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Application of Cultured Neuronal Networks for Use as Biological Sensors in Water Toxicology and Lipid Signaling.
This dissertation research explored the capabilities of neuronal networks grown on substrate integrated microelectrode arrays in vitro to be applied to toxicological research and lipid signaling. Chapter 1 details the effects of chlorine on neuronal network spontaneous electrical activity and pharmacological sensitivity. This study demonstrates that neuronal networks can maintain baseline spontaneous activity, and respond normally to pharmacological manipulations in the present of three times the chlorine present in drinking water. The findings suggest that neuronal networks may be used as biological sensors to monitor the quality of water and the presence of novel toxicants that cannot be detected by conventional sensors. Chapter 2 details the neuromodulatory effects of N-acylethanolamides (NAEs) on the spontaneous electrical activity of neuronal networks. NAEs are a group of lipids that can mimic the effects of marijuana and can be derived from a variety of plant sources including soy lecithin. The most prominent NAEs in soy lecithin, palmitoylethanolamide (PEA) and linoleoylethanolamide (LEA), were tested individually and were found to significantly inhibit neuronal spiking and bursting activity. These effects were potentiated by a mixture of NAEs as found in a HPLC enriched fraction from soy lecithin. Cannabinoid receptor-1 (CB1-R) antagonists and other cannabinoid pathway modulators indicated that the CB1-R was not directly involved in the effects of NAEs, but that enzymatic degradation and cellular uptake were more likely targets. The results demonstrate that neuronal networks may also be a viable platform for the elucidation of biochemical pathways and drug mechanisms of action.
Automated Low-cost Instrument for Measuring Total Column Ozone
Networks of ground-based and satellite borne instruments to measure ultraviolet (UV) sunlight and total column ozone have greatly contributed to an understanding of increased amounts of UV reaching the surface of the Earth caused by stratospheric ozone depletion. Increased UV radiation has important potential effects on human health, and agricultural and ecological systems. Observations from these networks make it possible to monitor total ozone decreases and to predict ozone recovery trends due to global efforts to curb the use of products releasing chemicals harmful to the ozone layer. Thus, continued and expanded global monitoring of ozone and UV is needed. However, existing automatic stratospheric ozone monitors are complex and expensive instruments. The main objective of this research was the development of a low-cost fully automated total column ozone monitoring instrument which, because of its affordability, will increase the number of instruments available for ground-based observations. The new instrument is based on a high-resolution fiber optic spectrometer, coupled with fiber optics that are precisely aimed by a pan and tilt positioning mechanism and with controlling programs written in commonly available software platforms which run on a personal computer. This project makes use of novel low-cost fiber optic spectrometer technology. A cost advantage is gained over available units by placing one end of the fiber outdoors to collect sunlight and convey it indoors, thereby allowing the spectrometer and computer to be placed in a controlled environment. This reduces the cost of weatherproofing and thermal compensation. Cost savings also result from a simplified sun targeting system, because only a small pan and tilt device is required to aim the lightweight fiber optic ends. Precision sun-targeting algorithms, optical filter selection, and software to derive ozone from spectral measurements by the spectrometer are a major contribution of this project. This system is a flexible platform …
Biogeography of Montane Mammals on the Colorado Plateau and Adjacent Regions
This study identifies the biogeographic factors that structure small mammal communities on mountains of the Colorado Plateau and adjacent regions. Forty six isolated ranges were characterized across a 5-state study area encompassing the Colorado Plateau, including the central high plateaus of Utah and the Basin and Range Province (i.e. the Great Basin and mountains of Arizona and New Mexico). Presence/absence data of 25 montane mammal species were used to explore the interactions between historical and ecological processes affecting local and regional diversity patterns. Multivariate analyses, such as non-metric dimensional scaling, were used to explore factors which influence community composition. Results of these analyses revealed the Colorado River as a significant biogeographic barrier that affects montane mammal community structure. MtDNA cytochrome b sequence variation was analyzed among populations of the long-tailed vole, Microtus longicaudus, sampled from five interior ranges of the Colorado Plateau- Abajo, LaSal, Henry, and Chuska Mts., and Boulder Mountain of the Aquarius Plateau-and analyzed using traditional phylogenetic approaches (parsimony and likelihood) as well as nested clade analysis. Results support previous documentation of a major east-west phylogeographic break occurring between populations southeast of the Colorado River (eastern Arizona, Colorado, Wyoming and New Mexico) and all other western populations, which include a central clade, a northwest clade, and an Alaskan island clade. Evidence also supports differentiation of a 'southern Rockies' clade and a distinct 'southwest island' clade. Populations of M. longicaudus north and west of the Colorado River (Boulder and Henry Mts.) share two haplotypes, form a well-supported subclade with populations from the Kaibab plateau, and are closely related to the Northwest clade. Past approaches to studying montane mammal communities utilizing theory based on island biogeography have overemphasized area and isolation as the only forces structuring insular communities. As a result, there has been a lack of recognition of the …
Ceramide Biosynthesis and NEET Proteins Impact Development, Function, and Maintenance of the Caenorhabditis elegans Germline
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 …
Chemotactic Response of Lumbricus terrestris Coelomocytes to Larval and Adult Stages of Rhabditis pellio
Experiments were performed to assess the suitability of Rhabditis pellio, a nematode found in earthworms, as a challenge organism for use in development of a biomarker assay to determine the potential of chemicals to suppress the immunocompetence of the non-specific immune system. To accomplish this goal, information on the life cycle of R. pellio was determined; including effects of incubation time and temperature on growth rates; along with information on the immune response elicited in the earthworm, Lumbricus terrestris. Immune parameters measured were coelomocyte migration toward and attachment to R. pellio larvae and adults. Preliminary background information showed that R. pellio has potential as a challenge organism for development of a biomarker assay.
Comparative Morphology of Sensilla Styloconica on the Proboscis of North American Nymphalidae and Other Selected Taxa: Systematic and Ecological Considerations.
Sensilla styloconica on the proboscis of 107 species of North American and tropical butterflies were comparatively studied using the scanning electron microscope. Focus was on 76 species of North American Nymphalidae representing 45 genera and 11 subfamilies. Nomenclature for generalized and specific types of nymphalid sensilla is proposed. Written descriptions and micrographs are presented for each species studied. Morphological features were generally consistent for all or most species within genera and sometimes within subfamilies, with specified exceptions. Statistical analysis revealed significant differences for six of eight variables tested between two distinct feeding guilds of North American Nymphalidae. Average number, density, extent of proboscis coverage with sensilla, their total length, and shoulder spine length were all significantly greater in the non-nectar feeding guild than in nectar feeders, and may indicate adaptation for greater efficiency in feeding on flat surfaces. The greater frequency of apical shoulder spines in non-nectar feeders may represent adaptation for protection of sensory pegs from mechanical abrasion during feeding, or for anchoring the flexible proboscis tip to the surface. Correlation analysis revealed 9 out of 28 positive correlations in nectar feeders and 5 out of 28 in non-nectar feeders. Results of preliminary cladistic analysis were not considered to be meaningfully robust due to few available characters. The stylar characters identified in this study should be more useful in future analyses when included with characters from other lines of evidence. The presence of sensilla styloconica in all subfamilies of Nymphalidae, except Danainae, largely supports Ehrlich's (1958) higher classification concept for the family. The presence of less conspicuous sensilla in the Danainae, and other characteristics are presented as further evidence that they should be reconsidered for full family status. Sensilla styloconica in nymphalid butterflies appear to function as extensions that provide greater sensory reach during feeding. The role of these …
Determination of Dissociation Constants for GABAA Receptor Antagonists using Spontaneously Active Neuronal Networks in vitro
Changes in spontaneous spike activities recorded from murine frontal cortex networks grown on substrate-integrated microelectrodes were used to determine the dissociation constant (KB) of three GABAA antagonists. Neuronal networks were treated with fixed concentrations of GABAA antagonists and titrated with muscimol, a GABAA receptor agonist. Muscimol decreased spike activity in a concentration dependent manner with full efficacy (100% spike inhibition) and a 50% inhibitory concentration (IC50) of 0.14 ± 0.05 µM (mean ± SD, n=6). At 10, 20, 40 and 80 µM bicuculline, the muscimol IC50 values were shifted to 4.3 ± 1.8 µM (n=6), 6.8 ± 1.7 µM (n=6), 19.3 ± 3.54 µM (n=10) and 43.5 µM (n=2), respectively (mean ± SD). Muscimol titration in the presence of 10, 20, 40 µM of gabazine resulted in IC50s values of 20.1 (n=2), 37.17 (n=4), and 120.45 (n=2), respectively. In the presence of 20, 80, and 160 µM of TMPP (trimethylolpropane phosphate) the IC50s were 0.86 (n=2), 3.07 (n=3), 6.67 (n=2) µM, respectively. Increasing concentrations of GABAA antagonists shifted agonist log concentration-response curves to the right with identical efficacies, indicating direct competition for the GABAA receptor. A Schild plot analysis with linear regression resulted in slopes of 1.18 ± 0.18, 1.29 ± 0.23 and 1.05 ± 0.03 for bicuculline, gabazine and TMPP, respectively. The potency of antagonists was determined in terms of pA2 values. The pA2 values were 6.63 (gabazine), 6.21 (bicuculline), and 5.4 (TMPP). This suggests that gabazine has a higher binding affinity to the GABAA receptor than bicuculline and TMPP. Hence, using spike rate data obtained from population responses of spontaneously active neuronal networks, it is possible to determine key pharmacological properties of drug-receptor interactions.
The Developmental Physiology of the Zebrafish: Influence of Environment and Cardiovascular Attributes
Temperature effects on the development of the zebrafish embryos and larvae and adults were examined. It was found that the earlier in development a temperature change was performed on an embryo, the more significant the change in survival and/or subsequent development. Thus, viable temperature ranges for zebrafish widened significantly as development proceeded. Adults reared and bred at 25oC produced embryos that were significantly more successful at the lower range of rearing temperatures compared to embryos produced from adults reared at 28oC. The majority of this study focused on the physiological effects of swim training during development in the zebrafish. The earlier in development the zebrafish larvae were trained, the greater the mortality. Trained free swimming larvae had a significantly higher routine oxygen consumption after 11 days of training, and a higher mass specific routine metabolic rate after 8 and 11 days of training. Trained free swimming larvae consumed significantly less oxygen during swimming and were more efficient at locomotion, compared to control larvae. Training enhanced survival during exposure to extreme hypoxia in all age groups. Performance aspects of training were investigated in attempt to quantify training effects and in most cases, trained fish performed significantly better than controls. As blood vessels formed during development, they decreased in cross sectional area from days two to six. It was also shown that the variability in visual stroke volume measurements could be reduced significantly by using a third dimension in the analysis with a more accurate volume equation. Finally, the ontogeny of cardiac control was evaluated. The adrenergic receptors were the first to respond to pharmacological stimulation but were closely followed by cholinergic pharmacological stimulation a few days later. There was a significant cholinergic tone present in day 15 zebrafish larvae which persisted. Although an adrenergic tone was not documented in this study, …
Evaluation of City of Denton Sub-Watershed by Benthic Macroinvertebrate Field Experimental Approach
In this study, two different field experiments were designed to assess the relative influence of urbanization on benthic communities. During spring and summer, four urban and one reference sites from Denton County, Texas were selected for benthic macroinvertebrate evaluation. Statistically significant differences in colonized benthic macroinvertebrate taxa on artificial substrates were observed among the four urban sites and the reference site. Oligochaetes and chironomids were the dominant taxa at all sites. Identification of chironomid larvae at the subfamily and genus level to detect differences between sites had higher statistical power than the evaluation based on total chironomids. At the reference site, Caenis, Cladotanytarsus, Orthocladius, and Ceratopogonidae were the dominant taxa, while the urban sites were dominated by Dero, Physella, Ancylidae, Chironomus, Dicrotendipes, Glyptotendipes, Polypedilum, Pseudochironomus, Stenochironomus, and Tanytarsus. These differences may have been dependent upon differences in hydrologic regime and water quality between sites. Significant differences (ANOVA, p < 0.01) in water quality parameters (alkalinity, hardness, nitrates, phosphates, chlorides, sulfates, calcium, magnesium, potassium, and triazine) were found among water samples collected from the reference and urban sites. During the transfer period, most of the Ephemeroptera and Trichoptera taxa and a few other taxa disappeared from artificial substrates that were colonized at the reference site and then moved to the urban sites. Also, local abundant taxa from the urban site significantly (t test, p < 0.05) increased in number on the transferred artificial substrates. Seasonal differences in colonization patterns were also observed between the spring and summer experimental periods, which indicate that temporal variation is equally important, as is the anthropogenic effect in benthic community evaluation. Field survival and growth experiments using Erpetogomphus designatus larvae were designed to detect differences between evaluated sites. Larvae were collected from the reference site, measured in the laboratory, and exposed at the urban sites for …
Evaluation of virulence in wild type and pyrimidine auxotrophs of Pseudomonas aeruginosa using the eukaryotic model system Caenorhabditis elegans.
The human opportunistic pathogen, Pseudomonas aeruginosa PAO1, has been shown to kill the nematode Caenorhabditis elegans. C. elegans has been a valuable model for the study of bacterial pathogenesis, and has reinforced the notion that common virulence and host defense mechanisms exist. Recently, the pyrimidine pathway was shown to regulate virulence levels. Therefore, mutations in the pyrimidine pathway of PAO1 showed decrease virulence in the nematode. When starving the nematode, bacterial resistance was also shown to increase. It was hypothesized that starvation induced the DAF pathway, which regulates the transcription of genes involved with the antibacterial defense mechanism. Further research will be conducted to test this theory by performing RNAi experiments for the genes functioning in the antibacterial defense mechanism.
Evidentiary Value of Condoms: Comparison of Durable Physical and Chemical Characteristics of Condoms
Condom trace evidence must not be overlooked in sexual assault cases; understanding the chemical and physical characteristics of condoms is imperative if condoms are to be useful evidence. Previous research shows that condom identification is possible, but it is equally important to evaluate durability of condom residues versus time. Using FT-IR, this study examined vaginal swabs from subjects who self-sampled at intervals for up to 72 hours after having intercourse with a condom. This study investigated whether age and the stage of the menstrual cycle affected the durability of residues in the vagina over time. This study revealed that condoms containing nonoxynol-9, silicone-based lubricants, and particulates provide valuable information for identification, and that nonoxynol-9 specifically withstands the vaginal environment for up to 72 hours. Additionally, age and menstrual cycle both appeared to have an effect on the durability of residues although larger sample size is desirable.
Inquiry-based science for high school students: a forensic unit
This project constitutes an instructional unit for honors biology that involves the use of science in the field of criminal investigation and forensics. Before beginning the unit, the learners should have mastered basic laboratory skills, including use of the microscope. They should also have an understanding of the basic structure and function of DNA and its role in heredity and protein synthesis. The standard time frame is 24 days with 70-minute periods, but can be easily adjusted to meet classroom needs. Several instructional strategies enhance student learning and make science fun. The unit is inquiry-driven and activity-based. Students are surprised by the crime, gather and analyze evidence, and work towards proposing an explanation. This real world problem involves the use of cooperative learning and a variety of assessment techniques.
A laboratory course in experimental genetics for the biology major.
This manual has been designed for a class of twenty- four students concurrently enrolled in the lecture course. The laboratory aids in the learning process and fosters an interest in the science of genetics. This manual and the experiments contained within are both informative and fun. The manual correlates with and expands upon the genetics course. Each investigation, with the exception of the Drosophila melanogaster project, can be completed in a 3-4 hour timeframe. This manual provides a “hands on” experience of theories simply discussed in the lecture course. This manual is intended to be a one-source manual where each investigation is designed to include an adequate introduction. Special attention has been given for each investigation with both the student and instructor in mind.
Life History and Case Building Behaviors of Phylloicus ornatus (Banks) (Trichoptera: Calamoceratidae) In Two Spring Fed Tributaries in the Central Edwards Plateau Bioregion of Texas
The life history and case-making behaviors of Phylloicus ornatus from two springfed first order streams in the Edwards Plateau Bioregion of Texas were studied from January 1998 to November 1999. Field larval, pupal and adult samples and laboratory rearings indicated a multivoltine cycle. First instars differ from late instars in number of labral setae and in having a unique spur-like claw on each lateral hump. Larval development was asynchronous with second through fifth instars and pupae present most months. First instars were present April through July, October and November. Case making of first instar and case reconstruction of later instars extracted from their cases was documented by videophotography.
Macroinvertebrate Community Structure as an Indicator of Watershed Health in the Upper Trinity River Basin, North Central Texas
This study describes macroinvertebrate community structure and assesses its potential in detecting point and non-point sources of disturbance associated with rural and urban areas in the Upper Trinity River Basin. Geospatial techniques were used to quantify landuse within the watershed in a GIS. At rural sites near the headwaters of the Trinity River, collector-gathering burrowers that are adapted to minimal flow comprised the majority of taxa. Destinies of taxa compositions at downstream sites increased and shifted toward psammophilic and rheophilic invertebrates, including primarily collector-filtering clingers, that are characteristic of shifting sand habitats in large prairie rivers. Benthic community structure generally benefited from point source impacts including wastewater treatment plant effluents that maintained higher flow. Community indices were negatively associated with forest landuse and positively associated with urban landuse. Partial CCA determined that flow and landuse contributed equally to species dispersions. Comparisons with historical biomonitoring studies in upper Trinity River Basin indicate improved watershed health.
The Metabolic Physiology of Planarian Flatworms
Using a high throughput closed respirometry method to measure oxygen consumption, I determined metabolic rates in asexual and sexual Schmidtea mediterranea and Girardia dorotocephala, as a function of temperature, taxon, stressors, reproductive mode, age, regeneration, and specific dynamic action. This study has shown that oxygen consumption can reliably be measured in planaria using optode closed respirometry, and also provided a reliable method for measuring wet mass in planaria, which has been a challenge to researchers in the past. This research revealed that oxygen consumption in S. mediterranea is 1.5-2.1X greater in the sexual strain over the asexual strain at 13-18°C. Within the sexual strain, oxygen consumption is 1.5 -2.2X greater in sexually mature adults over the sexually immature groups (hatchlings, juveniles, and regenerating sexuals). Furthermore, I was able to quantify differences in sexual morphology between these groups exhibiting significant differences in oxygen consumption. The results of this research supports a theory of higher metabolic costs with sexual maturity in S. mediterranea. Therefore, this study has established sexual and asexual S. mediterranea as simple, yet attractive models for investigating energetic costs between sexual and asexual phenotypes. This research also provided quantitative values for specific dynamic action in planaria, with a maximum increase in oxygen consumption of 160% induced by feeding, as well as metabolic relationships in planaria involving temperature, age, and regeneration. These values establish planaria as one of the simplest animal models in which common metabolic patterns, such as SDA and poikilothermic temperature sensitivity, have been demonstrated. Therefore, this research has contributed to the overall knowledge of the basic physiology in this animal, providing the framework for future metabolic studies in planaria involving environmental factors, reproduction, regeneration, development, and aging. Information from this study may supplement interpretation and understanding of modern cellular, molecular, and genomic studies in planaria.
A morphological study of the avian (Gallus domesticus) ductus arteriosi during hatching.
The ductus arteriosi (DA) are two blood vessels connecting the pulmonary arteries to the descending aorta in the avian embryo. Following hatching, the DA closes, separation of the systemic and pulmonary circulation. I present the morphological changes that occur in the chicken DA during prepipping, internal pipping, external pipping, and hatching. The avian DA consists of two distinct tissue types, a proximal and a distal portion. Histological examination shows developmental differences between the proximal and distal portions of the DA with regard to lumen occlusion, endothelial cells, smooth muscle and elastin. Endothelial cell proliferation begins to occur as early as external pipping, with the lumen almost completely occluded by the 3rd day of post-hatching life. Expression of vascular endothelial growth factor (VEGF) increases in avian endothelial cells during hatching. I provide a morphological timeline of changes in the DA as the chicken develops from embryo to hatchling.
A Multimedia Atlas of Dissection for Comparative Anatomy of the Vertebrates
Traditional methods of teaching the laboratory course for Comparative Anatomy of the Vertebrates could be improved by applying current computer technology to construct an interactive, multimedial atlas of dissection. Five specimens used in comparative anatomy courses at most institutions were chosen as representative members of the Phylum Chordata: amphioxus, lamprey, dogfish shark, mud puppy, and cat. Specimens were dissected according to the modified method of Wischnitzer, 1993, and each stage was photographed with a Kodak DC120 digital zoom camera. These images were processed on a Power Macintosh 7600 computer with Adobe Photoshop v. 5.0. The atlas was constructed from these images using Macromedia Authorware v. 4.0.3. Each image contains a series of interactive objects that display a highlight and descriptive text as the cursor passes over each object.
A Novel Phytoestrogen that Acts as an Agonist for Human Estrogen Receptors.
Estrogen is the natural agonist of the estrogen receptor (ER). However, certain plant-derived compounds or phytoestrogens have been identified that mimic estrogens and act as agonists and/or antagonists of ERs, depending on subtype and target tissue. Understanding how phytoestrogens interact with ERs, and therefore effect the estrogenic response, may prove beneficial in hormone replacement therapy and in the prevention and treatment of hormone-related diseases. Using Thin Layer Chromatography, gas chromatography/mass spectrometry (GC/MS), and proton nuclear nagnetic resonance (HNMR), I identified 4-ethoxymethylphenol (4EM) found in Maclura pomifera. While most phytoestrogens are heterocyclic compounds, 4EM is a simple phenol that acts as an agonist of ER-alpha and -beta in HeLa and MCF-7 cells. To study the effect of 4EM on ER-alpha and -beta activity, I performed transient transfection assays and showed that 4EM activates ER dependent gene transcription in a dose dependent manner in both ER subtypes. Further, 4EM- mediated transcription in ER-alpha, like estrogen, was enhance in the presense of co-activators, SRC-1 (steroid receptor coactivator-1), CBP (CREB binding proteins), and E6-AP (E6-associated protein) and inhibited by trans-4- hydroxytamoxifen (4HT). I found that 4EM was specific for ER and did not activate transcription of the progesterone receptor in HeLa cells.
Phenotype Analysis of the CISD Gene Family Relative to Mitochondrial Function in Caenorhabditis elegans
NEET proteins belong to a unique class of [2Fe-2S] cluster proteins that have been shown to participate in various biological processes such as regulating iron, reactive oxygen species and apoptosis within the cell and are localized to the mitochondria. Disruption of the mitochondrial NEET proteins are associated with different human diseases such as obesity, neurodegeneration, cancer and diabetes. In humans, a missense mutation in the CISD2 gene results in a heritable multisystem disorder termed Wolfram syndrome 2 (WFS2), a disease which displays an early onset of juvenile diabetes and various neuropsychiatric disorders. The C. elegans genome contains three previously uncharacterized cisd genes: cisd-1, which has homology to the human CISD1 and CISD2, and cisd-3.1 and cisd-3.2, both of which have homology to the human CISD3. Disruption of the cisd-3 gene(s) function results in mis-regulation of proteostasis in the mitochondria, whereas cisd-1 and cisd-3.1 disruption impacts proteostasis in the endoplasmic reticulum. Reduction of cisd-3.2 gene function also leads to a developmental delay in C. elegans. A knockout mutation of the cisd-3.2(pn68) gene function results in various germline defects including delayed development progression and morphological defects. Furthermore, I show the cisd gene(s) and protein expression profiles is present relative to sex, tissue type and developmental stages. This work is significant because it provides further insight of the essential role of CISD-3 relative to C. elegans. Furthermore, my studies can contribute to new genetic discoveries that will widen the scientific research relative to NEET protein family studies.
Production and Characterization of a Novel Extracellular Polysaccharide Produced by Paenibacillus velaei, Sp. Nov
Paenibacillus velaei, sp. nov. is a soil bacterium capable of producing an unusually large amount of exopolysaccharide (EPS). The EPS contains glucose, mannose, galactose and fucose in a molar ratio of 4:2:1:1. The molecular weight of the EPS is higher than 2x106. The viscosity of 1% EPS is 1300 cP when measured at a shear rate of 1 sec-1. Physiological parameters for optimal production of the EPS were studied and it was found that 1.4 g dry weight per 1 l of medium was produced when the bacteria were grown at 30EC and the pH adjusted at 7± 0.2 in a medium containing glucose as the carbon source. Growing the bacteria on different carbon sources did not alter the quantity or the composition of the EPS produced. No toxicity effects were observed in mice or rats when EPS was administered in amounts ranging from 20 to 200 mg per kg body weight. The data obtained from physical, chemical and biological properties suggest that the EPS may be employed in several industrial and environmental applications. It is an excellent emulsifier, it holds 100 times its own weight in water, it is not toxic, and it can be used to remove mercury, cadmium and lead from aqueous solutions.
Pyrimidine Enzyme Specific Activity at Four Different Phases of Growth in Minimal and Rich Media, and Concomitant Virulence Factors Evaluation in Pseudomonas aeruginosa
Pseudomonas aeruginosa is a Gram-negative rod, aerobic, non-fermenting, oxidase positive, pigment producing, and nutritionally versatile bacterium. Infections by P. aeruginosa are the most important cause of morbidity and mortality in immunocompromised patients, given virulence factor production that suppresses antibiotic therapy and promotes persistent infection. This research is the first comprehensive report of the pyrimidine biosynthetic pathway for all phases of growth in minimal and rich media coupled with the evaluation of virulence factor production of P. aeruginosa in comparison to four other bacterial species (Pseudomonas putida, Pseudomonas fluorescens, Burkholderia cepacia, and Escherichia coli wild-type strains). Cellular growth and passing genetic information to the next generation depend on the synthesis of purines and pyrimidines, the precursors of DNA and RNA. The pyrimidine biosynthetic pathway is essential and found in most organisms, with the exception of a few parasites that depend upon the pyrimidine salvage pathway for growth. Both the pyrimidine biosynthetic and salvage enzymes are targets for chemotherapeutic agents. In our laboratory, research on pyrimidine auxotrophic mutants showed the role of the pyrimidine biosynthetic pathway and its intermediates on P. aeruginosa metabolism and impaired virulence factors production. The present research shows that pyrimidine enzymes are active in all phases of growth, including the production of two forms of ATCase in the late log phase in P. aeruginosa. This finding may be explained by the displacement of the inactive PyrC' by the active PyrC or PyrC2 to form a new and larger pyrBC encoded ATCase. Pseudomonas aeruginosa wild-type appears to produce by far the most virulence factors, haemolysin, iron chelation, rhamnolipid, adherence, and three types of motility (swimming, swarming, and twitching) investigated in this study, when compared to the other four wild-type strains. Growth analysis was carried out as typically done in minimal medium but also in rich medium to simulate conditions …
Use of Geographic Information System and Remote Sensing Technologies to Describe Mosquito Population Dynamics in the Ray Roberts Greenbelt, Denton County, Texas
A population survey was conducted from April through September 2002 on mosquito species occurring on the Ray Roberts Greenbelt, a riparian corridor used for public recreation on the Elm Fork of the Trinity River, in Denton County, Texas. ArcGIS software was used to set up a stratified random sampling design based on habitat parameters. Multivariate analyses of sampling data and climatic variables were used to describe spatial and temporal patterns of mosquito species. A total of 33 species were collected during this study belonging to the following genera: Aedes, Anopheles, Coquillettidia, Culex, Mansonia, Ochlerotatus, Orthopodomyia, Psorophora, Toxorhynchites, and Uranotaenia. Seasonal distributions of the dominant species revealed population fluctuations. Aedes vexans was the primary species collected in April and May, occurring in low numbers throughout the rest of the sampling period. Psorophora columbiae reached its highest population density in June, with a smaller peak occurring in late July. Present from May through the end of September, Culex erraticus was the most abundant species collected with major peaks in mid-June and the end of July. Abundance of Culex salinarius followed the same general trend as that for Cx. erraticus, but with smaller numbers. The specimens were tested for a variety of arboviruses by the Texas Department of Health. One pool of Cx. erraticus and Cx. salinarius, collected in August 2002, tested positive for West Nile virus. Variables that were important factors for determining dominant species abundance were temperature, wind speed, rain accumulation occurring one-week and two-weeks prior to sampling, number of day since last rain event, dew point, and average canopy coverage.
Use of Satellite Imagery and GIS to Model Brood-Rearing Habitat for Rio Grande Wild Turkey Populations Occurring in the Western Cross Timbers Region of Texas
Remote sensing and GIS have become standard tools for evaluating spatial components of wildlife habitats. These techniques were implemented to evaluate Rio Grande wild turkey (Meleagris gallopavo) poult-rearing habitat in the Western Cross Timbers region of Texas. Texas Parks and Wildlife (TPWD) random roving turkey counts for 1987-1989 and 1998-2000 were selected, indicating locations where hens with poults were observed. Satellite imagery from 1988 and 1999 was classified and then processed with Patch Analyst. To add robustness, stream, road and census population densities were also evaluated for each turkey location. Analysis of the 1988 canopy cover image, comparing observed locations with randomly-selected habitat cells (N = 20) indicated significant differences (p <.05) for patch edge variables. Mean patch edge was significantly greater for habitat locations where hens with poults were observed than for those selected at random. Spatial data for 1999 did not indicate a significant difference (p < .05) between sampling groups (observed vs. random, N = 30). Significant differences (p <.05) did occur for turkey locations observed in both 1988 and 1999 (N = 7). This demonstrates the adaptability of wild turkey hens, as habitats change over time, hens continued to visit the same locations even though the habitat had significantly changed for select spatial variables.
Utilization of Corridor Habitat by White-tailed Deer (Odocoileus virginianus) in Denton County, Texas
White-tailed deer (Odocoileus virginianus) (N=15) movements were determined with use of radio telemetry techniques to determine the utilization of corridor habitat on the Lake Ray Roberts Greenbelt Corridor (RRGC) in north central Texas. Home ranges were calculated using three estimation types. Male white-tailed deer tend to have home ranges twice that of female home ranges. Seasonal home ranges were largest during spring (Feb. - April) and fall (Aug. - Oct.) seasons. Males had greater seasonal variation in utilization than females. No statistically significant difference (p=0.24) between white-tailed deer locations when the RRGC experiences heavy human traffic compared to days when there is light human traffic. Linearity indices indicated home ranges less linear than expected (LI = 3.02). The RRGC should be maintained at its current status to provide a variety of vegetational types and protective cover for white-tailed deer and other wildlife of Denton County.
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