hours of light period and 10 hours of dark period. Feeding was provided fed four times a day. Starved fish were measured. Measurements were held every two hours. Oxygen consumption was measured by multimeter and ammonia excretion using Nessler?s method. Average daily oxygen consumption in fed fish was dependent on temperature and average individual weight of fish, varied from 10.5 to 96.6 mg O2/kg/h and in hungry fish from 4.3 to 61.8 mg O2/kg/h. TAN excretion varied from 0 to 59.9 mg TAN/kg/h in fed fish and from 0 to 60.8 mg TAN/kg/h in hungry fish. Oxygen consumption increased with increasing temperature. Ammonia excretion was very unstable at all temperatures in all weight categories. Fish had higher ammonia excretion in light period of experiment.

Algumas atividades antrópicas podem provocar alterações nos recursos naturais, seja ela no rio, no solo ou no mar. Os efeitos destas alterações podem ser observados na biota, na qualidade das águas, na disponibilidade de nutrientes, interferindo em todos os elementos que compõem o ecossistema, influenciando-o em maior ou menor grau. O presente estudo procurou determinar o efeito dos poluentes sulfato de zinco, cloreto de amônia, dodecilsulfato de sódio e benzeno, numa espécie bioindicadora marinha, el larvas de mexilhão (Perna perna), além de acrescentar dados sobre a biologia e analisar quimicamente a presença de hidrocarbonetos nos tecidos dos animais adultos e das larvas. Para tanto foram utilizados testes de toxicidade, estudos sobre a bioenergética (consumo de oxigênio e excreção de amônia), enzimas biomarcadoras e excreção de fósforo e nitrogênio. Com relação à biologia, foram encontrados indivíduos sexualmente maduros a partir de 26,1 e 27 mm de comprimento para fêmeas e machos, respectivamente, e há diferença, ainda que esta não seja estatisticamente significativa, entre ovócitos e larvas submetidos a ação de poluentes. A análise de hidrocarbonetos indicou que existe uma contribuição biogênica e petrogênica no litoral norte do Estado de São Paulo. Para as demais análises verificou-se que, quando comparados aos grupos controles, as larvas foram sensíveis e responderam aos diferentes poluentes, geralmente com inibição da atividade. / Some antropic activities may be the cause fo alterations in natural resources, being it river, soil or sea. These alteration effects may be observed in the biota, water quality, nutrients disponibility, interfering with all elements that are part of a ecosystem, with greater or lesser influence degree. The present study was directed to determine the effect of pollutants, such as zinc sulphate, ammonia chlorate, sodium dodecilsulphate and benzene, acting over larvae of a marine bioindicator, the mussel (Perna perna), besides adding data on biology and chemically analysing hidrocarbon presence in larvae and adult animal tissues. In order to obtain these results, toxicity tests were used and bioenergetic (oxygen consumption and ammonia excretion), biomarkers, phosphorus and nitrogen excretion, were studied. As for biology, individuals sexually mature were found starting with 26,1 and 27 mm length females and males, respectively, and there is difference between larvae and ovocites, although statistically not significant, submitted to pollutants action. The hydrocarbon analysis indicates a biogenic and petrogenic contribution to north coast of São Paulo State. For the other analyses the observed results, when compared to control groups, showed that larvae are sensitive, and responded to different pollutants, generally with activity inhibition.

Algumas atividades antrópicas podem provocar alterações nos recursos naturais, seja ela no rio, no solo ou no mar. Os efeitos destas alterações podem ser observados na biota, na qualidade das águas, na disponibilidade de nutrientes, interferindo em todos os elementos que compõem o ecossistema, influenciando-o em maior ou menor grau. O presente estudo procurou determinar o efeito dos poluentes sulfato de zinco, cloreto de amônia, dodecilsulfato de sódio e benzeno, numa espécie bioindicadora marinha, el larvas de mexilhão (Perna perna), além de acrescentar dados sobre a biologia e analisar quimicamente a presença de hidrocarbonetos nos tecidos dos animais adultos e das larvas. Para tanto foram utilizados testes de toxicidade, estudos sobre a bioenergética (consumo de oxigênio e excreção de amônia), enzimas biomarcadoras e excreção de fósforo e nitrogênio. Com relação à biologia, foram encontrados indivíduos sexualmente maduros a partir de 26,1 e 27 mm de comprimento para fêmeas e machos, respectivamente, e há diferença, ainda que esta não seja estatisticamente significativa, entre ovócitos e larvas submetidos a ação de poluentes. A análise de hidrocarbonetos indicou que existe uma contribuição biogênica e petrogênica no litoral norte do Estado de São Paulo. Para as demais análises verificou-se que, quando comparados aos grupos controles, as larvas foram sensíveis e responderam aos diferentes poluentes, geralmente com inibição da atividade. / Some antropic activities may be the cause fo alterations in natural resources, being it river, soil or sea. These alteration effects may be observed in the biota, water quality, nutrients disponibility, interfering with all elements that are part of a ecosystem, with greater or lesser influence degree. The present study was directed to determine the effect of pollutants, such as zinc sulphate, ammonia chlorate, sodium dodecilsulphate and benzene, acting over larvae of a marine bioindicator, the mussel (Perna perna), besides adding data on biology and chemically analysing hidrocarbon presence in larvae and adult animal tissues. In order to obtain these results, toxicity tests were used and bioenergetic (oxygen consumption and ammonia excretion), biomarkers, phosphorus and nitrogen excretion, were studied. As for biology, individuals sexually mature were found starting with 26,1 and 27 mm length females and males, respectively, and there is difference between larvae and ovocites, although statistically not significant, submitted to pollutants action. The hydrocarbon analysis indicates a biogenic and petrogenic contribution to north coast of São Paulo State. For the other analyses the observed results, when compared to control groups, showed that larvae are sensitive, and responded to different pollutants, generally with activity inhibition.