Dindia, Laura Alexandria
Corticosteroids are key regulatory hormones involved in many aspects of physiology and have long been known to exert rapid and delayed effects. The delayed corticosteroid effects are mediated by transcriptional events downstream of glucocorticoid receptor activation. Conversely, the rapid effects are mediated independently of transcriptional regulation and are thought to involve non-classical steroid receptors and signaling pathways. Lately, research has begun to focus on delineating the rapid and nongenomic actions of glucocorticoids but most of these studies have been on mammalian models. Cortisol, the primary corticosteroid in teleosts, is an established genomic regulator of the physiological response to stress, but very little is known about either the rapid effects or their mechanisms of action in non-mammalian vertebrates. Additionally, nongenomic glucocorticoid action in the liver is poorly characterized in all animal species despite the importance of this organ in regulating glucocorticoid-mediated physiological adjustments during stress adaptation. The primary objective of this thesis is to investigate the rapid glucocorticoid effects, and their mode of action, associated with stressor-induced corticosteroid elevation in fish liver. The overriding hypothesis is that rapid effects of cortisol on acute stress adaptation involve changes to liver membrane order and rapid modulation of stress signaling pathways in rainbow trout (Oncorhynchus mykiss), a well studied teleost model. This hypothesis was tested by examining rapid plasma membrane and intracellular responses following stressor-induced cortisol elevations in vivo, as well as to cortisol treatment in vitro using liver plasma membrane, tissue slices, hepatocytes in suspension, or primary culture of hepatocytes. Steroid hormones are lipophilic molecules and freely incorporate into the plasma membrane. Through noncovalent interactions (hydrogen bonds and Van der Waal forces), glucocorticoids can potentially alter physical properties of the plasma membrane, thus leading to intracellular responses. The effect of stressor-induced cortisol elevations on physical changes to the hepatic plasma membrane was investigated by measuring the microviscosity of the plasma membrane. Plasma membrane fluidity (inverse of microviscosity) is an important determinant of transmembrane protein function, and changes to lipid order can transmit extracellular signals by activating membrane-associated signaling pathways. Fluidity of purified liver plasma membranes was monitored using steady-state fluorescence polarization of 1,6-diphenyl-1,3,5 hexatriene, a well characterized membrane probe. In addition to measuring lipid dynamics, the effect of cortisol on plasma membrane structure and surface properties were also investigated using atomic force microscopy. The effect of cortisol on the activation of key stress signaling pathways, including protein kinase (PKA), protein kinase (PKC), Akt, and mitogen-activated protein kinase (MAPK), was tested in fish liver. Also, as acute stress adaptation is regulated by an integrative hormonal response involving catecholamines (primarily epinephrine), the rapid effect of cortisol action on adrenergic signaling in the liver was evaluated in vitro. Finally, an attempt was made to identify cortisol-binding plasma membrane protein, as glucocorticoids are also thought to mediate rapid effects through a novel membrane glucocorticoid receptor. The results demonstrate for the first time that stressor exposure significantly increases liver plasma membrane fluidity (decreased microviscosity). A role for cortisol in mediating stressor-induced fluidization was confirmed in vitro, as physiological stress levels of this steroid (≥100 ng/ml) significantly increased liver plasma membrane fluidity. In addition to increasing lipid fluidity, acute stress and cortisol treatment altered membrane topography, including changes to membrane microdomains. The stressor-induced cortisol elevation also rapidly modulated major signaling cascades in rainbow trout liver, including PKA, PKC, and ERK1/2 MAPKs. A role for cortisol in the activation of these kinase pathways was confirmed in vitro. Specifically, cortisol rapidly and transiently increases cyclic AMP (cAMP) accumulation and induces the phosphorylation of PKA substrate proteins, including cAMP response element-binding (CREB) protein. In addition to activating PKA signaling, cortisol rapidly induced phosphorylation of PKC and Akt substrate proteins, while stimulating p38 MAPK dephosphorylation in vitro. Moreover, rapid cortisol signaling may stimulate metabolite oxidation in order to maintain the energy balance within liver tissue as cortisol acutely depleted key liver metabolites (including liver glucose), suggesting enhanced turnover without impacting the steady-state adenylate energy charge ratio (measure of the energy status of the cell). Also, rapid effects of cortisol alter the hormonal responsiveness of hepatocytes to adrenergic stimulation, including suppression of epinephrine-stimulated cAMP-CREB activation and glucose production. Preliminary results point to a plasma membrane protein that specifically binds cortisol in trout liver, but this remains to be characterized. Also, in addition the membrane-mediated response, mifepristone, a glucocorticoid receptor (GR)-antagonist, blocked some rapid cortisol effects suggesting the possible involvement of GR signaling pathway. Until now, cortisol has been primarily thought to play a role in the long-term recovery process to acute stress by enhancing plasma glucose levels through the upregulation of liver gluconeogenic capacity. The results presented in this thesis provide evidence for a novel role for rapid cortisol action on the acute metabolic adjustments that support liver function immediately following acute stressor exposure. Particularly, the results lead to the proposal that acute cortisol action stabilizes the energy status of the cell by maintaining ATP levels through increased metabolite turnover, suggesting enhanced metabolic activity of the liver immediately following acute stressor exposure. While the mechanism is unclear, plasma membrane alterations in response to cortisol intercalation may facilitate rapid steroid signaling either through mechanotransduction or by altering activity of plasma membrane proteins. The structural changes to the plasma membrane in response to acute stressor exposure and/or cortisol treatment highlight a novel membrane-mediated mechanism of rapid stress adaptation in hepatic tissue.
01 April 2013
Bacteria living within the cells of eukaryotic organisms can have profound effects on their hosts. One example is host sex-ratio distortion caused by bacterial endosymbionts, which can be induced through the killing of male progeny during their development. The mechanisms that underlie how bacteria can cause male death are poorly understood. Several previous studies suggested that the neural, and perhaps other, tissues are targeted by the male-killing bacterium, Spiroplasma.In this study, I tested this hypothesis, addressing whether tissues were specifically altered during male development and also when the initial defects manifested. These questions were addressed by analyzing developing tissues ofinfected and uninfected embryos with immunological reagents and confocal microscopy. My analyses revealed that Spiroplasma causes severe cellular defects in the male central nervous system (CNS) during mid embryogenesis. Specific CNS defects include disrupted neuronal packing as they differentiate and highly abnormal axon formation, with both phenotypes occurring at 6 hours and worsening throughout embryo development. Additionally, my analyses showed that during the onset of these CNS defects, other tissue types appeared normal. However, during late embryonic stages, male embryos became highly abnormal in tissue morphology across the whole animal. These observations suggest that Spiroplasma initially targets the CNS, which may, in turn, lead to secondary widespread defects in male embryos. I propose important experiments focusing on analysis of neuronal precursors to further explore the cellular basis of male killing, and I offer a model for how male CNS tissue can be targeted specifically at the molecular level.
The role of ACC deaminase in plant growth promotion by the endophytic bacterium Burkholderia phytofirmans PsJNSun, Yili 25 July 2008 (has links)
The endophytic bacterium Burkholderia phytofirmans PsJN has been previously shown to promote plant growth. This bacterium produces siderophores, indoleacetic acid (IAA) and the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, all of which have previously been implicated in the promotion of plant growth by bacteria. Following isolation of the ACC deaminase gene (acdS), AcdS deficient mutants of PsJN were generated. One mutant contains a tetracycline resistance gene inserted into acdS, and the other mutant contains a deletion in the acdS gene. Both of the mutants showed no detectable ACC deaminase activity, produced a decreased level of siderophores and an increased amount of IAA compared to the wild-type, and lost the ability to promote canola root elongation. In addition, the GFP-labeled acdS deletion mutant colonized plant interior surfaces somewhat less efficiently than the GFP-labeled wild-type strain.
Analysis of the structural determinants for voltage-dependent G protein modulation of synaptic Cav2 channelsHuang, Xuan Jay January 2008 (has links)
Specialized voltage-gated calcium channels in the Cav2 channel class (such as Cav2.2, N-type) mediate neurotransmitter release from presynaptic nerve terminals. Cav2.2 channels are exquisitely sensitive to inhibition by G protein-coupled receptors. The ubiquitous form of G protein modulation is a fast, membrane delimited, voltage-dependent form of regulation, which is relieved by strong depolarizations. LCav2, an invertebrate homolog from the pulmonate snail Lymnaea stagnalis, serves a similar function as a mediator of transmitter release in the nervous system. To examine the G protein modulation capacity in invertebrates, LCav2 was cloned to a bicistronic expression vector pIRES2-EGFP and expressed in HEK293T cells. Although LCav2 was almost indistinguishable from mammalian Cav2.2 in biophysical characteristics observed in vitro, snail LCav2 channel lacked the property of voltage dependent G protein modulation. The structural elements essential for the voltage sensitivity to G protein modulation were explored by swapping the N-terminus and I-II linker regions of rCav2.2 channels into LCav2 calcium channels. Functional comparisons were also made using both mammalian and invertebrate homologs of G protein beta subunits, Gβ1. Neither the N-terminus or I-II linker region of Cav2.2 alone, nor the invertebrate G protein beta subunit was sufficient for voltage-dependent G protein modulation. Further analyses using chimeric channels and G protein subunits will be required to find the minimal structural determinants for voltage-dependent G protein modulation.
Testate Amoebae as a Performance Indicator of Ecosystem Establishment in Wetlands Impacted By Oil Sands Processed MaterialsLegg, Allison 11 May 2009 (has links)
Mining for oil sands in the Athabasca Basin in northeastern Alberta is rapidly expanding. As economics continue to drive growing mining practices, waste management, reclamation and bio-monitoring strategies are becoming increasingly important. This project aims to determine the practicality of testate amoebae assemblages as an indicator of microbial community health and ecosystem establishment in wetlands impacted by oil sand processed materials (OSPM). Testate amoebae are unicellular, shelled protists that live in abundance in soils, leaf litter and in fresh water habitats. This group of protists forms shells (or tests) which makes them relatively easy to identify. Ecological studies have shown they occupy specific niches controlled by environmental parameters such as pH and moisture variables. These features make testate amoebae excellent bioindicators, and this project explores the potential of applying testate amoebae bioindicators in wetlands affected by OSPM. Using compound and epifluorescent microscopy techniques, testate amoebae species assemblages were identified and tabulated from a series of wetlands with different impacts by oil sands processed materials. Bacterial and fungal proportions were characterized to compare with the testate amoebae and identify possible links within the microbial community. A total of 44 species of testate amoebae were encountered in 24 wetlands, with Centropyxis platystoma and Centropyxis aculeata being the most common taxa. Natural peatland sites, not affected by OSPM contained the most diverse assemblage of testate amoebae containing Arcella, Assulina, Centropyxis, Englypha, and Heleopera. Open-water wetlands not impacted by OSPM were less diverse than peatland sites, but maintained between two and 12 taxa per site. Open-water sites amended with OSPM contained fewer taxa (between 0 and 4 taxa at any given site) and fewer individuals than any other site type, with Difflugia being most common. Bacteria contributed an average of 65% of the microbial community in open-water sites and an average of 80% in peatland sites. Peatland sites were significantly different (P<0.05) from all other site types in terms of testate amoebae, bacteria, and fungal biomass. This study demonstrated that differences exist in testate amoebae assemblages between site types do exist, this study establishes the fact that testate amoebae are too few (<1% of biomass), and not in sync with the other microbial facets studied (bacteria and fungi) limiting their use as bioindicators of microbial community establishment in wetlands impacted by oil sands processed materials. However the predominance of bacteria in all site types calls attention to their vital role in these sites and their importance in further research in oil sands reclamation.
The water-sediment interactions for Hyalella azteca exposed to uranium-spiked or contaminated sediments and different overlying water chemistriesAlves, Lara January 2009 (has links)
In comparison with other metals such as Cd, Cu, Pb, Ni, and Zn, little is known about uranium (U) toxicity to Hyalella azteca. There is even no national U water or sediment quality guideline yet for the protection of aquatic life in Canada, despite Canada being home to some of the biggest U producers in the world. In this context, the aim of this research was to determine the toxic effects of U concentrations in the water and sediment to H. azteca, and if these relationships can be modelled. This thesis demonstrated that U bioaccumulation was mainly via the water phase rather than the sediment phase. It showed that U bioaccumulation measurements in H. azteca were more reliable indicators of U toxicity than U concentrations in the water or sediment. A water-bioaccumulation saturation model was satisfactory at describing this relationship. Overlying water chemistry was found not only to influence U bioaccumulation and toxicity in the H. azteca but also the desorption of U in the sediment into the overlying water. A water-sediment partitioning saturation model was also satisfactory at explaining these interactions. Both body size and gut-content had an overall effect on U bioaccumulation in H. azteca exposed to water-only U concentrations in soft water. A saturation model was used not only to estimate the effect of gut-content on U bioaccumulation, but to predict the uptake and elimination rate constants for H. azteca exposed to water-only U concentrations. A field study was conducted to determine if the saturation models developed and applied in the laboratory could be used in the field to quantify U bioavailability, bioaccumulation and toxicity to H. azteca. Unfortunately, U concentrations in the water and sediment were below concentrations needed to validate these models. However, toxicity, not related to U concentrations in the field, was observed at some field sites. Overall this thesis not only encourages more work on U toxicity to H. azteca, but provides significant data and models to be used by risk assessors and regulators in the development of U water and sediment quality guidelines in the protection of aquatic environments in Canada.
Meyer, Christopher John
The thesis work concerns the multiseriate exodermis (MEX), an outermost cortical layer (two or more cell layers thick) characterized by Casparian bands and suberin lamellae. Diverse aspects of Iris germanica's MEX were examined including its maturation under differing growth conditions, and how this maturation affected water and solute permeability. Also, suberin metabolite profiles for the maturing MEX of I. germanica and the maturing uniseriate exodermis of Allium cepa were established. This multidisciplinary approach resulted in a comprehensive understanding of how anatomical and biochemical changes to the exodermis affect water and solute permeability of the MEX. Most previous studies of exodermal development have involved species with a uniseriate exodermis. To extend this work, the MEX in I. germanica roots was investigated. The outermost exodermal layer matured first with normal Casparian bands and suberin lamellae. But as subsequent layers matured, the Casparian band extended into the tangential and anticlinal walls of their cells. This atypical Casparian band was continuous around the root circumference. MEX maturation was influenced by the roots' growth medium. Plants were grown in soil or hydroponics (with and without a humid air gap), and their roots were sectioned and stained with various dyes to detect Casparian bands and suberin lamellae. In soil-grown roots, the exodermis started maturing (with concurrent deposition of Casparian bands and suberin lamellae) 10 mm from the root tip, and two layers had matured by 70 mm. In hydroponically grown roots, exodermal maturation was delayed. However, in basal regions exposed to an air gap in the hydroponic tank, maturation of the second exodermal layer was accelerated. Therefore, changes in growth conditions have striking effects on exodermal maturation in I. germanica. With respect to radial water and solute transport, I. germanica roots with a mature MEX had lower permeability rates compared with uniseriate exodermal roots or roots in which the endodermis represents the major transport barrier. Transport studies were conducted on completely submerged roots and air gap-exposed root regions using a pressure chamber whereby water permeability (Lppc) was measured quantitatively across the entire root. This instrument proved to be preferable because in large diameter roots (up to 2.5 mm in I. germanica), root hydraulics were affected by the large water storage capacity of the central cortex. Compared with regions of roots with no mature exodermal layers, the mature MEX reduced Lppc from 8.5 x 10-8 to 3.9 x 10-8 m s-1 MPa-1. Puncturing the MEX increased Lppc to 19 x 10-8 m s-1 MPa-1, indicating that the MEX is an important hydraulically resistant tissue. A root pressure probe was used to measure the permeability of roots to NaCl and ethanol; solute permeability was reduced in the presence of two mature MEX layers. The MEX of I. germanica should play an important role in survival under conditions of drought and salt stress. Suberin is a complex biopolymer with a poly(aliphatic) domain (SPAD) that, in the case of a suberin lamellae, is known to be located between the cell wall and plasma membrane. The location and lipophilic nature of the SPAD establishes it as a structure restrictive to radial water transport through the transcellular pathway. Synthesis of the SPAD in a maturing exodermis was not well understood. Hence, a suberin metabolite analysis during development was conducted on the maturing MEX of I. germanica grown in submerged and air gap hydroponic conditions. Suberin monomers of the soluble (unpolymerized) and insoluble (polymerized) fractions were chemically isolated, then quantified and identified by GC-MS. Interestingly, in air gap-exposed regions, there was an increased synthesis and deposition of insoluble SPAD monomers in the first two exodermal layers, compared with submerged regions. The SPAD fraction included fatty acids, α,ω-dioic acids, ω-OH fatty acids, and ferulic acid, with C18:1 α,ω-dioic acid and ω-OH fatty acid being the two most abundant monomers. Also, in tissue that matured in the air gap, the composition of the soluble fraction changed significantly among exodermal maturation stages and between growth conditions. Of particular significance, increased amounts of alkanes, the major component of waxes, accumulated in the first exodermal layer. Other monomers of the soluble fraction included fatty acids, fatty alcohols, and ferulic acid, that were SPAD biosynthetic precursors. It was postulated that the localized and abundant deposition of C18:1 α,ω-dioic acid and ω-OH fatty acid, along with high accumulation of intercalated waxes in the first mature exodermal layer, were more important than the overall number of suberized exodermal layers for reducing water loss from the root during drought. Lastly, hydroponically grown Allium cepa roots were used as models to analyze SPAD synthesis in a maturing uniseriate exodermis. Roots were divided into four maturation zones based on the growth rate and the deposition of suberin lamellae in maturing exodermal cells as determined by histochemical analyses. The chemical composition of the soluble fraction was essentially unchanged as the exodermis matured. In contrast, the SPAD composition differed during maturation, mainly due to significant increases in the deposition of C18:1 α,ω-dioic acids and C18:1 ω-OH fatty acids. It is proposed that the exodermal maturation zones with corresponding suberin metabolite profiles be used as targets for the functional enzymatic characterization of suberin biosynthetic pathways.
20 May 2010
Proteome-level changes of both the plant growth-promoting bacterium (PGPB) Pseudomonas putida UW4 and its plant host Brassica napus (canola) were profiled using two-dimensional (2-D) difference in-gel electrophoresis (DIGE) and mass spectrometry, to elucidate the proteins’ prospective of plant-bacterial interactions. This study was undertaken in an effort to elaborate how a plant growth-promoting bacterium and its plant host biochemically and physiologically influence one another. More specifically, the effects of the PGPB P. putida UW4 on the proteome of canola and vice versa were examined. In addition, environmental stresses including heavy metal and salt were incorporated into the system. Moreover, how the presence of a functional bacterial 1-aminocyclopropane-1-carboxylate (ACC) deaminase (AcdS), which can lower plant ethylene levels and hence promote plant growth under a variety of stresses, would affect protein expression in both the bacterium and the plant was investigated. First, 2-D DIGE was used to detect significantly up- or down- regulated proteins in P. putida UW4 and its AcdS minus mutant in response to the presence of 2 mM nickel. Thirty-five proteins whose expression was altered were successfully identified by mass spectrometry and sequence comparisons with related species. Nineteen of the identified proteins were detected as differentially expressed in both wild-type and AcdS minus mutant expression profiles. Functional assessment of proteins with significantly altered expression levels revealed several mechanisms involved in bacterial heavy metal detoxification, including general stress adaptation, anti-oxidative stress and heavy metal efflux proteins. In addition, by detection of bacterial protein expression changes in the presence of plant exudates, three unique P. putida UW4 proteins that mediate interactions between the bacterium and its plant host were identified. However many of the observed changes of protein expression elicited by nickel and plant exudates were similar for wild-type P. putida UW4 and the AcdS minus mutant, with the majority of identified significant protein expression changes occurring in both strains. This is not unexpected because the P. putida UW4 ACC deaminase is unlikely to be involved in bacterial perception and response to plant host signals and environmental stimuli, and it causes a noticeable difference only in plant growth. A comprehensive proteome 2-D reference map of the PGPB P. putida UW4 containing 326 2-D gel spots representing 275 different proteins was also constructed. A 2-D database containing all the mass spectrometric information of P. putida UW4 proteins has been constructed. The data set has been deposited into the World-2DPAGE database and is accessible at http://world-2dpage.expasy.org/repository/. On the plant side, ninety proteins with significantly altered expression levels in the presence of salt and/or bacteria were identified by mass spectrometry. Many of these proteins are involved in photosynthesis, anti-oxidative processes, salt transportation/accumulation and pathogenesis-related responses. Importantly, the presence of the bacterial ACC deaminase was observed to alter the plant’s protein expression in response to salt stress. The effects included enhanced photosynthesis and salt accumulation contributed by wild-type P. putida UW4. The work described in this thesis furthers our understanding of plant-bacterial interactions, and is also likely to be of importance to both organic agriculture and environmental remediation efforts.
Examination of curcumin-induced heat shock protein gene expression in Xenopus laevis A6 kidney epithelial cellsKhan, Saad January 2010 (has links)
The heat shock response is a cellular homeostatic mechanism that is activated in response to stressful stimuli (e.g. heat shock, heavy metals, disease states etc.), which causes an increase in unfolded protein, which triggers the expression of heat shock protein (hsp) genes. HSPs are molecular chaperones that assist in protein synthesis, folding and degradation and prevent stress-induced protein aggregation. Since stressor-induced tissue damage is associated with different disease states, indirect evidence suggested that HSP inducers may be therapeutically beneficial for certain diseases. Curcumin, a phenolic compound found in the Indian spice, Curcuma longa (Turmeric), was shown to have anti-inflammatory, anti-tumor and anti-amyloid properties. In the present study, it was determined that curcumin inhibited the ubiquitin-proteasome system (UPS) activity and induced the accumulation of HSPs in the frog model system, Xenopus laevis. Treatment of A6 kidney epithelial cells with curcumin enhanced ubiquitinated protein levels and inhibited chymotrypsin-like activity. Furthermore, HSP30 and HSP70 accumulation was observed in cells exposed to 10 - 50 μM curcumin for 24 h in a concentration-dependent manner with maximal levels of HSP30 and HSP70 in cells treated with 30 μM curcumin. Time-dependent increases in HSP30 and HSP70 accumulation were also observed in cells treated with 30 μM curcumin for 2 to 24 h. The accumulation of HSP30 and HSP70 in cells recovering from curcumin exposure increased up to 24 h after treatment. The simultaneous treatment of A6 cells with 10 μM curcumin and mild heat shock (30 ºC) for 6 h resulted in an enhanced accumulation of HSP30 and HSP70, which was greater than with each stressor alone. This pattern of combined stressor-induced HSP30 and HSP70 accumulation increased from 2 to 6 h, after which it decreased from 10 to 24 h. The activation of HSF1 may be involved in curcumin-induced hsp gene expression in A6 cells since KNK437, a heat shock factor-1 inhibitor, inhibited the accumulation of HSP30 and HSP70. Immunocytochemical analysis employing the use of laser scanning confocal microscopy (LSCM) revealed that curcumin-induced HSP30 was detectable primarily in the cytoplasm in a punctate pattern with minimal detrimental effects on the actin cytoskeleton. Elevation of the incubation temperature from 22 to 30 °C greatly enhanced the curcumin-induced cytoplasmic accumulation of HSP30 in a granular pattern. Lastly, curcumin treatment also conferred a state of thermotolerance in A6 cells such that they were able to maintain proper actin cytoskeleton in subsequent thermal challenges. This phenomenon was controlled at the transcriptional level since pretreatment of cells with KNK437, repressed HSP30 accumulation and cytoprotection. These findings are of importance given the interest in identifying agents that can upregulate HSP levels with minimal effects on cell structure or function as a therapeutic treatment of certain protein folding diseases.
Shallow urban impoundments are known for low water transparency, and generally low water quality. This thesis investigates the effects of land use, management regime, and carp density on the water quality in 4 shallow urban impoundments on the Laurel Creek system in Waterloo, Canada. The focus of this study is to look at how these factors affect suspended solids, total phosphorus, and phytoplankton assemblages within the impoundments. The land upstream of the impoundments is lightly to heavily urbanized. The impoundments differ in how they are managed as well, with the most upstream impoundment being completely drawn-down every year with drastic effects downstream. Clair and Silver Lakes received significantly higher total inorganic solids (TIS) concentrations compared to Laurel Creek Reservoir and Columbia Lake during the study period; however there were no significant differences between the impoundment inlets for total organic solids (TOS) or total phosphorus (TP) concentrations during the same period. Three of the four impoundments were found to export significantly higher concentrations of TIS; these were Columbia, Clair and Silver Lakes. Additionally, Clair and Silver Lakes exported significantly higher TIS concentrations than both Columbia Lake and Laurel Creek Reservoir. Laurel Creek Reservoir discharged very low TIS concentrations throughout the study period. The same pattern as TIS concentrations was seen for the TOS concentrations leaving the impoundments, with Columbia, Clair, and Silver Lakes having significantly higher outlet TOS concentrations when compared to Laurel Creek Reservoir; additionally Clair and Silver Lakes discharged significantly higher TOS concentrations when compared to Columbia Lake. There were no significant differences among TP concentrations leaving the impoundments during the study period. In general, Laurel Creek Reservoir and Silver Lake discharged the highest phytoplankton densities; while Columbia and Clair Lakes discharged lower densities; however, there was no significant difference in these discharge densities between the different impoundments.
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