The purpose of this study is to find out that if antibiotic streptomycin decreases neuronal network activity or affects the pharmacological responses. The experiments in this study were conducted via MEA (multi-electrode array) technology which records neuronal activity from devices that have multiple small electrodes, serve as neural interfaces connecting neurons to electronic circuitry. The result of this study shows that streptomycin lowered the spike production of neuronal network, and also, sensitization was seen when neuronal network pre-exposed to streptomycin.
Cannabinoid (CB) receptors have been found in most vertebrates that have been studied. The location of various CB receptors in the body and brain are known, but their physiological functions are not fully understood. The effects CBs have on the cardiovascular system have been of growing interest in recent years. Increasing reports from emergency departments and law enforcement agencies detail acute cardiovascular and psychological effects from synthetic CB intoxication, such as JWH-018. This major health concern is substantiated by governmental agencies like the CDC and NIDA. This pilot study investigates the acute toxic effects of the synthetic CB, JWH-018, on the cardiovascular and neuroendocrine systems in Ictalurus punctatus (channel catfish). Research in organisms besides the traditional mammal models can provide new insights into CB function and physiology. Ictalurus punctatus lend multiple benefits as a model organism that permits researchers to investigate in vivo effects of both cardiovascular and neuroendocrine systems without much influence from traditional sampling methods, and further more provide ample size and tissue to perform specific cardiovascular experiments. Multiple methods were used to assess cardiovascular function and sympathetic nervous system activation. Two different doses, low (500 µg/kg) and high 1,500 µg/kg, of JWH-018 were evaluated in the study. Delivery of JWH-018, via dorsal aorta cannulation, was administered to channel catfish in order to measure cardiovascular functions and sample blood. Plasma levels of the hypothalamus-pituitary-adrenal/interrenal (HPA/I) biomarkers; ACTH, cortisol, epinephrine, and norepinephrine, were measured using ELISAs. Myocardial and neural tissue was collected after the exposures for rt-PCR analysis on β2 adrenergic and glucocorticoid receptor density change. Acute exposure of JWH-018 in undisturbed channel catfish yielded several findings: (1) High dose of JWH-018 was responsible for cardio depressor effects in catfish with a tendency to produce tachycardia, (2) rt-PCR results showed a 2.7 fold increase of glucocorticoid receptor mRNA density …
Alpine zooplankton are exposed to a variety of stressors in their natural environment including ultraviolet radiation. Physiological coping mechanisms such as the accumulation of photoprotective compounds provide these zooplankton protection from many of these stressors. Elevated levels of carotenoid compounds such as astaxanthin have been shown to help zooplankton survive longer when exposed to ultraviolet radiation presumably due to the strong antioxidant properties of carotenoid compounds. This antioxidant capacity is important because it may ameliorate natural and anthropogenic stressor-induced oxidative stress. While previous researchers have shown carotenoid compounds impart increased resistance to ultraviolet radiation in populations of zooplankton, little work has focused on the toxicological implications of PAH induced phototoxicity on zooplankton containing high levels of carotenoid compounds. This thesis discusses research studying the role that carotenoid compounds play in reducing PAH induced phototoxicity. By sampling different lakes at elevations ranging from 9,500' to 12,700' in the front range of the Colorado Rocky Mountains, copepod populations containing different levels of carotenoid compounds were obtained. These populations were then challenged with fluoranthene and ultraviolet radiation. Results discussed include differences in survival and levels of lipid peroxidation among populations exhibiting different levels of carotenoid compounds, and the toxicological and ecological implications of these results.
Investigations of cholinergic and adrenergic tone on heart rate (fH) and mean arterial pressure (Pm) during embryonic development have been conducted on numerous avian species. While these investigations have documented that adrenergic tone, a continuous stimulation, on fH and Pm is vital to embryonic development in the birds studied to date, development of cholinergic tone on fH has been shown to vary even within species. Further, past studies have been bias to focus primarily on precocial species while altricial species remain poorly understood in this context. The goal of this investigation was to investigate the role of cholinergic and adrenergic tone on fH and Pm of an altricial species, the neotropic cormorant (P. brasilianus) to address this bias. The embryonic neotropic cormorant possesses B-and-a adrenergic tone on fH and Pm at 70% and 90% incubation while cholinergic tone on fH occurs at 90% incubation. This pattern of control is similar to that previously reported for several species of precocial birds suggesting the development of tonic cardiovascular regulation may be conserved across avian taxa.
Oxygen availability is critical for survival for most organisms. The nematode, C. elegans, has been useful for studying genetic regulation of anoxia tolerance due to the oxygen deprivation response mechanisms shared with other metazoans. Studies examining long-term anoxia (72h, LTA) tolerance have only been conducted at adult day 1. To investigate the effect of aging on anoxia tolerance wild-type and mutant strains were exposed to LTA between adult day 1 and day 9. Wild-type isolates and daf-16(mu86) (FOXO transcription factor regulated by insulin-signaling) and aak-2(gt33) (catalytic subunit of AMP-activated protein kinase) strains were anoxia sensitive at day 1 and displayed increased LTA tolerance with aging correlated with reproductive senescence followed by a decline in survivorhsip through day 9. The daf-2(e1370) (insulin receptor homologue of C. elegans), glp-1(e2141) (a lin-12/Notch receptor) and fog-2(q71) (required for spermatogenesis) strains were LTA-tolerant through day 5. I conclude that aging influences LTA-tolerance in a strain- and age-dependent manner. In addition to being LTA-tolerant the daf-2(e1370) and glp-1(e2141) strains have a longevity phenotype that is suppressed by loss of kri-1 or daf-12. While loss of kri-1 did not suppress the LTA-tolerant phenotype of glp-1(e2141) at day 1 the portion of impaired survivors increased at day 3 and by day 5 tolerance was suppressed. Similarly, when exposed to 4 days of anoxia the glp-1(e2141);daf-12(rh41rh611) double mutant had a reduced survivor rate at all ages analyzed compared to glp-1(e2141) controls. To better understand formation of an anoxia-tolerant physiology I exposed adults to one or more 24h bouts. Recurrent bouts increased LTA tolerance in wild-type hermaphrodites in a dose-dependent manner. Bout-treated daf-16(mu86) animals had increased survival rate compared to controls yet maximum survival remained below age-matched wild-type. Anoxia bouts decreased LTA-tolerance in aak-2(gt33) mutants, indicating the requirement for ATP regulation in establishing an LTA-tolerant phenotype. These data support the …
N-Acylethanolamines (NAEs) are a class of bioactive lipids, and FAAH is one of the enzymes responsible for degrading NAEs in both plants and animals. in plants, FAAH appears to be closely associated with ABA, a phytohormone which has long been associated with plant stress responses, since the overexpression of FAAH in Arabidopsis results in ABA hypersensitivity. Therefore, it is reasonable to speculate that alterations in FAAH transcript levels will result in altered stress responses in plants. to investigate this hypothesis experiments were carried out in which wild type (WT), FAAH-overexpressing (OE), and T-DNA insertional FAAH knockouts of Arabidopsis (faah) were grown in MS media under stress conditions. the stress conditions tested included chilling stress, heavy metal stress induced by cadmium or copper, nutrient limitations induced by low phosphorus or low nitrogen, salt stress induced with NaCl, and osmotic stress induced with mannitol. the OE plants were consistently hypersensitive to all stress conditions in relation to wild type plants. Inactive FAAH overexpressors did not have the hypersensitivity to the salt and osmotic stress of the active OE plants and were instead tolerant to these stresses. FAAH2 (faah2) knockouts and FAAH 1 and 2 double knockouts (faah 1+2) were based on some root development parameters somewhat more tolerant than WT plants, but more sensitive in terms of shoot growth. Collectively the data suggests that FAAH activity may interact with stress-responsive pathways in plants, perhaps including pathways involving ABA.
Urban land use and land cover has changed in the USA, giving rise to the American lawn – manicured, resource-intensive, and non-native. Green infrastructure design has been suggested in the literature as a potential alternative to the American lawn when managed as native xeriscapes, which require little to no irrigation after establishment. Given the influence of public preference on landscaping decisions, what is the relationship between the perceived value and ecological benefits of the American lawn compared to such alternatives? Few studies have explored this question in addition to the effects of college courses on influencing student preferences, as future stakeholders, towards native xeriscapes as alternatives to the American lawn. This research measured the effects of an introductory environmental education (EE) course on measurably influencing undergraduate student preferences for four xeriscapes as alternatives to the American lawn. To measure these effects, this study utilized the perceptions of 488 students enrolled in an indirect introductory EE course and 131 students enrolled in an introductory non-EE course. Three key results emerged from this research. Students preferred the American lawn more than xeriscape alternatives, irrespective to course enrolled. Introductory non-EE did not have an effect on student preferences, whereas indirect introductory EE did show some effects on student preferences. Lastly, student preferences were negatively associated with NPP per photosynthetically active square meter. The data from this study suggests that indirect introductory EE does not shift aesthetic landscape preferences towards pro-environmental alternatives. These results show promise for shifting such preferences via more direct EE approaches.
Eukaryotic platelets are small cell fragments that are released into the bloodstream from megakaryocytes, and their production is initiated in the bone marrow. They are mainly involved in blood hemostasis and thrombus formation. The newly synthesized platelets are called reticulated platelets or young platelets. Zebrafish thrombocytes are equivalent to mammalian platelets and have similar characteristics and functions. Likewise, zebrafish has both young and mature thrombocytes. Only young thrombocytes as reticulated platelets are labeled with thiazole orange. Similarly, labeling zebrafish thrombocytes with a specific concentration of DiI-C18 showed two populations of thrombocytes (DiI+ and DiI-). Again, only young thrombocytes showed DiI+ labeling. The mechanism of selective labeling of young thrombocytes by is unknown. Furthermore, there is no zebrafish line where young and mature thrombocytes are differentially labeled with fluorescence proteins. Therefore, in this study, we identified and confirmed that the RFP labeled cells of Glofish were young thrombocytes. In addition, we found that myosin light chain 2 (MLC2) promoter is expressed in young thrombocytes. We also generated a transgenic zebrafish line, GloFli fish, where the young and mature thrombocytes are labeled with red and green fluorescence proteins respectively. Furthermore, this study showed a two-fold increase in glycerol-phospholipids (GP) in mature thrombocytes compared to young thrombocytes suggesting the lipid composition may be important for differential labeling. Therefore, we tested the liposomes prepared with different ratios of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) and observed that the lower amounts of PE favor the DiI-C18 labeling whereas higher concentrations of PC are less efficient. Also, in both PE and PC, increased concentrations of both resulted in decreased binding. These results are consistent with our observation that mature thrombocytes have higher concentrations GP and thus DiI-C18 may not bind to them efficiently compared to young thrombocytes.
This study examined the impact of the National Science Foundation's Research Experience for Teachers (RET) in engineering at University of North Texas on students after their teachers' participation in the program. Students were evaluated in terms of self-efficacy, knowledge of engineering, perceptions of engineering, and interest in engineering. A 22-item Likert pre/post survey was used for analysis, and participants included 589 students from six high schools, one middle school, and one magnet school. Paired surveys were analyzed to determine if there was a statistically significant difference in attitudes and knowledge after teachers implemented lessons from their time at the RET. Surveys were also analyzed to determine if there was a statistically significant difference in student response based on gender or student school type. Results showed no statistically significant difference in the self-efficacy of students, however there was a statistically significant difference in knowledge, perceptions, and interest in engineering. In addition, there was a statistically significant difference between genders on an isolated question, and seven out of the 22 Likert questions showed a statistically significant difference between student school types.
C. halicacabum (Sapindaceae) is an invasive plant that is considered a nuisance species in Texas riparian environments. Little is known of the tolerance of C. halicacabum to flooding and drought; however, this information may provide insight into the characteristics that contribute to C. halicacabum purported invasiveness. C. halicacabum seedlings (n = 92) were exposed to one of four levels of water availability (flooded, saturated, intermediate and dry) over six weeks under greenhouse conditions. Plant performance was affected by water availability; however, there was no effect on survivorship. Flooded and saturated plants exhibited morphological adaptations; producing adventitious roots, hypertrophy, and aerenchyma tissue. Morphological measures, anatomical responses, and patterns of biomass allocation all indicate that C. halicacabum is able to survive periodic inundation, perform in saturation, and establish and thrive on the drier end of a moisture gradient.
The purpose of my research work was to assess a variety of Northern Thailand plants tissue extracts for antibacterial and anti-platelet aggregation activity. The Minimum Inhibitory Concentration assay method was used to assess antimicrobial activity of plant extracts, while the Zebrafish Platelet Aggregation Assay and the in vitro Whole Human Blood Impedance Aggregation Assay were used to study anti-platelet activity. Forty one plant extracts harvested from the tissues of 26 plants collected from Northern Thailand were assessed. Thirty-four plant extracts were found to have antibacterial activity against the Gram positive bacteria Staphylococcus aureus and/or Bacillus subtilis, while six plant extracts demonstrated activity against the Gram negative bacterium, Escherichia coli. Thirteen plant extracts exhibited anti-platelet aggregation activity better than the positive control. Two crude plant extracts, twigs from Garcinia sp. and twigs from Goniothalamus chilensis were selected for fractionation. Five of the 12 fractions showed anti-platelet activity. Four fractions (two from each plant extract) were selected for further sub-fractionation. Fourteen of 35 sub-fractions were selected for further testing of anti-platelet aggregation activity with 12 sub-fractions demonstrating positive antiplatelet activity. Positive sub-fractions were analyzed using liquid chromatography-mass spectrometry to determine their chemical properties. Three compounds that possessed anti-platelet activity were characterized by nuclear magnetic resonance. The compounds, all isolated from Garcinia sp., were identified as cambogin, isoxanthochymol and guttiferone F.
The goal of this study was to build on the historic use of the avian model of development and also to further the knowledge of autonomic nervous system (ANS) regulation of cardiovascular function in vertebrates. Vasoactive drugs sodium nitroprusside, a vasodilator and phenylephrine, a vasoconstrictor were used to study the correlation of cardiovascular function relationship with nerve activity, both sympathetic and parasympathetic (vagal). Additionally, ANG II was used to assess its effects on vagal inhibition. The present study shows that pharmacologically-induced hypertension is associated with a fall in mSNA, indicating that the capacity for sympathetic autonomic cardiovascular regulation is established by late incubation however, late-stage embryonic chickens did not show a significant increase in mSNA during hypotension. The hypotensive response of the embryo was not accompanied by the expected inhibition of vagal discharge; however a slight but insignificant reduction in vagal discharge was noted. When vagal efferent output was isolated, a significant drop in vagal efferent activity was noted in response to hypotension. The present study showed late-stage embryonic chickens lack a vagal response to hypertension in both efferent and sensory limbs. In this study, vagal discharge was reduced from baseline levels in response to Ang II. Collectively, the present study indicates that the lack of a decreased heart rate, in response to increases in Pm caused by Ang II, is due to a central inhibitory action of Ang II on the vagus. Data from the present study suggests that although autonomic interaction with the cardiovascular system in present in late-stage chicken embryos, it is still underdeveloped and possesses a limited capacity.
The Cape Horn Biosphere Reserve, within the remote Sub-Antarctic ecoregion is a reservoir of expressions of biological and cultural diversity. Although it is considered one of 24 wilderness areas remaining in the world, it is not free from local and global threats, such as invasive species, and climate change. Field biologists and philosophers associated to the Sub-Antarctic Biocultural Conservation Program and the Omora Ethnobotanical Park, have worked to describe the region’s biocultural diversity, linking ecological and philosophical research into education, ecotourism, and conservation, through a methodology called field environmental philosophy (FEP), which integrates ecological sciences and environmental ethics through a 4-step cycle consisting of: 1) interdisciplinary research; 2) composition of metaphors; 3) design of field activities with an ecological and ethical orientation; and 4) implementation of in situ conservation areas. In this context, the purposes of this dissertation were to: 1) provide a comprehensive review of publications regarding the conservation status of aquatic and terrestrial insects at a global scale and with an emphasis in southern South America; 2) study the distribution of benthic macroinvertebrates through the sharp altitudinal gradient of the Róbalo River watershed; 3) describe the life histories of Gigantodax sp (Simuliidae: Diptera) and Meridialaris chiloeense (Leptophlebiidae: Ephemeroptera) in the Róbalo River and to assess the potential effects of climate change on their phenology; and 4) to apply FEP methodology in order to better understand and communicate the intrinsic and instrumental values of freshwater invertebrates in the Cape Horn Biosphere Reserve.
Chicken embryos were chronically exposed to the ?1- blocker atenolol during one of three stages: mesonephros (E7-E9), mesonephros-metanephros (E11-E13), or metanephros (E15-E17). Mesonephros group hearts were larger than all other groups (P < 0.01). Mesonephros and metanephros group kidneys were larger than all remaining groups (P < 0.0001). The mesonephros group nephron number was ~40% lower than control values (P = 0.002). Glomerular areas were 26% and 18% larger than the control group in the mesonephros and metanephros groups, respectively (P < 0.001). These data suggest an E7-E9 critical window of cardiovascular and renal development for atenolol. Acute atenolol exposure in E15 embryos showed an increase in mean arterial pressure with all but the highest dose. All doses significantly decreased heart rate.
Triclosan (TCS) is an antimicrobial compound found in several types of common household products. After being washed down the drain, TCS will then end up in the local watershed. Although numerous studies have been conducted to evaluate the fate and effects of TCS in aquatic environments, there have been no studies evaluating the role arbuscular mycorrhizal fungi (AM) play in a plants response to TCS exposure. Three species of wetland plants native North Texas were inoculated with AM spores and exposed to 0, 0.4 g/L and 4.0 g/L TCS concentrations. Root morphology of E. prostrata and S. herbacea showed AM and exposure responses. S. herbacea produced the greatest amounts biomass and TCS bioaccumulation, in all but one treatment. It also displayed opposing results to E. prostrata in measures of root length, root surface area, relative root mass, relative shoot mass and shoot:root ratio. TCS root tissue concentrations increased with increased exposures for both E. prostrata and S. herbacea. Even though E. prostrata had the lowest levels in each measure of biomass production, it had the highest amount of root TCS bioaccumulation in the AM inoculated 4.0 g/L treatment. H. laevis was between the other two species in terms of biomass production, and did not demonstrate any exposure or inoculation effects in morphology or TCS accumulation. Overall, no clear patterns were detected, which highlights the fact that further study is required to completely understand the effects compounds like TCS can have on plant community structure, and ultimately ecosystem function.
Triclosan and triclocarban are antimicrobial compounds added to a variety of consumer products that are commonly detected in waste water effluent. The focus of this study was to determine whether the bioconcentration of these compounds in wetland plants and sediments exhibited species specific and site specific differences by collecting field samples from a constructed wetland in Denton, Texas. The study showed that species-specific differences in bioconcentration exist for triclosan and triclocarban. Site-specific differences in bioconcentration were observed for triclosan and triclocarban in roots tissues and sediments. These results suggest that species selection is important for optimizing the removal of triclosan and triclocarban in constructed wetlands and raises concerns about the long term exposure of wetland ecosystems to these compounds.
Altitudinal gradients in Sub-Antarctic freshwater systems present unique opportunities to study the effect of distinct environmental gradients on benthic macroinvertebrate community composition and dispersal. This study investigates patterns in biodiversity, dispersal and population genetic structure of benthic macroinvertebrate fauna across an altitudinal gradient between two watersheds on Navarino Island in southern Chile. Patterns in diversity, density, evenness and functional feeding groups were not significantly different across the altitudinal gradient in both the Windhond and Róbalo Rivers. Taxa richness in both rivers generally increased from the headwaters of the river to the mouth, and functional feeding group patterns were consistent with the predictions of the River Continuum Concept. Population genetic structure and gene flow was investigated by sampling the mitochondrial cytochrome oxidase I gene in two invertebrate species with different dispersal strategies. Hyalella simplex (Amphipoda) is an obligate aquatic species, and Meridialaris chiloeense (Ephemeroptera) is an aquatic larvae and a terrestrial winged adult. Contrasting patterns of population genetic structure were observed. Results for Hyalella simplex indicate significant differentiation in genetic structure in the Amphipod populations between watersheds and lower genetic diversity in the Róbalo River samples, which may be a result of instream dispersal barriers. Meridialaris chiloeense exhibited weak population structure but higher genetic diversity, which suggests this species is able to disperse widely as a winged adult.
Through the use of natural history records, published literature, and personal sampling (2011-2016) a total of 454 caddisfly species represented by 24 families and 93 genera were documented from the south-central United States. Two Hydroptilidae species were collected during the 2011-2016 collection efforts that are new to the region: Hydroptilia scheringi and Mayatrichia tuscaloosa. Eightteen species are endemic and 30 are considered species of concern by either federal or state agencies. The majority of each of these groups is Hydroptilidae, or microcaddisflies. Trichoptera community structure, by minimum number of species, was analysed in conjunction with large-scale geographical factors to determine which factor illustrated caddisfly community structure across the region. Physiographic provinces compared to other geographic factors analyzed best-represented caddisfly communities with a minimum of 10 or more species. Statistically, Hydrologic Unit Code 4 (HUC 4) was the most significant geographical factor but low number of samples representing this variable rendered it less representative of caddisfly community structure for the study area.
The northern bobwhite (Colinus virginianus) population decline is a severe, rangewide phenomenon beginning >150 years ago and continuing today. In this investigation, I: 1. document the timeline of bobwhite population decline and unintended genetic consequences of attempted remedies, 2) develop a model useful for predicting possible locations of potentially sustainable bobwhite populations in semiarid rangeland in Texas and Oklahoma, and 3) examine the relationship between population monitoring data and meteorological factors. While breeding season call counts of male bobwhite have been used for >70 years to provide estimates of fall populations for hunting, most studies of call counts have focused on mathematics and statistical accuracy of the count, largely overlooking the influence of meteorological factors on call counts. Here, I present the results of >4,400 individual point counts and examine their relationship with meteorological variables recorded at each stop. Humidity was positively correlated with the number of birds recorded (ρ = 0.275, p < 0.001) and temperature was negatively correlated (ρ = -0.252, p < 0.001). The number of birds recorded was significantly higher in wet years than in drought years. There was no significant correlation between wind velocity and number of birds recorded. These results suggest that, while weather does influence call counts and efforts should be made to record meteorological conditions when collecting call count data, the influence of weather may not easily factor into the analysis. These results also provide another line of evidence for decreased breeding behavior during high temperatures. With the increased focus on bobwhite habitat management on a regional scale, there is a need for reliable methods to identify potential bobwhite habitat. To identify bobwhite habitat in semiarid rangeland, I performed classification of LANDSAT scenes of Clay County, Texas from July and December 2015. Stands of mature little bluestem provide excellent bobwhite nesting cover …
Subsidiary mother cell (SMC) divisions during stomatal complex formation in Zea mays are asymmetric generating a small subsidiary cell (SC) and a larger epidermal cell. Mutants with a high number of abnormally shaped subsidiary cells include the brick1 (brk1) and discordia1 (dcd1) mutants. BRK1 is homologous to HSPC300, an ARP2/3 complex activator, and is involved in actin nucleation while DCD1 is a regulatory subunit of the PP2A phosphatase needed for microtubule generation (Frank and Smith, 2002; Wright et al. 2009). Possible causes of the abnormal SCs in brk1 mutants include a failure of the SMC nucleus to polarize in advance of mitosis, no actin patch, and transverse and/or no PPBs (Gallagher and Smith, 2000; Panteris et al 2006). The abnormal subsidiary mother cell division in dcd1 is due to correctly localized, but disorganized preprophase bands (PPBs; Wright et al. 2009). The observation that brk1 has defects in PPB formation and that the dcd1 phenotype is enhanced by the application of actin inhibitors led us to examine the dcd1; brk1 double mutant (Gallagher and Smith, 1999). We found that dcd1; brk1 double mutants demonstrate a higher percentage of aberrant SCs than the single mutants combined suggesting that these two mutations have a synergistic and additive effect on SC formation. Our observations and results are intriguing and the future step will be to quantitate the abnormal PPBs and phragmoplasts in the double and single mutants using immunolocalization of tubulin and actin as well as observations of live cells expressing tubulin-YFP.
I used the C. elegans genetic model to examine the role of ceramide biosynthesis (sphingolipid pathway) and iron regulation and found that each process impacts germline development and function. Using a sphingolipid specific antibody mAb15B4, I found that sphingolipids are associated with germ granules (P granules) within C. elegans and zebrafish; thus, suggesting conservation of macromolecules associated with germ granules. Phenotype analysis of ceramide biosynthesis mutants in C. elegans revealed that this pathway is essential for normal germline function in the aging adult hermaphrodite; specifically, precocious germline senescence was observed. Furthermore, I found that disruption of ceramide biosynthesis, via the hyl-2 deletion mutation, negatively impacts mAb15B4 localization at the P granules. Through genetic suppression analysis, I determined that insulin signaling and lipid biosynthesis can modulate the mAb15B4 localization to P granules. Additional, phenotype analysis showed that ceramide biosynthesis dysfunction decreased fecundity, and led to germline structure defects and uterine tumors. Through suppression analysis, I determined that modulation of the insulin signaling pathway suppressed the precocious germline senescence due to ceramide biosynthesis dysfunction. Since the presence of uterine tumors is associated with reproductive senescence I concluded that ceramide biosynthesis has a role in germline maintenance in the aging of the germline (germline senescence). The other important fate of a germ cell is programmed cell death. Apoptosis, which occurs through a highly conserved molecular pathway, is a normal component of growth and homeostatic processes. I used C. elegans to gain a greater understanding of the cisd gene function. The C. elegans genome has three previously uncharacterized cisd genes which code for CISD-1 (homology to vertebrate mitoNEET/CISD1 and NAF-1/CISD2) and CISD-3.1 and CISD-3.2 (homology to vertebrate Miner2/CISD3). I determined that independent disruption of the cisd genes resulted in a significant increase in the number of cell corpses within the adult hermaphrodite germline. Genetic …
Primary cilia are sensory organelles that are important for initiating cell division in the brain, especially through sonic hedgehog (Shh) signaling. Several lines of evidence suggest that the mitogenic effect of Shh requires primary cilia. Proliferation initiated by Shh signaling plays key roles in brain development, in neurogenesis in the adult hippocampus, and in the generation of glial cells in response to cortical injury. In spite of the likely involvement of cilia in these events, little is known about their characteristics. Centrosomes, which are associated with primary cilia, also have multiple influences on the cell cycle, and they are important in assembling microtubules for the maintenance of the cell’s cytoskeleton and cilia. The cilia of terminally differentiated neurons have been previously examined with respect to length, incidence, and receptors present. However, almost nothing is known about primary cilia in stem cells, progenitors, or differentiated glial cells. Moreover, it is not known how the properties of cilia and centrosomes may vary with cell cycle or proliferative potential, in brain or other tissues. This dissertation focuses first on neurogenesis in the hippocampal subgranular zone (SGZ). The SGZ is one of the few brain regions in mammals that gives rise to a substantial number of new neurons throughout adulthood. The neuron lineage contains a progression of identifiable precursor cell types with different proliferation rates. This present study found that primary cilia were present in every cell type in the neuronal lineage in SGZ. Cilium length and incidence were positively correlated among these cell types. Ciliary levels of adenylyl cyclase type III (ACIII) levels relative to ADP-ribosylation factor-like protein 13b (Arl13b) was higher in neurons than in precursor cells and glia, and also changed with the cell cycle. G-protein coupled receptors, SstR3, MCHR1, and Gpr161 receptors were only found in neuronal cilia. The levels …
This thesis consists of one major section with two subsections. The first subsection investigates the activity of Arthrobacter globiformis aspartate transcarbamylase's specific activity with increasing concentrations of the enzyme's substrate. Dihydroorotase (DHOase) activity was also measured with increasing concentrations of the substrate dihydroorotate. The second subsection collected data in order to classify the enzyme, resulting in a classification into the category of class A ATCases with bifunctional ATCase-DHOase complexes. The thesis provides evidence to broaden understanding of the ATCase and DHOase enzymes for members of the family that Arthrobacter belongs to.
Jackson Blue is a first magnitude spring in the karst terrane of northeast Florida. Previous studies have identified inorganic fertilizer as the source of high nitrate levels in the spring. Agricultural land use and karst vulnerability make Jackson Blue a good model for conservation concerns. This work offers an aggregation of studies relating to the springshed, providing a valuable tool for planning and conservation efforts in the region. An analysis of nitrate levels and other water quality parameters within the springshed did not reveal significantly different values between agricultural and forested land use areas. Confounding factors include: high transmissivity in the aquifer, interspersed land use parcels, and fertilizer application in forested areas due to commercial pine stand activity.
In the last decade emerging contaminants research has intensified in a bid to answer questions about fate, transport, and effects as these chemicals as they get released into the environment. The chemicals of interest were the antimicrobials; triclocarban (TCC) and triclosan (TCS), and a metabolite of triclosan, methyl triclosan (MTCS). This research was designed to answer the question: what is the fate of these chemicals once they are released from the waste water treatment plant into receiving streams. Three different assessment methods; field monitoring, in-situ experiments, and laboratory studies were used to answer the overall question. TCS, TCC, and MTCS levels were measured in surface water, sediment and the Asiatic clam Corbicula fluminea. Field studies were conducted using four sites at Pecan Creek, Denton TX. Levels of all three chemicals in clams were up to fives orders of magnitude the water concentrations but an order of magnitude lower than in sediment. Highest sediment levels of chemicals were measured in samples from the mouth of Pecan Creek (highest organic matter). TCC was the most and TCS was the least accumulated chemicals. In-situ and lab studies both indicated that uptake of these chemicals into the clams was very rapid and measurable within 24hours of exposure. The after clams were transferred into clean water most of the compounds were depurated within 14 days.
Austrofundulus limnaeus is a species of annual killifish which inhabits ephemeral ponds in South America. The species is able to survive seasonally desiccating ponds due to their ability to produce robust embryos. The embryos of this species are capable of entering a developmental arrest, termed diapause II, which precedes the onset of drought. While in this arrested state embryos exhibit the greatest tolerance to anoxia of any characterized vertebrate at 25ºC. Furthermore, when raised at 30ºC, embryos escape the entrance to diapause II and go on to develop directly. Currently, little is known about the molecular mechanisms which induce and maintain this developmentally arrested state. In this study I have developed methods to analyze changes in histone modifications in the context of diapause II. Histone modifications were chosen due to their extreme conservation and well characterized role as modulators of gene expression in other systems. Results utilizing adapted immunobased assays show significant changes in the global amount of H3S10P, H3K27me and H3K4me, as the embryos progress from early embryogenesis through the exit of diapause. Additionally, it is revealed that there exists a degree of phenotypic plasticity with regards to the entrance into diapause II which is modulated by the environment (severe hypoxia 0.1% O2). This work builds a foundation for future histone modification studies and contributes the development of several tools to the field. This study contributes to a greater molecular understanding of the cue(s) which influence the remarkable phenomenon of obligate developmental arrest in a vertebrate embryo.
This study determines effects of oxygen levels on morphology and VEGF expression of developing chicken lungs following incubation in normoxia (21% O2), hypoxia (15% O2) or hyperoxia (30% O2), until developmental days 16 or 18. Lung morphology was assessed using light microscopy, while VEGF expression was determined with ELISA. In hypoxia, the proportion of parabronchial tissue and parabronchi including lumina increased from day 16 to 18 (61 to 68% and 74.2 to 82.2%, respectively). Non-parabronchial tissue was higher in hypoxia than in hyperoxia on day 16 (26 to 20%). However, by day 18, there were no differences between groups. VEGF expression was 33% higher in hypoxia than in hyperoxia on day 16 (736 vs. 492 pg/ml). On day 18, VEGF expression was 43% higher in hyperoxia than in normoxia (673 to 381pg/ml), and remained elevated by 40% in hypoxia over normoxia (631 pg/ml). VEGF may be a mechanism by which parabronchial tissue is stimulated from day 16 to 18 following exposure to chronic hypoxia.
The Attwater’s prairie-chicken (Tympanuchus cupido attwateri; APC) is a species of grouse native to Texas coastal prairies and is on the critically endangered species list as a result of habitat destruction and overhunting. All of the current populations were captively bred and released into the wild. Survivorship for released APCs is very low, and individuals seldom survive to reproduce in the wild. One factor contributing to this may be an alteration in the gut microbiota as a result of captivity. Factors potentially influencing the gut microbial composition in captivity include antibiotic therapy, stress, and a predominantly commercially formulated diet. Recent studies have begun to shed light on the importance of the host microbial endosymbionts. Antibiotic administration, stress, diet, age, genotype and other factors have been shown to influence microbial populations in the gastrointestinal tracts of many different vertebrates. Sequencing of 16S rRNA gene amplicons on the Ion Torrent™ platform was used in this study to identify groups of bacteria in the cloacas as a surrogate for the gut microbiota in the APC. Antibiotic-treated and untreated birds, wild-hatched and captive-bred birds, and individuals sampled before and after release to the wild were examined. Significant differences were found between wild-hatched and captive raised birds both pre- and post release. In addition, there was extensive variation among the populations at the lower taxonomic ranks between individuals for each group of APCs. Principal coordinate analysis based on the weighted UniFrac distance metric further exhibited some clustering of individuals by treatment. These data suggest that captive breeding may have long-term effects on the cloacal microbiota of APCs with unknown consequences to their long-term health and survivorship.
Clustering techniques are important for gene expression data analysis. However, efficient computational algorithms for clustering time-series data are still lacking. This work documents two improvements on an existing profile-based greedy algorithm for short time-series data; the first one is implementation of a scaling method on the pre-processing of the raw data to handle some extreme cases; the second improvement is modifying the strategy to generate better clusters. Simulation data and real microarray data were used to evaluate these improvements; this approach could efficiently generate more accurate clusters. A new feature-based algorithm was also developed in which steady state value; overshoot, rise time, settling time and peak time are generated by the 2nd order control system for the clustering purpose. This feature-based approach is much faster and more accurate than the existing profile-based algorithm for long time-series data.
Few studies have been conducted on the effectiveness of phytoremediation of pharmaceutical compounds, although the persistent and non-acutely toxic nature of many of these compounds in today's water bodies may yield an ideal application for this practice. To quantify the potential effectiveness of plant uptake, kinetic and proportional bioconcentration factors (BCFk, and BCFp, respectively) in nanograms (ng) carbamazepine and amiodarone per gram (g) wet weight plant tissue for Lemna spp. were determined utilizing a 14-day continuous flow-through study. Samples were analyzed using isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) running in positive ion mode. Kinetic BCF was estimated at 0.538, while proportional BCF was estimated at 0.485. Kinetic BCF for the amiodarone study was estimated at 23.033, whereas proportional BCF was estimated at 41.340. Possible contamination of the C18 column and peristaltic pump failure may have impacted uptake results. In light of variability and current lack of research in the field, this work should be considered exploratory rather than conclusive.
Bacteriophages are viruses that specifically infect bacteria. When a phage infects a bacterium, it attaches itself to the surface of the bacteria and injects its DNA into the intracellular space. The phage DNA hijacks the cellular machinery of the bacteria and forces it to produce phage proteins. Eventually, the bacteria cell bursts or lyses, releasing new phage. The bacteria act as a host for phage reproduction. The ability for a phage to infect multiple bacterial species is known as host range. In siphoviridae bacteriophages, host range is thought to primarily be determined by proteins at the tip of their tail fibers. These proteins act as anti-receptors to specific receptors on the surface of bacteria. In siphoviridae Gram-positive infecting phages, the genes that code these proteins are typically located between the tape measure protein gene and the endolysin gene. It is hypothesized that phages that have similar anti-receptor proteins will have similar host range. In this study, the host ranges of 12 BD1 bacteriophages were tested on 9 different Streptomyces species. In these 12 phages, the genes between the tape measure protein gene and endolysin gene were compared. The 12 phages had high levels of variability in these genes. Five genes in this region had unknown functions and were called position A, B, C, D, and E. Position A-E were BLASTed on NCBI and Phages-DB and their results were recorded. The functions of position A, C, and E remain unknown. The function of position D is most likely a minor tail protein. Position B had BLAST hits for a collagen-like protein and a putative tail fiber protein. Position B was inspected further, and it was found that it contained Gly-X-Y repeats in its amino acid sequence. Position B also had some conservation in its N-terminal amino acid sequence, specifically where the …
The pyrimidine biosynthetic pathway is essential and similar in all bacteria. The pathway from Pseudomonas is regulated by nucleotides which bind to the upstream region of the pyrBC’ gene complex. Work in our lab mapped the genes and showed that the pyrB and pyrC’ were part of an overlap complex. The Pseudomonas aeruginosa has one circular chromosome. A former Pseudomonas now called Burkholderia cepacia is similar to P. aeruginosa except that it contains three circular chromosomes (CI, CII, CIII) and one large plasmid. The primary chromosome named CI contains the pyrBC’. To our knowledge there has been no report of the activity of ATCase in Pseudomonas and contrasted with that of Burkholderia. Here, we compare the activity of ATCase in P. aeruginosa and B .cepacia. Cells of both organisms were grown in Pseudomonas minimal medium and in Enriched medium. The ATCase was extracted and partially purified from each sample. It is hypothesized that the B. cepacia has greater activity for ATCase than do the Pseudomonas.
Macroinvertebrates associated with the aquatic plant, water stargrass (Heteranthera dubia), were sampled from 12 waterbodies in four regions of the United States from June to August 2005. Taxa richness, evenness, and diversity were lowest in the Lower Midwest (LMW) region, and higher in Northern sites, especially the Upper Midwest (UMW), and Northeast (NE). While relative abundance varied from site to site and region to region, utilization of the plant by functional groups remained fairly constant. Collector-gatherers consistently comprised the largest portion of invertebrates sampled. The shredder/ herbivore functional group comprised an average of 17 % of total groups. Through an exhaustive literature review, it was found that shredder/ herbivores of water stargrass have not been reported in the literature. Because of this, the herbivore group was analyzed separately and consisted of 2,383 specimens representing 23 species. The most common groups were Rhopalosiphum sp., Nectopsyche spp. and chironomids. No differences were found in herbivore diversity or evenness between sampling regions, but species richness was significantly different.
This work explores the metabolism of mercury in liver and spleen tissue of fish from a methylmercury contaminated wetland. Wild-caught bass, catfish, bowfin and gar were collected. Macrophage centers, which are both reactive and primary germinal centers in various fish tissues, were hypothesized to be the cause of demethylation of methylmercury in fish tissue. Macrophage centers are differentially expressed in fish tissue based on phylogenetic lineage, and are found primarily in the livers of preteleostean fish and in the spleen of teleostean fish. Histology of liver and spleen was examined in both control and wild-caught fish for pathology, size and number of macrophage centers, and for localization of mercury. Total mercury was estimated in the muscle tissue of all fish by direct mercury analysis. Selenium and mercury concentrations were examined in the livers of wild-caught fish by liquid introduction inductively coupled plasma mass spectrometry (ICP-MS). Total mercury was localized in histologic sections by laser ablation ICP-MS (LA-ICP-MS). Mercury speciation was determined for inorganic and methylmercury in liver and spleen of fish by bas chromatography-cold vapor atomic fluorescence spectroscopy (GC-CVAFS). Macrophage center tissue distribution was found to be consistent with the literature, with a predominance of centers in preteleostean liver and in spleens of teleostean fish. Little evidence histopathology was found in the livers or spleens of fish examined, but differences in morphology of macrophage centers and liver tissue across species are noted. the sole sign of liver pathology noted was increased hepatic hemosiderosis in fish with high proportions of liver inorganic mercury. Inorganic mercury was found to predominate in the livers of all fish but bass. Organic mercury was found to predominate in the spleens of all fish. Mercury was found to accumulate in macrophage centers, but concentrations of mercury in this compartment were found to vary less in relation …
Seeds from the desert shrub Simmondsia chinensis (jojoba) are one of the only known natural plant sources to store a majority of its oil in the form of liquid wax esters (WE) instead of triacylglycerols (TAGs) and these oils account for ~55% of the seed weight. Jojoba oil is highly valued as cosmetic additives and mechanical lubricants, yet despite its value much is still unknown about its neutral lipid biosynthetic pathways and lipid droplet packaging machinery. Here, we have used a multi-"omics" approach to study how spatial differences in lipid metabolites, gene expression, and lipid droplet proteins influence the synthesis and storage of jojoba lipids. Through these studies mass spectrometry analyses revealed that WEs are compartmentalized primarily in the cotyledonary tissues, whereas TAGs are, surprisingly, localized to the embryonic axis tissues. To study the differences in gene expression between these two tissues, a de novo transcriptome was assembled from high throughput RNAseq data. Differential gene expression analysis revealed that the Jojoba Wax Synthase, which catalyzes the formation of wax esters, and the Diacylglycerol O-Acyltransferase1, which catalyzes the final acylation of triacylglycerol synthesis, were differentially expressed in the cotyledons and embryonic axis tissues, respectively. Furthermore, through proteomic analysis of lipid droplet proteins from lipid droplets of the cotyledons and embryonic axis, it was estimated that each of these tissues contains a different proportion of the major lipid droplet proteins, oleosins, steroleosins, caleosins, and lipid droplet associated proteins. The Jojoba Olesosin1, Lipid Droplet Associated Protein 1, and Lipid Droplet Associated Protein 3, were identified as potential lipid droplet proteins that could be important for storage of wax esters. The coding sequences of these genes were transiently expressed in N. benthamiana leaves individually, and with co-expression of Mus musculus diacylglycerol acyltransferase 2, and in all cases were able to induce neutral lipid accumulation. …
A study of Asian clam (Corbicula fluminea Müller) colonization in relation to changes in aquatic vegetation community as a result of management of Hydrilla verticillata (L. f.) Royle with grass carp was conducted at the Lewisville Aquatic Ecosystem Research Facility (LAERF), Lewisville, TX, from April 2015 through October 2016. Percent vegetation cover, C. fluminea abundance and water quality metrics (pH, turbidity, conductivity, DO, calcium, chlorophyll a) from 16 experimental subjects were analyzed. Treatments included four replicated grass carp stocking densities; 1-control with no fish stocked (n = 4), 2-low density of 40-43 fish per vegetated ha (n = 4), 3-medium density of 72-81 fish per vegetated ha (n = 4) and 4-high density of 110-129 fish per vegetated ha (n = 4). Data analysis showed statistical significance in the relation of C. fluminea abundance to percent vegetation cover (multiple linear regression, r2 = 0.820), grass carp stocking densities (two-way analysis of variance, p = <0.001) and chlorophyll a (multiple linear regression, r2 = 0.339). Findings of this research indicate the possibility that management of hydrilla had enabled establishment of secondary invasive species.
With an increasing population suffering from obesity or Diabetes Mellitus (DM), it is more pertinent than ever to understand how physiological changes impact cellular processes. Patients with DM often suffer from obesity, hyperglycemia, altered fatty acids that contribute to vascular dysfunction, and increased risk to ischemia. Caenorhabditis elegans is a model system used to study the conserved insulin signaling pathway, cellular responses in whole organisms and the impact a glucose diet has on oxygen deprivation (anoxia) responses. RNA-sequencing (RNA-Seq) was used to analyze the expression of genes in the anoxia sensitive populations of N2 (wild-type) fed glucose and hyl-2(tm2031), a mutant with altered ceramide metabolism. Comparison of the altered transcripts in the anoxia sensitive populations revealed 199 common transcripts- 192 upregulated and 7 downregulated. One of the gene families that have altered expression in the anoxia sensitive populations encode for Cytochrome P450 (CYP). CYPs are located both in the mitochondria and endoplasmic reticulum (ER), but the CYPs of interest are all predicted to be mainly subcellularly localized to the ER. Here, I determined that knock-down of specific cyp genes, using RNA interference (RNAi), increased anoxia survival in N2 animals fed a standard diet. Anoxia sensitivity of the hyl-2(tm2031) animals was supressed by RNAi of cyp-25A1 or cyp-33C8 genes. These studies provide evidence that the CYP detoxification system impacts oxygen deprivation responses. using hsp-4::GFP animals, a transcriptional reporter for ER unfolded protein response (UPR), I further investigated the impact of cyp knock-down, glucose, and anoxia on ER UPR due to the prediction of CYP-33C8 localization to the ER. Glucose significantly increased ER UPR and cyp knock-down non-significantly increased ER UPR. Measurements of ER UPR due to anoxia were made difficult, but representative images show an increase in ER stress post 9-hour anoxia exposure. This study provides evidence that glucose affects ER …
Mefloquine hydrochloride is an antimalarial agent that has been used for the past 40 years. Numerous reports of neurological side effects have recently led the FDA to issue a strong warning regarding long-term neurological effects. This warning lead to the U.S. Army’s Special Forces and other components to discontinue its use in July of 2013. Despite reported adverse side effects, mefloquine remains in circulation and is recommended to travelers going to specific Asian countries. Mefloquine has been used as a treatment for those already infected with the malaria parasite (blood concentrations ranging from 2.1 to 23 µM), and as prophylaxis (blood concentrations averaging 3.8 µM) (Dow 2003). The purpose of this study was to quantify Mefloquine’s toxicity using spontaneously active nerve cell networks growing on microelectrode arrays in vitro and to identify compounds that alleviate or reduce toxic effects. The current literature on mefloquine toxicity is lacking electrophysiological data. These data will contribute to research on the mechanism of adverse side effects associated with mefloquine use. Sequential titration experiments were performed by adding increasing concentrations of mefloquine solution to cultured neurons. Network responses were quantified and reversibility was examined. In each network, activity decreases were normalized as a percent of reference activity yielding a mean IC50 value of 5.97 ± 0.44 (SD) µM (n=6). After total activity loss, no activity was recovered with two successive medium changes. To test for network response desensitization resulting from sequential applications over 5-6 hr periods, one-point titrations at varying concentrations were conducted with fresh networks. These experiments yielded a single concentration response curve with an IC50 value of 2.97 µM. This represents a statistically significant shift (p < 0.0001) to lower concentrations of mefloquine, demonstrating that sequential applications result in network desensitization. After mefloquine exposures, cells were evaluated for irreversible cytotoxic damage. Over a …
One of the challenges in biology is placing a function on the myriad of gene sequences having become available from rapid advances in genome sequencing. One such example is a gene cluster (Nit1C) found in bacteria that is tied to the unusual ability of certain bacteria to grow when supplied cyanide as the sole nitrogen source. The term cyanotrophs has been applied to such bacteria, for which a genetic linkage between cyanotrophy and Nit1C was demonstrated for 10 separate bacteria. In addition to growth, cyanide induced the expression of Nit1C genes in all organisms tested, and in one case, deletion of one of the Nit1C genes (nitC) caused a loss of growth. Of the ten bacteria able to grow cyanotrophically, all gave evidence of harboring Nit1C on their genome except for two (Pseudomonas fluorescens Pf11764 and P. monteilii BCN3), which were sequenced and the presence of Nit1C was also confirmed. A broader search of bacteria identified 270 separate strains with the cluster, all limited to bacteria spanning the phyla Firmicutes, Actinobacteria, Proteobacteria and Cyanobacteria. Remarkably, many examples of a single representative of a given taxon contained Nit1C, most poignantly displayed by Pf11764 and PmBCN3; the interpretation being the cluster was likely acquired by horizontal gene transfer in response to cyanide as an environmental cue. Consistent with its absence in Archaea is the time line for the emergence of cyanide producing organisms (cyanogens) on earth dating back only 400-500 million years.
Factor VIII is a clotting factor found on the intrinsic side of the coagulation cascade. A mutation in the factor VIII gene causes the disease Hemophilia A, for which there is no cure. The most common treatment is administration of recombinant factor VIII. However, this can cause an immune response that renders the treatment ineffective in certain hemophilia patients. For this reason a new treatment, or cure, needs to be developed. Gene editing is one solution to correcting the factor VIII mutation. CRISPR/Cas9 mediated gene editing introduces a double stranded break in the genomic DNA. Where this break occurs repair mechanisms cause insertions and deletions, or if a template oligonucleotide can be provided point mutations could be introduced or corrected. However, to accomplish this goal for editing factor VIII mutations, a way to deliver the components of CRISPR/Cas9 into somatic cells is needed. In this study, I confirmed that the CRISPR/Cas9 system was able to create a mutation in the factor VIII gene in zebrafish. I also showed that the components of CRISPR/Cas9 could be piggybacked by vivo morpholino into a variety of blood cells. This study also confirmed that the vivo morpholino did not interfere with the gRNA binding to the DNA, or Cas9 protein inducing the double stranded break.
Plants possess unique features in many aspects of development. One of these features is seen in cell wall placement during cytokinesis, which is determined by the position of the preprophase band (PPB) and the subsequent expansion of the phragmoplast that deposits the new cell wall. During phragmoplast expansion, the phragmoplast tracks to the cortical division site, which was delineated by the PPB. Thus the position of the PPB determines the orientation of the division plane. In Arabidopsis thaliana, TONNEAU2 (TON2) is required for PPB formation and has been shown to interact with a type A subunit of the PP2A phosphatase in the yeast two-hybrid system. In Arabidopsis tonneau2 (ton2) mutants, abnormalities of the cortical microtubule cytoskeleton, such as disorganization of the interphase microtubule array and lack of PPB formation before mitosis markedly affects cell shape and arrangement as well as overall plant morphology. Loss of dcd1/add1, the maize ton2 homologues gives rise to a similar phenotype in Zea mays. The TON2 protein has two EF hand domains which are calcium-binding sites. Since calcium has been known to play key roles in several areas of plant functioning, the following question was raised: “Does calcium binding contribute to the localization and function of TONNEAU at the PPB?” To address this question, a series of constructs were generated to determine if TON2 binds calcium. Additionally, Ca2+ binding sites were mutated in constructs containing the TON2 gene fused to GFP or YPF. These constructs were then transformed into ton2 mutant plants and the localization of TON2 fusion protein and whether the construct is capable of rescuing the mutant phenotype were observed. Although, localization of TON2 to the PPB was not observed, the presence of the constructs were confirmed in the transformed plants using selection markers and by observing fluorescence under a confocal microscope.
The primary objective of this work was to determine the tissue-specific bioconcentration factors (BCFs) of the selected pharmaceuticals: norethindrone (NET), ibuprofen (IBU), verapamil (VER), clozapine (CLZ) and fenofibrate (FFB) in two freshwater fishes: fathead minnow and channel catfish. BCF tests on fathead followed the standard OECD 42-day test while a 14-day abridged test design was used in catfish exposures. Additional objectives included a) comparing the measured BCFs to the US EPA's BCFWIN model predicted values, b) comparing the BCF results from the standard and reduced tests, and c) prediction of chronic risk of the pharmaceuticals in fish using the human therapeutic plasma concentrations. Each test included uptake and depuration phases to measure tissue-specific kinetic BCFs. The results indicated that all the pharmaceuticals, except IBU, have the potential for accumulation in fish. Estimated BCFs for NET, VER and FFB may not be significant in view of the current regulatory trigger level (BCF ≥ 2000); however, CLZ's BCF in the liver had approached the criterion level. Significant differences were noticed in the tissue-specific uptake levels of the pharmaceuticals with the following general trend: (liver/kidney) > (gill/brain) > (heart/muscle) > plasma. IBU uptake was highest in the plasma. When compared to the measured BCFs, predicted values for NET, IBU, VER and FFB were slightly overestimated but did not differ largely. However, the measured BCF of CLZ in the liver was approximately two-orders of magnitude higher than the predicted level. The tissue-BCFs for the two species were not widely different indicating the potential usefulness of the reduced BCF test. Comparison of fish and human plasma levels indicated that NET, CLZ and VER have the potential to cause chronic effects in fish.
Natural history collections house immense amounts of data, but the majority of data is only accessible by locating the collection label, which is usually attached to the physical specimen. This method of data retrieval is time consuming and can be very damaging to fragile specimens. Digitizing the collections is the one way to reduce the time and potential damage related to finding the collection objects. The Elm Fork Natural Heritage Museum is a natural history museum located at the University of North Texas and contains collections of both vertebrate and invertebrate taxa, as well as plants. This project designed a collection digitization workflow for Elm Fork by working through digitizing the Benjamin B. Harris Herbarium. The collection was cataloged in Specify 6, a database program designed for natural history collection management. By working through one of the museum’s collections, the project was able to identify and address challenges related to digitizing the museum’s holdings in order to create robust workflows. The project also produced a series of documents explaining common processes in Specify and a data management plan.
A method was developed for extending a fine-scaled forest gap model to a watershed-scaled landscape, using the Eastern Cross Timbers ecoregion as a case study for the method. A topographic wetness index calculated from digital elevation data was used as a measure of hydrologic across the modeled landscape, and the gap model modified to have with a topographically-based hydrologic input parameter. The model was parameterized by terrain type units that were defined using combinations of USDA soil series and classes of the topographic wetness index. A number of issues regarding the sources, grid resolutions, and processing methods of the digital elevation data are addressed in this application of the topographic wetness index. Three different grid sizes, 5, 10, and 29 meter, from both LiDAR-derived and contour-derived elevation grids were used, and the grids were processed using both single-directional flow algorithm and bi-directional flow algorithm. The result of these different grids were compared and analyzed in context of their application in defining terrain types for the forest gap model. Refinements were made in the timescale of gap model’s weather model, converting it into a daily weather generator, in order to incorporate the effects of the new topographic/hydrologic input parameter. The precipitation model was converted to use a Markov model to initiate a sequence of wet and dry days for each month, and then daily precipitation amounts were determined using a gamma distribution. The output of the new precipitation model was analyzed and compared with a 100-year history of daily weather records at daily, monthly, and annual timescales. Model assumptions and requirements for biological parameters were thoroughly investigated and questioned. Often these biological parameters are based on little more than assumptions and intuition. An effort to base as many of the model’s biological parameters on measured data was made, including a new …
Metagenomics is the study of the totality of the complete genetic elements discovered from a defined environment. Different from traditional microbiology study, which only analyzes a small percent of microbes that could survive in laboratory, metagenomics allows researchers to get entire genetic information from all the samples in the communities. So metagenomics enables understanding of the target environments and the hidden relationships between bacteria and diseases. In order to efficiently analyze the metagenomics data, cutting-edge technologies for analyzing the relationships among microbes and communities are required. To overcome the challenges brought by rapid growth in metagenomics datasets, advances in novel methodologies for interpreting metagenomics data are clearly needed. The first two chapters of this dissertation summarize and compare the widely-used methods in metagenomics and integrate these methods into pipelines. Properly analyzing metagenomics data requires a variety of bioinformatcis and statistical approaches to deal with different situations. The raw reads from sequencing centers need to be processed and denoised by several steps and then be further interpreted by ecological and statistical analysis. So understanding these algorithms and combining different approaches could potentially reduce the influence of noises and biases at different steps. And an efficient and accurate pipeline is important to robustly decipher the differences and functionality of bacteria in communities. Traditional statistical analysis and machine learning algorithms have their limitations on analyzing metagenomics data. Thus, rest three chapters describe a new phylogeny based machine learning and feature selection algorithm to overcome these problems. The new method outperforms traditional algorithms and can provide more robust candidate microbes for further analysis. With the frowing sample size, deep neural network could potentially describe more complicated characteristic of data and thus improve model accuracy. So a deep learning framework is designed on top of the shallow learning algorithm stated above in order to further …
Liquid chromatography-mass spectrometry (LC-MS) based proteomic methods have provided archaeologists with a powerful tool for the discovery and identification of proteins within artifacts. Traditionally, discovery-based methods have utilized a non-targeted full mass scan method in an attempt to identify all proteins present within a given sample. However, increased sensitivity is often needed to target specific proteins in order to test hypotheses. Proteins present within archaeological materials present a unique challenge, as they are often subjected to a variety of chemical transformations both before and after burial. Any preserved proteins will be present within a complex mixture of compounds, and full mass scans often fail to detect less abundant proteins of interest. Consistent and reliable targeted methods are needed to detect protein biomarkers. Taphonomic experimentation was employed as a means to identify the effect of particular processes and conditions on the preservation of mare's milk proteins. In addition, three LC-MS methods were evaluated for their efficiency in identifying mare's milk-specific peptide biomarkers from experimental pottery samples. The ability to reliably detect the presence of these species-specific peptides can help provide evidence about past cultural groups, including the origins of dairying and animal domestication.
Improvements in mass spectrometry (MS)-based strategies for characterizing the plant lipidome through quantitative and qualitative approaches such as shotgun lipidomics have substantially enhanced our understanding of the structural diversity and functional complexity of plant lipids. However, most of these approaches require chemical extractions that result in the loss of the original spatial context and cellular compartmentation for these compounds. To address this current limitation, several technologies were developed to visualize lipids in situ with detailed chemical information. A subcellular visualization approach, direct organelle MS, was developed for directly sampling and analyzing the triacylglycerol contents within purified lipid droplets (LDs) at the level of a single LD. Sampling of single LDs demonstrated seed lipid droplet-to-droplet variability in triacylglycerol (TAG) composition suggesting that there may be substantial variation in the intracellular packaging process for neutral lipids in plant tissues. A cellular and tissue visualization approach, MS imaging, was implemented and enhanced for visualizing the lipid distributions in oilseeds. In mature cotton seed embryos distributions of storage lipids (TAGs) and their phosphatidylcholine (PCs) precursors were distribution heterogeneous between the cotyledons and embryonic axis raising new questions about extent and regulation of oilseed heterogeneity. Extension of this methodology provides an avenue for understanding metabolism in cellular (perhaps even subcellular) context with substantial metabolic engineering implications. To visualize metabolite distributions, a free and customizable application, Metabolite Imager, was developed providing several tools for spatially-based chemical data analysis. These tools collectively enable new forms of visualizing the plant lipidome and should prove valuable toward addressing additional unanswered biological questions.
von Willebrand factor (VWF) protein acts in the intrinsic coagulation pathway by stabilizing FVIII from proteolytic clearance and at the site of injury, by promoting the adhesion and aggregation of platelets to the exposed subendothelial wall. von Willebrand disease (VWD) results from quantitative and qualitative deficiencies in VWF protein. The variability expressivity in phenotype presentations is in partly caused by the action of modifier genes. Zebrafish has been used as hemostasis animal model. However, it has not been used to evaluate VWD. Here, we report the development of a heterozygote VWF mutant zebrafish using the genome editing CRISPR/Cas9 system to screen for modifier genes involved in VWD. We designed CRISPR oligonucleotides and inserted them into pT7-gRNa plasmid. We then prepared VWF gRNA along with the endonuclease Cas9 RNA from Cas9 plasmid. We injected these two RNAs into 1-4 cell-stage zebrafish embryos and induced a mutation in VWF exon 29 of the zebrafish with a mutagenesis rate of 16.6% (3/18 adult fish). Also, we observed a germline transmission with an efficiency rate of 5.5% (1/18 adult fish). We obtained a deletion in exon 29 which should result in truncated VWF protein.
The question of post-mortem interval (PMI) or time since death is often the most sought after piece of information associated with a medical death investigation. Based on the observation that DNA degradation disproportionately affects the analysis of larger genetic loci, it was proposed that DNA degradation, as a result of autolysis or putrefaction, could prove suitable as a potential rate-of-change indicator of PMI. Nine randomly amplified polymorphic DNA (RAPD) analysis primers and three sets of directed amplification primers were evaluated to determine their suitability for use in assessing the degree of DNA fragmentation in tissue samples. They were assessed for amplicon specificity, total DNA target sensitivity, allele monomorphism and the observance of degradation-based profile changes. Markers meeting the requisite criteria were then used to assess a range samples degraded under controlled and uncontrolled conditions. Tissue samples collected from seven domestic pigs (Sus scrofa) were incubated under controlled laboratory or uncontrolled field conditions to produce samples simulating those potentially collected in a forensic case. DNA samples isolated from these specimens were then analyzed at those loci which had been determined to meet the requisite criteria. Collectively, data generated from these analyses indicate that genetic profiles generated by this approach can provide information useful for estimating the post-mortem interval, with the locus and amplicons used being most useful during the first 72 hours after death.
This dissertation investigates the spatio-temporal dynamics of intermittent stream fish metacommunities in response drought-induced fragmentation and re-connectivity using both field and experimental approaches. A detailed field study was conducted in two streams and included pre-drought, drought, and post-drought hydrological periods. Fish assemblages and metacommunity structure responded strongly to changes in hydrological conditions with dramatic declines in species richness and abundance during prolonged drought. Return of stream flows resulted in a trend toward recovery but ultimately assemblages failed to fully recover. Differential mortality, dispersal, recruitment among species indicates species specific responses to hydrologic fragmentation, connectivity, and habitat refugia. Two manipulative experiments tested the effects of drought conditions on realistic fish assemblages. Fishes responded strongly to drought conditions in which deeper pools acted as refugia, harboring greater numbers of fish. Variability in assemblage structure and movement patterns among stream pools indicated species specific habitat preferences in response predation, resource competition, and desiccation. Connecting stream flows mediated the impacts of drought conditions and metacommunity dynamics in both experiments. Results from field and experimental studies indicate that stream fish metacommunities are influenced by changes in hydrological conditions and that the timing, duration, and magnitude of drought-induced fragmentation and reconnecting stream flows have important consequences metacommunity dynamics.
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Each of the Months listed note their name and the number of records that will be limited down to if you choose that option.
This dialog allows you to filter your current search.
Each of the Rights Access listed note their name and the number of records that will be limited down to if you choose that option.
This dialog allows you to filter your current search.
Each of the Discipline listed note their name and the number of records that will be limited down to if you choose that option.
The list can be sorted by name or the count.
This dialog allows you to filter your current search.
Each of the Degree Level listed note their name and the number of records that will be limited down to if you choose that option.