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Bottom-Up Impacts of Grazing Disturbance on Ground-Nesting Bee Assemblages: Do They Dig It?
In my thesis, I examined impacts of nesting and floral resources on ground-nesting bee assemblages at nine ranch sites with differing grazing histories. Because ecological disturbances can strongly affect the availability of foraging and nesting resources over time, I also examined the impacts of grazing history on nesting and floral resources for ground-nesting bees. I sampled bee assemblages, a comprehensive array of vegetation and soil measures, and floral abundance and richness. I used these data to determine (1) the importance of different nesting habitat resources and livestock grazing history for ground-nesting bees and (2) how livestock grazing history influenced the availability of nesting and floral resources. I found that sites with sandier soils had greater abundance and richness of ground-nesting bees, and sites with less compacted soils had greater bee abundance. Contrary to many other studies, the availability of bare ground was not important for ground-nesting bee abundance or richness. Grazing history did not affect any measures of nesting or floral habitat, and had negligible effects on ground-nesting bee abundance, richness, and community composition. My results suggest that the availability of sandier or less compacted soils may be limiting for ground-nesting bee abundance or richness, especially in areas with predominately clay soil types. In this context, the availability of such nesting resources may be more limiting than floral resource availability. More research is necessary to tease apart the effects of different nesting and floral resources for bees, and how different grassland management types affect bees through those resources. My findings highlight the importance of considering nesting habitat resources in bee conservation and restoration efforts.
Hypoxia-Induced Cardiac Arrest Alters Central Nervous System Concentrations of the GLYT2 Glycine Transporter in Zebrafish (Danio rerio)
Hypoxia as a stressor has physiological implications that have been a focal point for many physiological studies in recent years. In some studies, hypoxia had large effects on the organ tissue degeneration, which ultimately effects multiple ecological processes. These organ tissue studies played a part in the development of new fields like neurocardiology, a specialty that studied the relationship between the brain and the heart. This thesis focuses on how hypoxia-induced cardiac arrest alters the amounts of GLYT2, a glycine reuptake transporter, in the central nervous system of zebrafish, Danio rerio. At 7 days post-fertilization (dpf), zebrafish were exposed to acute, severe hypoxia until they lost equilibrium, and minutes later, subsequent cardiac arrest occurred. Zebrafish were then placed into recovery groups to measure the GLYT2 levels at multiple points in zebrafish recovery. Fish were then sacrificed, and their brains dissected. Using immunofluorescence, the outer left optic tectum of the zebrafish was imaged, and mean image pixel fluorescent intensity was taken. There were significant changes (one-way ANOVA) in the levels of GLYT2 compared to that of the control groups during the course of recovery. GLYT2 levels continued to rise through the 24-hour recovery mark but did not show significant difference after 3 hours of recovery. This suggest that GLYT2 levels increased rapidly in the first 3 hours of recovery and continued to increase through 24 hours at a slower rate. Changes in GLYT2 levels may affect motor and sensory information, movement, visualization, and audition in these zebrafish. Further research should be conducted to determine how long it takes for GLYT2 levels to return to baseline, as well as behavioral measurements through each recovery period as it relates to glycine function.
Air Breathing Fish: Development of Air Breathing in Bristlenose Plecos (Ancistrus cirrhosus)
The bristlenose pleco (Ancistrus cirrhosus) is a species of armored catfish in the Loricariidae family that breathes air facultatively when the aquatic environment becomes hypoxic. The bristlenose pleco uses its highly vascularized stomach as an air breathing organ. The two main goals of this developmental study were to determine the size of onset of air breathing and to determine the frequency of air breathing behavior in bristlenose plecos from juveniles to adults. Developing juveniles reach functional maturity within four to six months of hatching and grow to an adult size of eight to ten cm in length. To examine the developmental timing for the onset of air breathing, we tested different sized juveniles beginning at one cm up until 8 cm in length. The developmental timing for the onset of air breathing was measured by exposing each fish to a slowly decreasing aquatic oxygen content from 100% air saturation down to 8% air saturation. Fish were first able to breathe air at just over 2 cm and 1 gram in mass. There was a weak negative correlation between fish length and % air saturation at which air breathing began. When exposed to 15% air saturation, frequency of air breathing was negatively correlated with fish length. Armored catfish are becoming an invasive species in the southern US, outcompeting local fauna potentially because of this adaptation. This research provides important insight into the development of the air breathing adaptation that may allow these fish to outcompete the others.
Secondary Production of Dragonflies: Comparing Ecosystem Function of Ponds within an Urban Landscape in North Central Texas
The change of land use to include more urban areas is considered one of the main threats to biodiversity worldwide. Urban stormwater retention ponds have been built to collect storm runoff intensified by the increase in impervious surfaces. Although subject to environmental pressures like habitat degradation and pollution, these stormwater retention ponds are diversity hotspots by providing habitat for several aquatic and semi-aquatic species, including dragonflies. Previous research in Denton, Texas, has demonstrated that urban stormwater retention ponds support high taxa richness of adult dragonflies, but not for the aquatic nymphs. The current study builds on what we have seen by focusing on the immature aquatic stage as nymphs using secondary production of dominant dragonfly taxa and community structure to compare ecosystem function in three ponds with differing intensities of land use. Comparing communities and secondary production resulted in specific conductivity, dissolved oxygen, complex vegetation, and abundance explaining the differences between dragonfly communities. Secondary production was dependent on abundance which followed the intensity of urban land use surrounding the pond. This study supports that urban land use does have an effect on the functioning of the ponds and shows the importance of studying the communities over a year to get a clearer picture of what is happening in the ponds.
Conservation, Connectivity, and Coexistence: Understanding Corridor Efficacy in Fragmented Landscapes
Conservation corridors, areas of land connecting patches of natural land cover, are frequently cited and implemented as a restorative strategy to counteract fragmentation. Current corridor ecology focuses on experimental corridor systems or designed and built conservation corridors to assess functionality. Such systems and designs are typically short, straight swaths of homogenous land cover with unambiguous transitions between patches. Quantifying the degree to which amorphous landscape configurations, tortuosity, and heterogeneity of land cover and land uses within the corridor has on functional connectedness is a crucial yet overlooked component of corridor efficacy studies. Corridor literature lacks a robust and repeatable methodology for delineating existing landscape elements, recognizing arbitrary edges, and identifying the start and end of ambiguous transitions between the patches and corridor. Using a set of landscapes being studied as part of a global assessment of corridor efficacy, I designed a workflow that standardizes the boundary of corridor-patch interfaces. The proposed method is a quantitative and repeatable approach that minimizes the subjectivity in corridor delineations. This research investigates the degree to which the existence of a corridor modifies the structural and functional connectivity between patches connected by a corridor compared to an intact reference area.
Combined Effects of Polycyclic Aromatic Hydrocarbons and Ultraviolet Light on Benthic and Pelagic Macroinvertebrates
Crude oil commonly enters freshwater aquatic ecosystems as thin sheens forming on the water surface. Oil contains mixtures of toxic compounds called polycyclic aromatic hydrocarbons (PAHs), some of which are known to be photodynamic, increasing toxicity when combined with ultraviolet radiation. Benthic macroinvertebrate communities are commonly utilized as bioindicators, and as such rely on abundant data in literature concerning benthic macroinvertebrates' relative tolerances to a wide range of pollutants. A series of 10 plastic traps, half of which were filtered from UV radiation, were deployed in an urban pond for 27 days to determine colonization preferences of benthic macroinvertebrates to UV exposure. Results of this in situ experiment indicated that the majority of aquatic insects collected from traps inhabited the UV exposed treatment group, particularly the nonbiting midge, Chironomidae. A series of bioassays were then completed to investigate the sensitivities of a Chironomidae species to thin sheens of crude oil in the presence and absence of UV radiation. All bioassays were conducted using 10 day old Chironomus dilutus larvae cultured in the lab. The series of C. dilutus bioassays were all conducted under the same water quality parameters, temperatures, and oil sheen dosing methods, under a 16:8 photoperiod and exposed to 16 h UV per day. Five replicates (n = 20) were loaded into 350 mL glass crystallizing dishes and exposed to four treatments for 96 hours: no UV/with sheen, with UV/no sheen, both UV and oil sheen, and a control. Three assays with 175 mL water volume were completed, one with no sediment, one with silica sand, and one with fine sand. Sediment type had a significant effect on mortality (p < 0.0001), but significant effects of UV or PAHs were not found. Two more C. dilutus assays were completed with identical parameters as the latter two with sediment, …
Greater, Lesser, Guessers: A Look into the Hybridization of Greater and Lesser Prairie-Chickens
My thesis focuses on the conservation consequences of the hybridization of Lesser Prairie-Chickens in Kansas. Specifically, examining how past land management practices altering the species ranges impact the distinctiveness of Lesser Prairie-Chickens. Each chapter is an individual publication that addresses if the Greater and Lesser Prairie-Chicken are distinct when applying the morphological and biological species concepts. Chapter 2 compares the evolutionary history and morphological construct of Lesser Prairie-Chickens and other Galliformes using morphometric analysis. Chapter 3 uses low-resolution microsatellite data to reflect recent changes at the population level. This study aims to observe the Greater and Lesser Prairie-Chicken using the morphological and biological species concepts, two of the many species concepts, to determine the distinctiveness and rate of hybridization for these closely related species.
Data Mining Using Direct Injection Triple Quadrupole Mass Spectrometry, Infrared Spectroscopy, Inductively Coupled Plasma Optical Emission Spectroscopy, and Polymerase Chain Reaction for the Rapid Identification of Nutraceuticals and Contaminants
There has been a rapid surge toward "organic" products devoid of GMOs, MSGs, and other common compounds found in processed foods that continue to indicate an association with an increased risk for disease. These consumers seek nutrients and vitamins that are lacking in their diet and lifestyle in the form of nutraceuticals for disease prevention and treatment as well as overall lifestyle enhancement. However, these products generally lack clinical evidence as well as legal definition. Due to this ambiguity, nutraceuticals are neither considered a food product nor a pharmaceutical product. Furthermore, due to their alleged natural properties allowing for safe, therapeutic effects, nutraceuticals are being eagerly sought after by consumers in the place of pharmaceuticals. Additionally, since nutraceutical substances are "naturally" derived, there is a general lack of regulation regarding the manufacturing and distribution process. This mismanagement leads to lack of quality assurance (QA) and quality control (QC) protocols strictly implemented to define appropriate production and storage parameters. Without these critical measures, consumers are subjected to contamination of their products resulting from improper storage conditions and unmanaged production. These contaminants often include heavy metal impurities, pesticides, bacterial activity, and may also be adulterated with illicit drugs, all leading to detrimental health and environmental effects.
Influence of Hypoxia on Acute Lead Toxicity and Calcium Homeostasis in Early Life Stage Zebrafish (Danio rerio)
The purpose of this study was to investigate the effects of Pb and hypoxia co-exposure on Pb toxicity and Ca homeostasis in early life stage (ELS) zebrafish (Danio rerio). Previous evidence indicates that exposure of ELS zebrafish to hypoxia (~20% air saturation) reduces Ca uptake, likely through down-regulation of the apical epithelial Ca channel (ECaC). Considering that Pb and Ca are known antagonists and compete for uptake pathways, it was hypothesized that co-exposure of Pb with hypoxia would decrease Pb toxicity by reducing Pb uptake (likely mediated through a reduced number of ECaCs). However, it was shown that at 96 hpf, whole body accumulation of both Pb and Ca was lower at 40% air saturation compared to 100% and 20% air saturation. This result closely aligned with the 96h LC50 results which showed the highest mortality of zebrafish at 40% compared to the other air saturation levels. This suggests that toxicity is likely the result of exacerbated hypocalcemia at 40% air saturation due to both Pb competition for Ca binding to Ca uptake channels/transporters, such as ECaC, and potentially reduced expression of such channels/transporters in response to this level of hypoxia. Overall, it appears that ELS zebrafish respond differentially to the 40% and 20% hypoxia levels when co-exposed with Pb. Further investigation is needed to illustrate the physiological and molecular mechanisms underlying this response.
Reduced Visceral Fat and Biological Indices of Inflammation Following Combined Prebiotic/Probiotic Supplementation in Free Living Adults
Probiotics/prebiotic supplementation represents a viable option for addressing systemic inflammation and chronic disease risk resulting from excessive body weight. The purpose of this feasibility study was to determine if 90-d of supplementation with prebiotic and probiotic could alter mRNA responsible for inflammation and subsequently metabolic health in weight stable overweight adults. Participants were advised to not change their diet or exercise habits during the study. All protocols were approved by the University IRB and participants gave written informed consent. Participants were randomly assigned to either placebo (N=7; rice flour) or combined (N=8) prebiotic (PreticX® Xylooligosaccharide; 0.8 g/d; ADIP) and probiotic (MegaDuo® Bacillus subtilis HU58 and Bacillus coagulans SC-208; 3 Billion CFU/d) and measurements were made at baseline, 30, 60, and 90-d. Whole body DXA scans (GE iDXA®) and blood 574-plex mRNA analysis (Nanostring®) were used to generate primary outcomes. Compared to placebo, supplementation was associated with a 36% reduction in visceral adipose tissue (p = 0.001). Supplement resulted in significant, differential expression of 15 mRNA associated with adipose tissue inflammation, systemic inflammation, and/or chronic disease risk. The key findings support that 90-d prebiotic/probiotic supplementation may be associated with an improved metabolic health, reduced adipose tissue inflammation, reduced systemic inflammation, and reduced chronic disease risk. Collectively these findings demonstrate the potential of a prebiotic/probiotic supplement to impact metabolic health risk independent of weight loss in free-living individuals.
Flow-Recruitment Relationships of Smallmouth Buffalo (Ictiobus bubalus) in Three Texas River Basins
This project focused on the relationship between instream flows and smallmouth buffalo (Ictiobus bubalus) recruitment in the Gulf Coastal Plain of Texas. The flow regime is the dominant factor in lotic systems and, consequently, the relationship between instream flows, including impacts to natural flow regimes, and life-history is a subject of growing interest. Smallmouth buffalo is a good model to investigate the relationship between river flows and variable interannual recruitment success of periodic life-history strategist fish species. Smallmouth buffalo were collected from the Brazos, Colorado, and Guadalupe Rivers of Texas, U.S.A., and otoliths were extracted from individuals in the field and sectioned and photographed in the lab. Photographs of sectioned otoliths were used to estimate age and thus the year in which the individual was spawned by counting back from the time of capture. Population age structure (i.e. a ‘state' or condition at a point in time) was used to infer effects of flow variation on a rates-based process (i.e. recruitment). After controlling for mortality using recruitment index values, interannual variation in recruitment was modeled using multiple components of the flow regime quantified as indicators of hydrologic alteration (IHA) variables based on daily discharge data from USGS gaging stations in each river system. Model selection followed a two-tier approach, first fitting models using only flow attributes associated with the spawning season then adding additional informative parameters from the pre-spawn and post-spawn periods. The primary finding from model selection was that duration of high flow pulses during the spawning season is a critical component of the flow regime associated with successful Smallmouth Buffalo recruitment. These findings have implications for river management and conservation of ecological integrity, in particular populations of periodic life-history strategist species.
Sociality in Harris's Hawks Revisited: Patterns of Reproductive Output and Delayed Dispersal
In the lower Rio Grande Valley of south Texas, more than half the nesting groups of Harris's hawks (Parabuteo unicinctus) include at least one auxiliary group member in addition to a breeding pair. To provide further insight into cooperatively breeding raptors, I evaluated sociality in Harris's hawks through the dual benefits framework. I explored the formation, structure, and stability of cooperative group formation across a spatially variable study area, which includes high levels of urbanization and development as well as remote, undisturbed native habitats with low anthropogenic impact. I used color banding, regular censuses of active territories, and a microsatellite relatedness analysis to examine patterns of sociality, including delayed dispersal, the effect of auxiliary group members on reproductive output, parentage of broods, and the relatedness of auxiliaries compared to the nestlings in their territories. I confirmed cooperative polygamy with genetic techniques for the first time in Harris's hawks and found 58% of juvenile hawks delayed dispersal for at least 6 mo. Using the dual benefits framework, I found social associations that formed through delayed dispersal followed predictions for resource-defense benefits, but sociality among mature non-related hawks more closely followed predictions associated with collective action benefits, specifically reproductive output was significantly reduced in undeveloped habitats, presumably due to a less predictable prey-base.
A Genetic Assessment of the Mating System of a Suburban Red-Shouldered Hawk Population in Southwest Ohio
Considering the high reproductive investment of the social male and the cost to the female of losing this benefit by soliciting copulations outside the social pair bond, it is expected that most raptor populations would exhibit low to no occurrence of extra-pair paternity (EPP). This holds true for the majority of raptor species studied to date with only one exception of an urban Cooper's hawk (Accipiter cooperii) study which reported an unexpectedly high extra-pair young frequency of 19.29%. In our study we examined the frequency of EPP within a red-shouldered hawk (Buteo lineatus) population residing in the suburban/urban matrix of southwest Ohio. During the breeding seasons of 2018 and 2019, 181 breeding age and nestling individuals were color-banded and sampled for genetic analysis using nine microsatellite loci. After genotyping a total of 40 broods (with at least two nestlings per brood) and both presumptive parents of each brood, no clear evidence of EPP was detected. However, at one nest site, the entire brood of four chicks was not sired by the adult male observed during the courtship period, nor another adult male observed tending the chicks later in the season. We suspect that this particular nest represented two instances of rapid mate replacement rather than extra-pair fertilization by a third unsampled male, because none of the chicks were sired by either of the two adult males observed at the nest. We also reviewed potential factors contributing to our finding of overall genetic monogamy in our study population in comparison to other raptor taxa EPP studies. Our results suggested that factors other than habitat composition alone play an important role in determining the type of breeding strategy exhibited by different raptor populations.
A Test of the Female Mimicry Hypothesis in Painted Buntings (Passerina ciris)
While female mimicry and lower status signaling hypotheses of delayed plumage maturation have received much discussion in the literature, the experimental tests of these hypotheses have been infrequent. Those experimental tests often use a simulated intruder method with artificial model intruders rather than using live conspecific birds as intruders. Subadult male painted buntings (Passerina ciris) possess delayed plumage maturation where they appear visually identical to adult females during their first potential breeding season, while adult males are strikingly different in plumage coloration. Here I test the behavioral responses in a territorial population of painted buntings that exhibits extreme delayed plumage maturation using a simulated territorial intrusion experiment to measure territorial male behavioral response when presented with live caged intruders of both subadult and adult males. Territorial adult males were significantly more likely to initiate an attack and continue to attack caged adult male intruders than compared to caged subadult male intruders. This result supports both the female mimicry and status signaling hypotheses, and does not support the cryptic hypothesis. Additionally, in anecdotal observations, territorial males occasionally performed mating display behaviors to caged subadult male intruders. These results further suggest that territorial male painted buntings may identify subadult males as potential mates, supporting the female mimicry hypothesis for subadult males in this species. To what degree subadult males may benefit from DPM deserves further study.
Peregrine Falcon (Falco peregrinus) Subspecies Phylogenomics Using Whole Genome Re-Sequencing
Peregrine falcon subspecies taxonomy is widely debated due to uncertainty in their evolutionary history and unresolved phylogenetic reconstruction using both morphological and molecular data. Previous genetic work has shown limited support for subspecies taxonomy largely as a result of molecular markers used, potential contemporary gene flow, incomplete lineage sorting, and ancestral polymorphisms. With the advent of next-generation sequencing, the cost of generating large amounts of sequence data has dropped significantly, making whole genome re-sequencing (WGR) studies of non-model organisms more tangible. In this study, WGR methods have been utilized to investigate the phylogenetic relationships among all 20 currently recognized peregrine falcon subspecies. By generating whole-genome data for all 20 subspecies, subspecies specific diagnostic SNPs have been identified to aid in subspecies delimitation. Results of this study broadly support current subspecies, however, reveal that further study is needed to investigate regional relationships among subspecies in Asia, Australia, and western North America. With these results, conservation efforts can be further supported by allowing for accurate delimitation of local subspecies and subspecies boundaries.
Metabolic Responses to Crude Oil during Very Early Development in the Zebrafish (Danio rerio)
The present study sought to determine some morphological and physiological critical windows during very early development in zebrafish exposed to crude oil. I hypothesized that exposed zebrafish would present a decrease in survival rate and body mass, and an increase in routine oxygen consumption (ṀO2), and critical oxygen tension (PCrit). To test these hypotheses, zebrafish were acutely exposed (24 h) during different days of development (1 to 6 days post-fertilization, dpf) to different concentrations of high-energy water-accommodated fractions (HEWAFs). The endpoints of survival, body mass, routine oxygen consumption, and critical oxygen partial pressure were measured at 7 dpf. Survival rate decreased based on the exposure concentration but not as a function of the day of crude oil exposure. No significant effects were found in PCrit. Body mass was reduced by the different concentrations of HEWAF, with the size of the effect varying with exposure day, with the effect strongest on when exposure occurred at 2 and 3 dpf. Oxygen consumption (ṀO2) differed significantly depending upon the day of exposure in fish exposed to crude oil. Specifically, HEWAF exposure significantly increased ṀO2 in larvae exposed at 3 dpf (9.081 µmol O2/g/h, ±0.559) versus 2 dpf (6.068 µmol O2/g/h, ±0.652) and 6 dpf (6.485 µmol O2/g/h, ±0.609). Overall, the main effects on body mass and ṀO2 occurred at crude oil exposures during 3 dpf. The presence of a critical window in fish is proposed at this developmental time, which coincides with the hatching period.
The Effects of Probiotics on Growth, and Metabolism in Juvenile Oreochromis mossambicus (Mozambique Tilapia)
Improving growth, lowering mortality rates, and having a faster turnaround to harvest is essential for the future of commercial aquaculture. The primary goal of this study was to determine if introducing a single strain probiotic Lactobacillus rhamnosus IMC 501 into the feed regimen of a commercially important aquaculture freshwater fish, Mozambique tilapia (Oreochromis mossambicus), would decrease mortality; change metabolic rates; and increase tissue wet mass (MW), standard length, growth rate and feed conversion rate (FCRs). IMC501 was added to the fishmeal in four increasing concentrations and compared to a control without probiotics. Results from two-way ANOVAs showed that both treatment levels and elapsed time had a significant effect on both mean standard length and wet mass; in the latter case, time points and treatments interacted with one another, showing that tilapia grew best with a moderate level of probiotics present. The growth benefits of probiotics continued for months after the initial treatments. Oxygen consumption (metabolic rate) was measured using closed respirometry and resulted in recording the first values for juvenile tilapia treated with probiotics. For oxygen consumption, there were significant treatment and time effects with significant interactions, indicating that metabolism increased with probiotics once the dosage exceeded three times the industry level. These results are consistent with the observed increases in mass, length and growth rates. These results demonstrate the importance of conducting dose-response experiments in order to determine the most effective concentration of probiotics in juvenile freshwater fish. Importantly, probiotics at the right concentration increase metabolic rates and can positively influence tilapia growth, which is of interest for the optimization of Mozambique tilapia production in aquaculture.
Storm Water Retention Ponds: An Important Source of Aquatic Macroinvertebrate Diversity in a Semi-Arid Urban Landscape of Denton, Texas
The City of Denton, located in a semi-arid region of Texas, has over 200 manmade ponds within its city limits. Many of these ponds, located in densely populated areas, are engineered to control storm water runoff. There is a general lack of recognition of the value these waters contribute to regional biodiversity and as greenspaces. This study, conducted in Denton, is monitoring habitat variables and macroinvertebrate diversity in a series of ponds selected to represent a gradient of urban influences. The objective of this study is to identify the variables associated with the highest diversity. Using drone imagery and a meter square box sampler, the quantitative approach allowed for delineation of three habitat types and area. The macroinvertebrates where identified to the genus level which allowed for higher resolution and resulted in stronger comparisons of the communities and conditions of the ponds. Taxa richness was positively correlated to pond size and trees along shoreline and negatively associated with average depth. Overall, submerged vegetation supported highest diversity and abundance, especially genera of Chironomidae (Diptera). Conductivity was associated with urban influences and the most urban influenced pond had the lowest taxa richness, but also reduced habitat area. Results of this study conclude that these stormwater ponds benefit to the ecology of the city and provide beautiful, green spaces. If managed correctly, these systems can be incorporated into sustainable development in the future of the City of Denton.
Optical Brain Imaging of Motor Cortex to Decode Movement Direction using Cross-Correlation Analysis
The goal of this study is to determine the intentional movement direction based on the neural signals recorded from the motor cortex using optical brain imaging techniques. Towards this goal, we developed a cross-correlation analysis technique to determine the movement direction from the hemodynamic signals recorded from the motor cortex. Healthy human subjects were asked to perform a two-dimensional hand movement in two orthogonal directions while the hemodynamic signals were recorded from the motor cortex simultaneously with the movements. The movement directions were correlated with the hemodynamic signals to establish the cross-correlation patterns of firings among these neurons. Based on the specific cross-correlation patterns with respect to the different movement directions, we can distinguish the different intentional movement directions between front-back and right-left movements. This is based on the hypothesis that different movement directions can be determined by different cooperative firings among various groups of neurons. By identifying the different correlation patterns of brain activities with each group of neurons for each movement, we can decode the specific movement direction based on the hemodynamic signals. By developing such a computational method to decode movement direction, it can be used to control the direction of a wheelchair for paralyzed patients based on the changes in hemodynamic signals recorded using non-invasive optical imaging techniques.
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.
Isolation and Bioinformatic Characterization of Four Novel Bacteriophages from Streptomyces toxytricini
Six initial phage isolates with high titer lysates were obtained using Streptomyces toxytricini B-5426 as the host bacterium. These isolates were named Goby, Toma, Yosif, Yara, Deema, and Hsoos. However, upon completion of the sequencing, it was found that the Yara and Hsoos isolates were identical, as were Goby and Deema. As a result, final analysis was completed on only the four unique isolates. All of the phages mentioned above were isolated from soil samples from different locations. Also, they had different sizes of plaques, ranging from 0.3 – 0.9mm. Yosif had the largest plaque size. Yara's head diameter was 79nm with tail diameter of 94nm.
Autonomic Nerve Activity and Cardiovascular Function in the Chicken Embryo (Gallus gallus)
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 Role of Thyroid Hormone on the Development of Endothermy in White Leghorn Chickens (Gallus gallus)
As chickens hatch, there is a rapid change in their physiology and metabolism associated with attaining endothermy. It is thought that thyroid hormones (TH) play a major role in regulating developmental changes at hatching. In birds, TH regulates skeletal muscle growth, which has a direct impact on the chick's ability to thermoregulate via shivering thermogenesis. To better understand the role of TH in the timing of hatching, development of thermogenic capacity, and metabolic rate, we manipulated plasma TH levels in chicken embryos beginning at 85% development (day 17 of a 21 day incubation) with either thyroperoxidase inhibitor methimazole (MMI) or supplemental triiodothyronine (T3). After TH manipulation, we characterized O2 consumption and body temperature in the thermal neutral zone and during gradual cooling. Externally pipped embryos and 1 day post hatch (dph) chicks were cooled from 35 to 15°C. Manipulation of TH altered the timing of hatching, accelerating hatching under hyperthyroid conditions and decelerating hatching with hypothyroid conditions. Cohen's d revealed a large effect size on body temperature (Tb) of EP embryos of hypothyroid animals when compared to euthyroid animals in environmental temperatures of 32°C to 15°C, which was not seen in 1dph animals. Hyperthyroid EP animals were able to maintain metabolic rate over a wider range of ambient temperatures compared to control and hypothyroid animals, but these differences disappeared in 1dph animals. Here, we find that elevating TH levels prior to hatching accelerated hatching and the animal's thermogeneic ability to respond to cooling, but these differences disappear with age.
The Role of Transmembrane Protein 59 in Thrombocyte Function and the Effect of MS-222 on Hemostasis in Zebrafish
Transmembrane protein 59 (tmem59) is a gene that encodes a protein involved in autophagy and apoptosis in human. A previous study in zebrafish showed that tmem59 mRNA was several folds higher in thrombocytes than those found in red blood cells (RBCs). Therefore, we hypothesized that tmem59 has a role in thrombocytes function. We injected a hybrid of control vivo-morpholino (cVMO) and tmem59 specific antisense standard oligonucleotide (tmem59SO) into adult zebrafish to knockdown tmem59.This piggyback knockdown approach resulted in fish that had more bleeding in gill bleeding assay than the control fish. The thrombocytes fromtmem59 knockdown zebrafish aggregated faster with ADP and collagen agonists. Also, the number of blood cells was reduced after the knockdown of tmem59. We also found the effects of MS-222 anesthesia on hemostasis and found that the bleeding was reduced yielding less blood and the blood cell counts increased probably due to vasoconstriction of the blood vessels. In summary, we found tmem59 is a negative regulator of hemostasis and inferred that anesthesia should be avoided in hemostasis studies.
Superbursts: Investigation of Abnormal Paroxysmal Bursting Activity in Nerve Cell Networks In Vitro
Superbursts (SBs) are large, seemingly spontaneous activity fluctuations often encountered in high density neural networks in vitro. Little effort has been put forth to define and analyze SBs which are paroxysmal bursting discharges. Through qualitative and quantitative means, I have described specific occurrences of superbursting activity. A complex of paroxysmal bursting has been termed a "superburst episode," and each individual SB is a "superburst event" which is comprises a fine burst structure. Quantitative calculations (employing overall spike summations and coefficient of variation (CV) calculations), reveal three distinct phases. Phase 1 is a "build up" phase of increasingly strong, coordinated bursting with an average of a 17.6% ± 13.7 increase in activity from reference. Phase 2, the "paroxysmal" phase, is comprised of massive coordinated bursting with high frequency spike content. Individual spike activity increases by 52.9% ± 14.6. Phase 3 is a "recovery phase" of lower coordination and an average of a 50.1% ± 35.6 decrease in spike production from reference. SBs can be induced and terminated by physical manipulation of the medium. Using a peristaltic pump with a flow rate of 0.4ml/min, superbursting activity ceases approximately 28.3 min after the introduction of flow. Alternatively, upon cessation of medium flow superbursting activity reemerges after approximately 8.5 min. Additionally, this study explored other methods capable of inducing superbursting activity using osmotic shocks. The induction and termination of SBs demonstrates that the cell culture environment plays a major role in generating this phenomenon. The observations that high density multi-layer neuronal networks in culture are more likely to enter paroxysmal bursting also supports the hypothesis that enrichment and depletion layers of metabolites and ionic species are involved in such unusual activity. The dynamic similarity of the SB phenomenon with epileptiform discharges make further quantification on the spike pattern level pertinent and important.
In Vitro Exploration of Functional Acrolein Toxicity with Cortical Neuronal Networks
Acrolein is produced endogenously after traumatic brain injury (TBI) and is considered a primary mechanism for secondary damage occurring after TBI. We are using frontal cortex networks derived from mouse embryos and grown on microelectrode arrays in vitro to monitor the spontaneous activity of networks and the changes that occur after acrolein application. Networks exposed to acrolein exhibit a biphasic response profile. An initial increase in network activity, followed by a decrease to 100% activity loss in applications ≥ 50 µM. In applications below 50 µM, acrolein was not toxic but generated activity instability with coordinated but irregular population busts lasting for up to 6 days. The increase in activity preceding toxicity may be linked to a decrease in free spermine, a free radical scavenger that modulates Na+, K+, Ca+ channels as well as NMDA, Kainate, and AMPA receptors. Action potential wave shape analysis after 20 and 30 µM acrolein application revealed a concentration-dependent 15-33% increase in peak to peak amplitude within minutes after exposure. For the same concentrations of acrolein (50 µM), the time required to reach 100% activity loss (IT100) was longer in serum-free medium than in medium with 5% serum, in which IT100 values were reduced by a factor of 4. The greater toxicity in the presence of serum may be explained by acrolein adducts on serum proteins. These reaction products have been shown by other labs to be toxic in cell culture. This in vitro system could be used to expand biochemical analyses such as acrolein-induced spermine depletion and may provide an effective platform for investigating cell culture correlates of secondary TBI damage.
Population Dynamics and Community Structure of Mosquitoes (Diptera: Culicidae) Recorded in Denton, Texas from 2005 to 2015
A population survey was conducted on the mosquito species recorded in Denton, Texas for the years of 2005 to 2015. Data used in this project were obtained from an ongoing, long-term surveillance program led by the City of Denton and conducted through the University of North Texas. Research focused on the population dynamics and community structure of mosquitoes collected within urban areas of Denton, Texas in relation to certain environmental variables. A total of 80,837 female mosquitoes were captured and represented 38 species found under the following genera: Aedes, Anopheles, Coquillettidia, Culex, Culiseta, Mansonia, Orthopodomyia, Psorophora, Toxorhynchites, and Uranotaenia. Culex quinquefasciatus was the most abundant species followed by Aedes vexans. Seasonal patterns of the most abundant species revealed high variability throughout the study. Container breeders were most abundant in August and those that breed in floodwaters were most abundant in the months of May and September. Samples were tested for arbovirus presence through the Texas Department of State Health Services in Austin, Texas and multiple pools tested positive for West Nile virus throughout the study. Stepwise multiple regression and Spearman's rank correlation analyses were performed to examine the relationship between the mosquito community and environmental variables. Data revealed that temperature, precipitation, and dew point were the most important variables influencing the mosquito population in the City of Denton.
Simultaneous Electrophysiological and Morphological Assessment of Impact Damage to Nerve Cell Networks
A ballistic pendulum impulse generator was used to impact networks in primary culture growing on microelectrode arrays. This approach has the advantage of imparting pure tangential acceleration insults (50 to 300 g) with simultaneous morphological and electrophysiological multichannel monitoring for days before and after the impact. Action potential (AP) production, network activity patterns, and cell electrode coupling of individual units using AP waveshape templates were quantified. Network adhesion was maintained after tangential impacts up to 300g with minimal loss of pre-selected active units. Time lapse phase contrast microscopy revealed stable nuclei pre-impact, but post impact nuclear rotation in 95% of observations (n= 30). All recording experiments (n=31) showed a repeatable two-phase spike production response profile: recovery to near reference in 1-2 hrs, followed by a slow activity decay to a stable, level plateau approximately 30-40% below reference. Phase 1 consisted of a complex two-step recovery: rapid activity increase to an average 23.6% (range: 11-34%) below reference, forming a level plateau lasting from 5 to 20 min, followed by a climb to within 20% of reference where a second plateau was established for 1 to 2 hrs. Cross correlation profiles showed changes in firing hierarchy after impact, and in spontaneous network oscillatory activity. Native oscillations were found in the Delta band (2 to 3 Hz), and decreased by approximately 20% after impact. Under network disinhibition with bicuculline, oscillations were slower (0.8-1Hz) and decreased 40% after impact. These data link network performance deficits with microscopically observable subcellular changes.
Phenotypic Analysis of Medicago truncatula NPF1.7 Over-Expressing Plants Grown under Different Nitrate Conditions
Plants have many nitrate transporters; in the model legume Medicago truncatula, MtNPF1.7 is among them. MtNPF1.7 is important for M. truncatula growth and it has been established that MtNPF1.7 is a high affinity nitrate transporter. M. truncatula plants with mutations in MtNPF1.7 gene show defects during plants growth, with striking abnormalities in nodule development and root architecture. Nitrogen fixation is an energy expensive process; when legumes have sufficient bioavailable nitrogen like nitrate available, it suppresses nodulation and nitrogen fixation. Previous preliminary results in our lab showed that plants constitutively expressing MtNPF1.7 have a growth phenotype in the absence of nitrate, but no data was available on how M. truncatula plants constitutively expressing MtNPF1.7 are affected by the presence of nitrate. For my research, I confirmed the preliminary results on the growth of M. truncatula plants overexpressing NPF1.7 and examined these plants' phenotypes when nitrate was not provided in the growth media and when it was provided at two different concentrations. Compared with wild type A17, plants constitutively expressing MtNPF1.7 gene grow larger, have more lateral roots and more nodules when grown in the absence of nitrate and when 0.2 mM KNO3 was provided. At 1 mM KNO3, there are fewer differences between wild type A17 and plants constitutively expressing the MtNPF1.7 gene. Compared with wild type A17, plants constitutively expressing the MtNPF1.7 gene flower earlier, which indicates MtNPF1.7 gene may have a function in plant flowering.
Evaluating the Role of UV Exposure and Recovery Regimes in PAH Photo-Induced Toxicity to Daphina Magna
Polyaromatic hydrocarbons (PAHs) are contaminants synthesized through incomplete combustion of carbon based substances. PAHs are known to be photodynamic and toxicity increases exponentially when in contact with ultraviolet radiation (UV). The effect of UV absent recovery periods and potential for latent toxicity during photo-induced toxicity are previously unknown and are not included within the toxicity model. Results of equal interval tests further support the current reciprocity model as a good indicator of PAH photo-induced toxicity. Interval test results also indicate a possible presence of time-dependent toxicity and recovery thresholds and should be included into toxicity risk assessments. Moreover, results of latent effects assays show that latent mortality is a significant response to PAH photo-induced toxicity and should be included into toxicity risk assessments. The present research demonstrates that UV exposure time rate is a significant driving force of PAH photo-induced toxicity.
Neurological Responses to a Glucose Diet in Caenorhabditis elegans
TRPV channels play a role in both mammalian insulin signaling, with TRPV1 expression in pancreatic beta-cells, and in C. elegans insulin-like signaling through expression of OSM-9, OCR-1, and OCR-2 in stress response pathways. In response to a glucose-supplemented diet, C. elegans are know to have sensitivity to anoxic stress, exhibit chemotaxis attraction, and display reduced egg-laying rate. Transcriptome analysis reveals that glucose stimulates nervous system activity with increased transcript levels of genes regulating neurotransmitters. Ciliated sensory neurons are needed for a reduced egg-laying phenotype on a glucose-supplemented diet. Egg-laying rate is not affected when worms graze on glucose-supplemented Delta-PTS OP50 E. coli, which is defective in glucose uptake. This suggests a possible sensory neuron obstruction by exopolysaccharides produced by standard OP50 E. coli on glucose, eliciting a starvation response from the worm and causing reduced egg-laying rate. Glucose chemotaxis is affected in specific TRPV subunit allele mutants: ocr-2(vs29) and osm-9(yz6), serotonin receptor mutants: ser-1(ok345) and mod-1(ok103), and G-alpha protein mutant: gpa-10(pk362). TRPV deletion mutants had no effect on glucose chemotaxis, alluding to the modality role pf TRPV alleles in specific sensory neurons. The role of serotonin in a reduced egg-laying rate with glucose remains unclear.
Generating Molecular Biology Tools to Investigate the Ca2+ Binding Ability of Arabidopsis TON2
The position of the cell division plane in plants is determined by the position of the preprophase band. The pre prophase band (PPB) is a ring of microtubules centered around the nucleus on the inner side of plasma membrane that establishes the cortical division site. The PPB forms at the end of G2 and breaks down at the end of prophase leaving behind protein markers of its position that are collectively called the cortical division site. During cytokinesis the phragmoplast expands towards the cortical division site and mediates the fusion of the new cell plate with the mother cell at that position. Several proteins necessary for PPB formation in plants have been identified, including maize DCD1 and ADD1 and Arabidopsis TON2, which are all type 2A protein phosphatase (PP2A)B" regulatory subunits. DCD1, ADD1, and TON2 localize to the PPB and the cortical division site through metaphase. The PP2A subunits each have two EF-hand domains, which are predicted to bind calcium ions. Since calcium ions are important for some aspects of cell division, we designed a series of constructs to test if TON2 binds calcium. TON2 protein was cloned into expression vectors, pET42a, and expression of TON2 protein was confirmed via Western blotting and immunodetection using a GST antibody. Site directed mutagenesis was used to mutate the TON2 EF-hand domains and mutated cDNAs were also cloned into expression vectors. These were then expressed in bacterial systems. Finally, the GST tagged proteins were purified. In the future, wild-type and mutated proteins TON2 proteins will used in calcium binding assays to determine if TON2 binds calcium.
Delivery of CRISPR/Cas9 RNAs into Blood Cells of Zebrafish: Potential for Genome Editing in Somatic Cells
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.
Investigating Human Gut Microbiome in Obesity with Machine Learning Methods
Obesity is a common disease among all ages that has threatened human health and has become a global concern. Gut microbiota can affect human metabolism and thus may modulate obesity. Certain mixes of gut microbiota can protect the host to be healthy or predispose the host to obesity. Modern next-generation sequencing technique allows accessing huge amount of genetic information underlying microbiota and thus provides new insights into the functionality of these micro-organisms and their interactions with the host. Multiple previous studies have demonstrated that the microbiome might contribute to obesity by increasing dietary energy harvest, promoting fat deposition and triggering systemic inflammation. However, these researches are either based on lab cultivation studies or basic statistical analysis. In order to further explore how gut microbiota affect obesity, this thesis utilize a series of machine learning methods to analyze large amount of metagenomics data from human gut microbiome. The publicly available HMP (Human Microbiome Project) metagenomic sequencing data, contain microbiome data for healthy adults, including overweight and obese individuals, were used for this study. HMP gut data were organized based on two different feature definitions: taxonomic information and metabolic reconstruction information. Several widely used classification algorithms: namely Naive Bayes, Random Forest, SVM and elastic net logistic regression were applied to predict healthy or obese status of the subjects based on the cross-validation accuracy. Furthermore, the corresponding feature selection algorithms were used to identify signature features in each dataset that lead to the differences between healthy and obese samples. The results showed that these algorithms perform poorly on taxonomic data than metabolic pathway data though lots of selected taxa are still supported by literature. Among all the combinations between different algorithms and data, elastic net logistic regression has the best cross-validation performance and thus becomes the best model. In this model, several important …
Effect of pH on the Persistence and Toxicity of Cyfluthrin to Chironomus Tentans
The effect of pH upon the aquatic toxicity of cyfluthrin was determined in 48 h static acute toxicity tests using 2nd instar Chironomus tentans larvae. Tests were conducted in both dechlorinated tap water and lake water of pH 8.0, 7.2, and 6.6. After 48 h, immobilized and dead larvae were removed and replaced with 2nd instar larvae to assess the persistence of toxicity. Midges were cultured in water adjusted to the pH values used in testing. Toxicity if cyfluthrin varied inversely with pH. An increase in the pH of tap water by 2 units resulted in a 2-decrease in toxicity. Toxicity of cyfluthrin also varied between tap and lake water of the same pH. EC50 values in lake water were approximately 2-3 times lower than those in tap water. Toxicity in the lake water and tap water at every pH tested was also significantly different when regression line elevations were compared. Natural waters amended with cyfluthrin were consistently more toxi to the chironomids than tap water of the same pH. Persistence of cyfluthrin at low pH also influenced chironomid behavior. Recovery of normal behavior generally began within 24 h at pH 8.0 At pH 6.0, recovery did not begin until one week after dosing. The persistence of cyfluthrin also varied with pH. Averaged across all concentrations, 30% of the initial dose remained in tap water (pH 8.0) after 48 h, compared with 45% (pH 7.2), and 75 % (pH 6.6).
Advanced Techniques in Microbial and Molecular Biology: Laboratory Procedures for a Graduate Level Course
Advanced laboratory techniques for Microbial and Molecular Biology at the graduate level are presented in this thesis. The procedures for the laboratory experiments are set forth in detail. This laboratory is conducted as two parts, each by a different professor. Part 1 covers the experiments conducted by Dr G.A.O. Donovan. These experiments include an introduction, staining procedures, biochemical reactions, mutagenesis experiment, essays,. preparation and analysis of plasmid DNA and various other topics. Part 2 covers the experiments conducted by Dr. Daniel Kunz and includes various topics like media preparation, phenotyping strains, conjugative transfer of plasmids, SDS-PAGE, induction and measurement of enzyme and transposon mutagenesis
A Water Quality Study of Lake Texoma
A series of four stations for sampling the waters of the Red Rivers and Lake Texoma was established. Water samples from top, middle, and bottom levels of these four stations were taken on monthly intervals during periods of homothermousity and on two-week intervals during periods of thermal stratification. These samples were analysed for the cations sodium, potassium, calcium, and magnesium, and for the anions normal carbonate, bicarbonate, sulfate, chloride, nitrate, water hardness (expressed as calcium carbonate), and phosphates. Determinations for dissolved oxygen, carbon dioxide, pH, temperature, and specific conductance were performed. Using these data, vertical profiles of the lake at these four stations were established. From the study of these data it was concluded that the waters of the Red River improve in quality coincident with impoundment, and that these improvements are sufficient to warrant the use of these waters.
Effects of Nutrient Media on Growth and Morphology of Azotobacter Vinelandii
The work described in this thesis was undertaken to study the reasons why Azotobacter vinelandii ATCC 12837 after incubation in Burk's nitrogen-free liquid will not form as many colonies when plated on Difco Tryptic Soy Agar as when planted on Burk's nitrogen-free agar. The difference in growth of A. vinelandii on the two agars was established by performing viable cell-plate counts. The difference in growth was most apparent at 24-hours incubation of the Burk's liquid-media cultures. Phase contrast microscopic observations of Tryptic Soy media cultures of A. vinelandii disclosed the regular formation of fungoid cells at early stages of growth of the bacteria, 18 to 24 hours.
A Comparison of Thermogenesis by Selected Substrates on Hypothermic Rat Liver
The thermogenic effects in hypothermia of four substrates--alanine, glycine, ethano, and pyruvate - were studied in seventeen experiments. Albino rats were decapitated, and their livers were removed. The livers were homogenized with phosphate buffer at -5° C. After equilibration in a refrigerated Warburg apparatus at 20° C, the substrates were added and tissue respiration was recorded over three hours. Heat production was calculated from O2 uptake and CO2 production. Results showed that alanine, glycine, and pyrvate yielded 93.19, 89.86, and 89.89 x 10^6 kg-cal compared to a control value of 86.11 x 10^-6 kg-cal. Ethanol provided 110.31 x 10^-6 kg-cal, a value significantly greater than for the other substrates. The substrates studied, especially ethanol, did, therefore increase heat production in an artificially hypothermic environment in homogenized rat livers.
Respiratory Responses in the Freshwater Snail (Pomacea Bridgesii) are Differently Affected by Temperature, Body Mass,and Oxygen Availability
Pomacea bridgesii is a snail species native to tropical and sub-tropical regions, where it usually faces variability in water, temperature and oxygen level. This study of the effect of temperature on mass-specific oxygen consumption (ṀO2) and its relation to body weight shows that the ṀO2 of juvenile snails in normoxia (18-21 kPa) acclimated at temperature of 25°C ranged from 5 to 58 µMol O2/g/h, with a mean of 41.4 ± 18.3 µMol O2/g/h (n=7). Adult snails in normoxia at 25°C show less variation, ranging from 13 to 23 µMol O2/g/h , with a mean of 24.4± 6.1 µMol O2/g/h (n=12). The Q10 value for juvenile snails was higher in the interval 25-30°C (Q 10=5.74) than in the interval 20-25°C (Q10= 0.286). In adult snails, Q10 was higher in the interval 20-25°C (Q10=3.19). ṀO2 of P. bridgesii in relation to body weight showed a negative linear correlation between metabolic rate and body weight with b values between 0.23 and 0.76. Also, both juvenile and adult snails exhibited weak O2 regulation. In general, the different respiratory characteristics between juvenile and adult snails might be related to the differences of individual life history, which caused them to perform differently in face of temperatures change. Additionally, Pomacean snails species originated in tropical habitats where there is a lack of thermal fluctuation. For this reason, Pomacean snails may be less likely to have evolved effective thermal acclimation capabilities.
The Role of Cysteinyl Leukotriene Receptor 2 in Thrombocyte Aggregation
Cysteinyl leukotriene receptor 2, a G-protein coupled receptor known to be expressed and functional on human platelets. However, it seems that upon ligand activation the cysteinyl leukotriene receptor 2 activates a variety of signaling pathways in multiple cell types among different species. Previously, a former laboratory member Vrinda Kulkarni found cysteinyl leukotriene receptor 2 to be expressed on the surface of adult zebrafish thrombocytes. In this work I studied the characteristics of aggregation in adult zebrafish thrombocytes with the knockdown of cysteinyl leukotriene receptor 2. I used a newly developed knockdown method to study the function of cysteinyl leukotriene receptor 2. Knockdown of the cysteinyl leukotriene was confirmed using RT-PCR results showed p=.001, reduced sell surface level of expression of the cysteinyl leukotriene receptor 2 results showed that p=.002. I found that the knockdown of cysteinyl leukotriene receptor 2 results in prothrombotic thrombocytes by using flow cytometry p=.0001.
Phenotypic Morphological Plasticity Induced by Environmental Salt Stress in the Brine Shrimp, Artemia franciscana
Phenotypic plasticity is the ability of an organism to express different phenotypes in response to biotic or abiotic environmental cues. The ability of an organism to make changes during development to adjust to changes in its environment is a key to survival. Sexually reproducing organisms that have short life cycles and that are easy to raise in the laboratory are more conducive for developmental phenotypic plasticity. Considerable research has already been carried out on the brine shrimp, Artemia franciscana, regarding its morphology due to changing salinities. There is, however, little research considering subsequent generations and how there morphology might be affected by parental experiences. This study has examined: 1) the morphological effects of different rearing regimes of different salinity levels, and 2) the epigenetic transgenerational transfer of these morphological traits in A. franciscana. Measurements included rate of growth (as measured by instar), body size, body length, and other morphological traits. A gradual increase to more hyperosmotic conditions during development produced brine shrimp that were larger in size and also more developmentally advanced. Salinity stress experienced by adults had increased the growth rate in the F1 offspring of A. franciscana. Collectively, these data indicate that Artemia franciscana is a tractable model for investigating phenotypic plasticity. These findings have added to the ever-growing field of developmental phenotypic plasticity while also providing more information on the natural history and adaptive abilities of A. franciscana.
A High-fat Meal Alters Post-prandial mRNA Expression of SIRT1, SIRT4, and SIRT6
Sirtuins (SIRT) regulate the transcription of various genes involved in the development of diet-induced obesity and chronic disease; however, it is unknown how they change acutely following a high-fat meal. The purpose of this study was to determine the effect of a high-fat meal (65% kcals/d; 85% fat recommendation), on SIRT1-7 mRNA expression in blood leukocytes at 1, 3, and 5-h post-prandial. Men and women (N=24) reported to the lab following an overnight fast (>12H). Total RNA was isolated and reverse transcribed prior to using a Taqman qPCR technique with 18S rRNA as a normalizer to determine SIRT1-7 mRNA expression. An additional aliquot of serum was used to measure triglycerides. Data was analyzed using a RM ANOVA with P<0.05. Triglycerides (P<0.001; 124%) peaked at 3-h. SIRT 1 (P=0.004; 70%), and SIRT 6 (P=0.017; 53%) decreased expression at 3-h. SIRT4 (P=0.024) peaked at 5H relative to baseline (70%) and 3-h (68%). To our knowledge, this is the first study to report that consumption of a high-fat meal transiently alters SIRT mRNA expression consistent in a pattern that mirrors changes in serum triglycerides. Decrease in expression of SIRT1 and SIRT6 combined with an increased SIRT4 would be consistent with an increase in metabolic disease risk if maintained on a chronic basis.
The Effect of Post-exercise Ethanol Consumption on the Acute Hormonal Response to Heavy Resistance Exercise in Women
The purpose of this study was to examine the hormonal response to acute ethanol ingestion following a bout of heavy resistance exercise in women. Eight resistance trained women completed two identical acute heavy resistance exercise tasks (AHRET). From 10-20 minutes post-AHRET, participants consumed either a grain ethanol or a placebo beverage. Blood was collected before (PRE) and immediately after the AHRET (IP) and then every 20 minutes for five hours. Blood collected after beverage ingestion was pooled into 3 batches (phases: 20-40 minutes, 60-120 minutes, and 140-300 minutes post-exercise) and analyzed for serum total testosterone (TT), free testosterone (FT), insulin-like growth factor-I (IGF-I), human growth hormone (GH), cortisol (COR), and estradiol (E2) concentrations. Circulating concentrations of TT were significantly greater at P20-40 than at PRE, P60-120, and P140-300. Circulating concentrations of FT were significantly greater at P20-40 than at all other times. Circulating concentrations of GH were significantly greater at IP than at PRE, P60-120, and P140-300. Circulating concentrations of COR were significantly greater at P20-40 than at all other times. Additionally, COR concentrations at P140-300 were significantly lower than at all other times. Circulating concentrations of IGF-1 were significantly greater at P20-40 than at P60-120 and P140-300. Circulating concentrations of E2 were significantly greater at P20-40 than at all other times. In summary, the present study demonstrated an acute modulation of the neuroendocrine milieu following a heavy resistance exercise bout in women. Ethanol ingestion appeared to have no significant effect on the characteristics of acute hormonal augmentation in TT, FT, GH, COR, IGF-1, or E2.
Cytotoxicity and Functional Toxicity of Mefloquine and the Search for Protective Compounds
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 …
Neurotoxicity of the Industrial Solvent 4-Methylcyclohexanemethanol: Involvement of the GABA Receptor
A recent chemical spill of 4-Methylcyclohexanemethanol (4-MCHM) in West Virginia left 300,000 people without water. Officials claimed that this compound is not lethally toxic, but potentially harmful if swallowed or inhaled, and can cause eye and skin irritation. Sittig's Handbook of Toxic and Hazardous Chemical Carcinogens reports high exposures from skin contact or inhalation may cause damage to the heart, liver, kidneys, and lungs, and may result in death. However, no quantitative data seem to exist and no references can be found on neurotoxicity. We have investigated the neurotoxicity of 4-MCHM using mammalian nerve cell networks grown on microelectrode arrays. Network spontaneous activity from multiple units (range 48 – 120 per network) were used as the primary readout. Individual units were followed based on spike waveforms digitized at 40 kHz (Plexon MNAP system). Dose response curves show the effective inhibitory concentration at 50 percent decrease (EC50) to average 27.4 microM SD±6.17. However, in the presence of 40 microM bicuculline, a competitive GABAA antagonist, the EC50 shifts to 70.63uM SD ±4.3; implying that early, low concentration exposures to 4-MCHM involve GABA activation. Initial activity loss occurs without active unit loss (defined as 10 or more template threshold crossing per min), indicating functional interference with spike production. Full recovery has not been seen at concentrations above 130 microM, unless the culture was given bicuculline. Direct exposure to 400uM results in immediate, irreversible loss of spike production, followed by necrosis of glia and neurons.
A Study of Some Congenital Anomalies of the Teeth
Anomalies of the teeth include a variety of expression such as: anodontia, hyperdontia, peg teeth. opalescent dentine, mottled enamel, and other imperfections of the teeth. This paper is concerned primarily with anodontia in human subjects, although other abnormalities of interest are mentioned. Insufficient datum is the principal difficulty encountered in determining the mechanism and means of transmission in human genetics. Small families, slow breeding, and inability to subject humans to experimentation account, in part, for this difficulty. The lack of reliable data is the second major difficulty in determining the mode of transmission. Often normal or unaffected members of a family are forgotten due to insignificance placed on them--thus an untrue pedigree is the result. The fact that one must rely on information from one or two members of a family lends itself to error. Therefore, an absolutely accurate determination of Mendelian ratios in human genetics would be impossible. This error, however, can be reduced by the gathering and compilation of large numbers of family pedigrees from which calculations can be made and results determined which will reveal the mode of transmission of various hereditary traits. With this view in mind, this paper is presented, first, to add several more cases to the literature, and, second, by analysis of the pedigrees to determine the possible modes of transmission of these anomalies.
Use of Remote Sensing for Cover Type Interpretation Over the Ray Roberts Reservoir Area
As part of a pre-impoundment study for the Ray Roberts Reservoir Area, Landsat-5 multi-spectral scanner (MSS) imagery was used for cover type interpretation. This research was concerned with analysis techniques for MSS images and a comparison of results obtained using computer assisted interpretatin of MSS images and a comparison of results obtained using computer assisted interpretation of MSS images with those obtained using manual interpretation of false color infrared (CIR) photographs.
Investigations on Abundance, Habits, and Distribution of Amphibians and Reptiles of Denton County, Texas
The purpose of the present study of the herpetofauna was to obtain additional information regarding the vertebrates of Denton County, and to produce a well-preserved, cataloged collection of the amphibians and reptiles for the Museum of Zoology, North Texas State University. An understanding of the vertebrate life of the county also involves an investigation of the habitats within the county that may, in part, count for the distribution of these animals. It is well recognized that the environmental areas of the county have altered vastly during the last one hundred years. This alteration is due largely to agriculture and industry. However, there are adequate numbers of natural environments, as well as newly created ones that may contribute to the distribution of the vertebrates at the present time. Therefore, the problem not only concerned the collection of specimens, but also the identification, abundance classification, general habitat classifications, and county distribution.
Fumarase From Ascaris Suum: Partial Purification and Characterization
One molecular form of fumarase from Ascaris suum was demonstrated by cellulose acetate electroporesis and isoelectric focusing. The enzyme was partially purified by ammonium sulfate fractionation and ion-exchange chromatography to a specific activity of 49 units per mg protein. Enzymatic assay of the partially purified by ammonium sulfate fractionation amd ion-exchange chromatography to a specific activity of 49 units per mg protein. Enzymatic assay of the partially purified preparation showed glyceraldehyde-3-phosphate dehydrongenase to be the major preparative contaminant.
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