This study analyzes the remaining suitable habitat of the one-horned rhinoceros, Rhinoceros unicornis, in Royal Chitwan National Park of Nepal. An April 2003 Landsat image was classified into eight land cover types: wetland, sand, water, mixed forest, sal forest, agriculture, settlement, and grassland. This image was converted into habitat suitability maps using cover, food, and water. The rhinoceros prefers grassland habitat with oxbow lakes and closed canopy during the monsoon season. Nominal values of five parameters were used to create a map of habitat suitability index. The map was categorized into four habitat classes: highly unsuitable, unsuitable, moderately suitable habitat, and suitable. Landscape metrics, patch metrics and class metrics associated with habitat were determined through the use of FRAGSTATS.
During the spring 2004 a pilot school recycling program was implemented within Robert E. Lee Elementary. The primary goal of the program was to determine how recycling education in the school would affect curbside recycling rates within the surrounding community. The program was a cooperative effort between the University of North Texas, City of Denton Solid Waste Department and Keep Denton Beautiful. Throughout the first months of the study during the spring 2004, an increase in curbside recycling within the Robert E. Lee Elementary attendance zone was observed, with a dramatic decrease in participation over the summer and a rapid increase once again during the second full semester of the study. In a survey conducted with 3rd and 5th grade students at the pilot project school, most students expressed positive attitudes about recycling. Students whose survey responses indicated a high level of knowledge about what could be recycled were 37% more likely to claim to recycle regularly, than those students that scored low on the knowledge portion of the survey. Although the total amount of waste generation (recyclable and non-recyclable) at Robert E. Lee Elementary did not decrease during the study, the campus was able to divert recyclable material from their trash at a much higher rate than two other local elementary campuses with paper-only recycling and no associated recycling education program. Based upon the success of the recycling program at Robert E. Lee Elementary, the City of Denton Recycling Division has agreed to move forward with offering recycling to more schools within the Denton Independent School District during the 2005-2006 school year.
In response to compatible rhizobia, leguminous plants develop unique plant organs, root nodules, in which rhizobia fix nitrogen into ammonia. During nodule invasion, the rhizobia gain access to newly divided cells, the nodule primordia, in the root inner cortex through plant-derived cellulose tubes called infection threads. Infection threads begin in curled root hairs and bring rhizobia into the root crossing several cell layers in the process. Ultimately the rhizobia are deposited within nodule primordium cells through a process resembling endocytosis. Plant host mechanisms underlying the formation and regulation of the invasion process are not understood. To identify and clone plant genes required for nodule invasion, recent efforts have focused on Medicago truncatula. In a collaborative effort the nodulation defect in the lin (lumpy infections) mutant was characterized. From an EMS-mutagenized population of M. truncatula, two non-allelic mutants nip (numerous infections with polyphenolics) and sli (sluggish infections) were identified with defects in nodule invasion. Infection threads were found to proliferate abnormally in the nip mutant nodules with only very rare deposition of rhizobia within plant host cells. nip nodules were found to accumulate polyphenolic compounds, indicative of a host defense response. Interestingly, nip was also found to have defective lateral root elongation suggesting that NIP has a role in both nodule and lateral root development. NIP was found to map at the upper arm of chromosome 1. In sli, infection threads were observed to bring rhizobia from infection threads to newly divided nodule primordium cells in the roots inner cortex. Polyphenolic accumulation in sli nodule/bumps was found. Lateral roots in sli were found to be clustered at the top of the root, indicating that sli like nip may be defective in lateral root development.
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 Rocker b Ranch on the southern Rolling Plains has one of the last sizeable populations of pronghorn (Antilocapra americana) in Texas. To investigate habitat utilization on the ranch, pronghorn were fitted with GPS/VHF collars and were released into pastures surrounded by a variety of fences to determine how fence types affected habitat selection. Habitat parameters chosen for analysis were vegetation, elevation, slope, aspect, and distances to water, roads, and oil wells. Results showed that pronghorn on the ranch crossed modified fencing significantly less than other types of fencing. Pronghorn selected for all habitat parameters to various degrees, with the most important being vegetation type. Habitat selection could be attributed to correspondence of vegetation type with other parameters or spatial arrangements of physical features of the landscape. Seasonal differences in habitat utilization were evident, and animals tended to move shorter distances at night than they did during daylight hours.
Effects of episodic exposures have gained attention as the regulatory focus of the Clean Water Act has shifted away from continuous-flow effluents. Standardized laboratory toxicity tests require that exposure be held constant. However, this approach may not accurately predict organism responses in the field following episodic exposures such as those associated with rain-driven runoff events or accidental pollutant discharge. Using a modified version of the 7-day short-term chronic test recommended by the US Environmental Protection Agency, Ceriodaphnia dubia were exposed to copper or cadmium for durations ranging from 1 minute to 24 hours. In addition, adult reproductive recovery and effects on second generation individuals was assessed following select copper exposures. Finally, cadmium exposures were compared in reconstituted hard water (RHW) and municipal treated wastewater effluent (TWE). Following exposure, organisms were transferred to clean RHW or TWE and maintained for the remainder of the test. No- and lowest observed effect concentrations (NO- and LOECs) increased logarithmically with respect to logarithmic decreases in duration regardless of metal, endpoint or water type. Effective concentrations of cadmium however, were usually higher than those of copper, especially in TWE. LOECs for C. dubia survival following 24-hour and 5-minute exposures to copper were 116 and 417 µg/L, respectively. LOECs for fecundity were 58 and 374 µg/L, respectively. Neonate production of first generation adult C. dubia appeared to recover from pulsed copper exposure upon examination of individual broods. Cumulative mean neonate production however, showed almost no signs of recovery at exposure durations ≥3 hours. Pulse exposure to copper also resulted in diminished fecundity of unexposed second generation individuals. Such effects were pronounced following parental exposure for 24 hours but lacking after parental exposures ≤3 hours. LOECs for C. dubia survival following 24-hour and 5-minute exposures to cadmium in RHW were 44 and 9000 µg/L, respectively. LOECs for …
Dissociated neuronal cell cultures grown on substrate integrated microelectrode arrays (MEAs) generate spontaneous activity that can be recorded for up to several weeks. The signature wave shapes from extracellular recording of neuronal activity display a great variety of shapes with triphasic signals predominating. I characterized extracellular recordings from over 600 neuronal signals. I have preformed a categorical study by dividing wave shapes into two major classes: (type 1) signals in which the large positive peak follows the negative spike, and (type 2) signals in which the large positive peak precedes the negative spike. The former are hypothesized to be active signal propagation that can occur in the axon and possibly in soma or dendrites. The latter are hypothesized to be passive which is generally secluded to soma or dendrites. In order to verify these hypotheses, I pharmacologically targeted ion channels with tetrodotoxin (TTX), tetraethylammonium (TEA), 4-aminopyridine (4-AP), and monensin.
The industrial uses of cottonseed oil are limited by its fatty acid composition. Genetic modification of cotton lipid profiles using seed-specific promoters could allow cotton growers to produce valuable new oils in the seed without adverse effects on fiber quality and yield, therefore making this crop more commercially profitable. Transgenic cotton callus harboring a diverged fatty acid desaturase gene (FADX) from Momordica charantia was characterized for production of alpha-eleostearic acid (conjugated double bonds: 18:3 D9 cis, 11 trans, 13 trans), not normally found in cotton. Gas chromatography (GC) in conjunction with mass spectrometry (MS) confirmed production of alpha-eleostearic acid in the transgenic cotton tissues. A second series of transformation experiments introduced the cotton fatty acid thioesterase B (FATB) cDNA, fused to the seed-specific oleosin promoter into cotton to promote the over-expression of FATB, to generate cotton with increased palmitate in the cottonseed. PCR amplification, as well as fatty acid analysis by gas chromatography, confirmed introduction of the FATB cDNA in transgenic tissues. Collectively, these results demonstrate the feasibility of manipulating the fatty acid composition in cotton via transgenic approaches and form the basis for continued efforts to create novel oils in cottonseed.
Previous research in our laboratory showed that when knockout mutations were created in the pyrB and pyrC genes of the pyrimidine pathway in Pseudomonas aeruginosa, not only were the resultant mutants auxotrophic for pyrimidines but they were also impaired in virulence factor production. Such a correlation had not been previously reported for P. aeruginosa, a ubiquitous opportunistic pathogen in humans. In an earlier study it was reported that mutants blocked in one of the first three enzymes of the pyrimidine pathway in the non-pathogenic strain P. putida M produced no pyoverdin pigment while mutants blocked in the later steps produced copious amounts of pigment, just like the wild type. This study probed for the same connection between pyrimidine auxotrophy and pigment production applied in P. aeruginosa. To that end a knockout mutation was created in pyrD, the fourth step in the pyrimidine pathway which encodes dihydroorotate dehydrogenase. The resulting mutant required pyrimidines for growth but produced wild type pigment levels. Since the pigment pyoverdin is a siderophore it may also be considered a virulence factor, other virulence factors were quantified in the mutant. These included casein protease, hemolysin, elastase, swimming, swarming and twitching motility, and iron binding capacity. In all cases these virulence factors were significantly decreased in the mutant. Even supplementing with uracil did not attain wild type levels. Starvation of the pyrimidine mutant for uracil caused increased specific activity of the pyrimidine enzymes, suggesting that regulation of the pyrimidine pathway occurred at the level of transcription. This effect has also been reported for P. oleovorans. The present research consolidates the idea that pyrimidine auxotrophs cause decreased pathogenicity in P. aeruginosa. Such a finding may open the search for chemotherapy targets in cystic fibrosis and burn victims where P. aeruginosa is an infecting agent.
Methods to determine genetic diversity and relatedness within populations are essential tools for proper wildlife management. Today the approach of choice is polymerase chain reaction-based microsatellite analysis. Seven new polymorphic loci were isolated from a microsatellite-enriched Caribbean flamingo genomic library and used to characterize survey populations of Caribbean and African greater flamingos. In addition, four of these loci were used to verify parentage relationships within a captive-breeding population of African greater flamingos. Parentage predictions based upon gamekeeper observations of breeding and nesting did not always agree with genetic-based parentage analyses of the nine suggested family groups. Four family groups were supported (groups I, II, III and VI) by there results. However, an analysis of the remaining five suggested groups, with a total of eight offspring/dam and eight offspring/sire suggested relationships, yielded seven exclusions of the suggested dam and six exclusions of the suggested sire. This put the overall suggested dam exclusion rate at 35% and exclusion rate for suggested sires at 29%. Although the keeper observation data for our family groups must be considered a variable of concern at this time, these findings are certainly suggestive that more carefully controlled studies may reveal that flamingos are not monogamous as long accepted, but rather socially monogamous or even promiscuous. Thus we have now been able to both characterize and demonstrate the utility of our polymorphic microsatellite loci. We hope these results will interest additional wildlife facilities in further parentage and behavioral studies that will collectively aid to improve monitoring and maintenance of genetic diversity, and as provide better insight into breeding habits of both wild and captive populations.
A study of the life history and ecology of Camelobaetidius variabilis was conducted in Honey Creek, OK from February 2003-April 2004. Nymph development was assessed using changes in external morphology. Laboratory reared nymphs were used to calculate number of degree days to complete development (772 degree days at 20.8° C ±.38° C), which was used to determine voltinism. Field collected nymph microhabitat distribution was used in assessing microhabitat distribution. Nymphal thermoregulation was assessed during the winter and spring by comparing nymphal numbers present in shaded and un-shaded habitats. Camelobaetidius variabilis nymphs showed preference for algal microhabitats during the spring and leaf packs in the winter. Nymphs inhabited leaf packs to increase metabolic rate during the winter. Increased temperatures aid in development of nymphs. Camelobaetidius variabilis exhibited a multivoltine life cycle with six overlapping generations.
Flamingo species generate tremendous interest whether they are small captive groups or wild populations numbering in the thousands. Genetic pedigrees are invaluable for maintaining maximum genetic diversity in captive, as well as wild, populations. However, presently there is a general lack of genetic data for flamingo populations. Microsatellites are loci composed of 2-6 base pair tandem repeats, scattered throughout higher eukaryotic genomes, often exhibiting high levels of polymorphism and heterozygosity. These loci are thus important genetic markers for identity, parentage and population studies. Here, six microsatellite loci were isolated from a microsatellite-enriched Caribbean flamingo partial genomic library. Two are compound complex repeats and four are perfect trinucleotide repeats. Each locus was amplified from Caribbean, African greater, Chilean and lesser flamingo genomic DNAs. Heterozygosity frequencies were calculated for Caribbean (range 0.12-0.90) and African greater flamingos (range 0.23-0.94) loci. All six microsatellite loci were found to be in Hardy-Weinberg equilibrium and linkage disequilibrium analyses did not suggest linkage for any pair of two greater flamingo subspecies (African and Caribbean) loci. At least five of the loci also exhibit polymorphism in Chilean and lesser flamingos, but due to small sample numbers, relevant allele/heterozygosity frequency calculations could not be estimated. Nucleotide sequence comparisons of the amplicons derived from the four flamingo groups reveal a high level of sequence conservation at all loci. Although small sample numbers again limit the data for lesser flamingos and to some degree for the Chilean birds, the sequences of the two greater flamingo subspecies were identical and the number of nonconserved nucleotides appears to be higher for lesser/greater comparisons than for Chilean/greater comparisons. This is consistent with Chilean flamingos being a different species within the same genus as the greater flamingos, while lesser flamingos belong to a separate genus. Parentage analyses on suggested African greater flamingo family groups from …
N-Acylethanolamines (NAEs) are enriched in seed-derived tissues and are believed to be formed from the membrane phospholipid, N-acylphosphatidylethanolamine (NAPE) via the action of phospholipase D (PLD). In an effort to identify a functional NAPE-PLD in cotton seeds and seedlings, we have screened a cotton seedling cDNA (cotyledon mRNA from 48 h dark grown seedlings) library with a 1.2 kb tobacco partial cDNA fragment encoding the middle third of a putative PLDβ/γ (genbank accession, AF195614) isoform. Six plaques were isolated from the Uni-ZAP lambda library, excised as pBluescript SK(-) phagemids and subjected to nucleotide sequence analysis. Alignment of derived sequences with Arabidopsis PLD family members indicated that the cDNAs represent six different PLD gene products -three putative PLD β isoforms and three putative PLD δ isoforms. The PLD β isoforms, designated Ghpldβ1a, GHpldβ1b and a truncated Ghpldβ1b isoform. Both the full-length PLD β proteins contained characteristic HKxxxxD catalytic domains, a PC-binding domain, a PIP2-binding domain and a C2 domain. In addition both cotton PLD β isoforms had a N-terminal "SPQY" rich domain which appeared to be unique to these PLDs. The three PLD δ isoforms, designated Ghpldδ1a, Ghpldδ1b and Ghpldδ1b-2 encode full-length PLDδ proteins, and like the above PLDs, contained the characteristic catalytic and regulatory domains. The expression of Ghpldδ1b showed hydrolytic and transphosphatidylation activity toward radiolabelled phosphatidylcholine (PC) but it appears Ghpldδ1b does not utilize NAPE as a substrate to produce NAEs nor does it seem to be suppressed by NAEs.
The development of cholinergic neurons in mouse frontal cortex was studied both in vivo and in vitro by immunocytochemistry with an antibody to choline acetyltransferase (ChAT), the enzyme responsible for acetylcholine synthesis. While cortical cholinergic neurons have previously been characterized in rat cortex, up until very recently, intrinsic cortical cholinergic neurons were considered to be absent in mouse, and little is known about their development or phenotypic characteristics. The present study found no ChAT-positive neurons in mouse frontal cortex on postnatal day 0 (P0, the day of birth). On P7 there were few, faintly stained, ChAT-positive neurons. The numerical density of ChAT-positive neurons increased substantially with age, from none on P0, to 9.2 + 1.4 on P7, to 14.8 + 0.9 on P16, and 41.6 + 3.9 in adulthood. Considering that the numerical density of total neurons decreases during this postnatal period, the data represent a marked developmental increase in the percentage of cholinergic neurons. The development of cholinergic neurons showed very similar timelines in rat and mouse frontal cortex. Cultures prepared from mouse frontal cortex on embryonic day 16 were maintained for 25, 76, or 100 days in vitro (div). The percentage of ChAT-positive neurons was considerably higher than in vivo, ranging from a mean 28% to 31% across the three age (div) groups. With increasing age of the cultures, the numerical density of total neurons and ChAT-positive neurons decreased while the percentage of ChAT-positive neurons did not change significantly. These observations suggest some temporal stability in the cultures. Using dual immunofluorescence, ChAT-positive neurons were tested for colocalization with GAD or TH. The majority of ChAT-positive neurons colocalized with GAD, both in vitro and in vivo. However, ChAT did not colocalize with TH, either in vitro or in vivo. Our comparison of intact frontal cortex and cultures suggest that …
Primary cilia are non-motile organelles having 9+0 microtubules that project from the basal body of the cell. While the main purpose of motile cilia in mammalian cells is to move fluid or mucus over the cell surface, the purpose of primary cilia has remained elusive for the most part. Primary cilia are shortened in the kidney tubules of Tg737orpk mice, which have polycystic kidney disease due to ciliary defects. The product of the Tg737 gene is polaris, which is directly involved in a microtubule-dependent transport process called intraflagellar transport (IFT). In order to determine the importance of polaris in the development of neuronal cilia, cilium length and numerical density of cilia were quantitatively assessed in six different brain regions on postnatal days 14 and 31 in Tg737orpk mutant and wildtype mice. Our results indicate that the polaris mutation leads to shortening of cilia as well as decreased percentage of ciliated neurons in all brain regions that were quantitatively assessed. Maintainance of cilia was especially affected in the ventromedial nucleus of the hypothalamus. Furthermore, the polaris mutation curtailed cilium length more severely on postnatal day 31 than postnatal day 14. These data suggests that even after ciliogenesis, intraflagellar transport is necessary in order to maintain neuronal cilia. Regional heterogeneity in the effect of this gene mutation on neuronal cilia suggests that the functions of some brain regions might be more compromised than others.
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 …
Hydrologic models were used to examine the effects of land cover change on the flow regime of a watershed located in North-Central Texas. Additionally, the effect of spatial resolution was examined by conducting the simulations using sub-watersheds of different sizes to account for the watershed. Using the Army Corps of Engineers, Hydrologic Engineering Center Hydrologic Modeling System (HEC-HMS), two different modeling methods were evaluated at the different sub-watershed resolutions for four rainfall events. Calibration results indicate using the smaller spatial resolutions improves the model results. Different scenarios for land cover change were evaluated for all resolutions using both models. As land cover change increased, the amount of flow from the watershed increased.
Managing water quality aspects requires the use of integrative tools that allow a holistic approach to this problem. Water quality models coupled to hydrodynamic models are these tools. This study presents the application of the water quality model WASP coupled to the hydrodynamic model DYNHYD for two distinct reservoirs: Lake Texoma and Tocoma Reservoir. Modeling the former included simulations of water velocities, water level, and four chemical and physical compounds: chlorides, dissolved oxygen (DO), biochemical oxygen demand (BOD), and total suspended solids (TSS); and validation of the results by comparing with observed values during March - May, 1997. The latter is still under project status and the simulation was performed in a prospective way. The analysis included simulations of water velocities under current and for expected conditions, DO and BOD. Both models, DYNHYD and WASP, fitted pretty well to observed conditions for Lake Texoma and for where Tocoma Reservoir has been planned. Considering management and decision support purposes, the role of boundary and loading conditions also was tested. For Lake Texoma, controlling boundary conditions for chlorides is a determinant factor for water quality of the system. However, DO and TSS in the reservoir are governed by additional process besides the condition of the boundary. Estimated loadings for this system did not provided significant effects, even though the allocation of a load for chlorides resulted in significant changes in the trend for expected chloride concentrations at the Washita River Arm of Lake Texoma. For Tocoma Reservoir, the expected concentration of DO all over the reservoir is going to driven by boundary conditions, as well as by the management of autochthonous BOD loadings provided by vegetation decomposition. These two factors will be determinant for the resulting water quality of the future reservoir.
The stonefly genus Isogenoides is revised following a holomorphological approach utilizing traditional morphology and behavioral lines of evidence. Species keys are provided for all life stages. One species, I. krumholzi (Ricker) is considered a synonym of I. doratus (Frison). Detailed species descriptions are provided for males, females, nymphs and ova. Distributions are updated utilizing all known published accounts and materials examined. The vibrational communication (drumming) behavior is reported for males and all but one species for females. The signals were species-specific and ranged in complexity from ancestral sequenced duets to derived grouped exchanges. I. olivaceus is least specialized, having mostly sequenced duets, and I. zionensis most specialized, displaying ancestral sequenced, derived grouped and complex derived exchanges containing both sequenced and grouped elements. Laboratory egg incubation experiments over a 2-4 year period show that Isogenoides has a great capacity for extended, sometimes-asynchronous diapause and hatching. The eggs of six species were incubated at a single, ca. simulated San Miguel River, Colorado, seasonal temperature regime. Direct hatch within two weeks occurred only for I. zionensis (Leopard Creek, Colorado) with small numbers hatching again after one, two, and four years. Eggs of I. doratus and I. varians hatched after an over summer, 3-5 month diapause, and I. varians again in August the next year. Populations of I. colubrinus, I. elongatus, I. frontalis and San Miguel River, I. zionensis began hatching after a 9-11 month diapause and again during spring-summer temperatures in 2nd through 4th years. I. zionensis in the San Miguel River, Colorado, exhibited a semivoltine life cycle over the two-year study period. Adults emerged in June-July when stream temperature reached ca. 11-17°C. Reared females at Quartz Creek, Pitkin, Colorado, deposited up to three egg batches. Mean fecundity of females was 691 egg/female. Oviposition in the field was observed and described. Nymphal growth …
Numerous troponin T (TnT) isoforms are generated by alternative RNA splicing primarily in its NH2-terminal hypervariable region, but the functions of these isoforms are not completely understood. In this dissertation work, calcium and terbium binding behavior of several forms of TnT were investigated by spectroscopic and radioactive techniques. Chicken breast muscle TnT binds calcium and terbium through its NH2-terminal Tx motif (HEEAH)n with high affinity (10-6 mM) and fast on-rate (106 - 107 M-1 s-1). Chicken leg muscle TnT and a human cardiac TnT NH2-terminal fragment, which both lack the Tx motif on their NH2-terminal regions, do not have affinities for calcium in the physiological range. Computational predictions on TnT N47 suggest that the TnT NH2-terminal region might fold into an elongated structure with at least one high affinity metal ion binding pocket comprised primarily of the Tx motif sequence and several lower affinity binding sites. In addition, calcium binding to TnT N47 might alter its conformation and flexibility. Luminescence resonance energy transfer measurements and other experimental observations are consistent with the computational predictions suggesting the computational simulated atomic model is reasonable. TnT mutations are responsible for 15% of familiar hypertrophic cardiomyopathy (FHC) cases with a phenotype of relatively mild hypertrophy, but a high incidence of sudden death. Detection of those genetic mutations would facilitate the clinical diagnosis and initiation of treatment at an early stage. This dissertation also investigated a novel hybridization proximity assay (HYPA) combining molecular beacon and luminescence resonance energy transfer (LRET) technologies. Experimental results suggest that a shared stem probe design produces a more consistent response upon hybridization, whereas the internally labeled probe was less consistent, but can yield the highest responses. Using the optimally designed molecular probes, the HYPA provides a detection of alterations in nucleic acid structure of as little as a single nucleotide. …
Both the upper and lower temperature tolerances of red fluorescent protein transgenic zebra danios and wildtype zebra danios, Danio rerio, were significantly different via two different methods; however, all differences are small (< 1°C) and probably not ecologically important. The U.S. geographic distributions of both transgenic and wildtype zebra danios will not be restricted by their upper thermal tolerances, but will be limited to the southern and western portions of the U.S. by their lower thermal tolerances. Largemouth bass did not preferentially prey upon transgenic zebra danios compared to wildtype danios or wildtypes relative to a native fish. If transgenic or wildtype zebra danios are released into southern or western U.S. waters, it is possible they could be eliminated by predation.
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