The effect of gregarine parasite infections, age, and diet on calling song structure and mating behaviour in the Texas field cricket, Gryllus integer

In the past ten years, many researchers have focussed their attention on parasites regarding the role they may play in causing variations in male secondary sexual traits and subsequent effects on female choice. Male age has also been suggested to be an important factor in female choice if old age reflects superior genes. This study investigated the effects that gregarine gut parasites, age, and diet have on the calling and mating behaviour of the male Texas field cricket, Gryllus integer. Male calling songs were recorded in the laboratory using a Digital Signal Processing Network. The song parameters measured were: pulse rate, pulse width, burst duration, pulses per burst, interburst interval, and percent missing pulses. The effects of parasite load and age on the various calling song parameters was investigated in crickets that were fed two different diets varying in nutritional quality. None of the calling song parameters were affected by either parasite load or age in either diet grou p. Courtship behaviour was ob served and recorded using an Eventlog recorder on an IBM computer in the laboratory. Females mated equally with paras(tized and unparasitized males and with old and young males The total duration and proportion of time spent performing each of 9 courtship displays were recorded for males on each diet. Only one display was affected by parasite load. Highly parasitized males fed the nutritionally inferior diet juddered for a proportionately shorter time than males with low parasite loads. Also, older males performed juddering and shaking antennae proportionally longer and juddering and raising wings for longer durations than younger males. Males that successfully mated were observed for performance of 8 post-copulatory guarding behaviour displays. None of the guarding behaviours were affected by parasite load. However, one display was affected by age, with older males performing guard turning for shorter durations than younger males. Results are discuss,ed in terms of the influence of parasites and age on female choice.

The effect of viticultural and oenological treatments on fruit and wine composition of Chardonnay musqué /

The effect of viticultural and oenological treatments on fruit and wine composition of Chardonnay musque Study I: Effect ofveraison leafremoval and cluster thinning A one-year study was performed analysing die effects of leaf removal, cluster thinning, yeast strain selection, and enzyme usage on the chemical composition and sensory properties of Chardonnay musque wine. A number of substantial differences were found between treatments in °Brix, TA, pH, and in free and potentially volatile terpene concentrations. Greatest variations in sensory attributes were created however through use of different viticultural practices.Study II: Effect ofcluster thinning timing A two year study was conducted investigating the effect of cluster thinning timing, yeast strain selection, and enzyme usage on the chemical composition and sensory attributes of Chardonnay musque wine. Time of thinning was found to impact °BrLx, titratable acidit}% pH, and free and potentially volatile terpene concentrations, as well as, a number of yield parameters.Yeast strain selection and enzyme usage also impacted wine composition, andwas found to exhibit a greater effect on sensory properties than application of cluster thinning.

Effect of pH on non-photochemical quenching in spinach photsystem 2 particles as measured by picosecond fluorescence decay kinetics

Single photon timing was used to study picosecond chlorophyll a fluorescence decay kinetics of pH induced non-photochemical quenching in spinach photosystem 2 particles. The characteristics of this quenching are a decrease in chlorophyll a fluorescence yield as well as a decrease in photochemistry at low pH. Picosecond kinetics of room temperature fluorescence temporally resolve the individual components of the steady state fluorescence yield into components that are related to primary energy conversion processes in photosystem 2. Four components were resolved for dark adapted (Fo), light saturated (Fm), and chemically reduced (Nadithionite) photosystem 2 reaction centres. The fastest and slowest components, indicative of energy transfer to and energy capture by the photosystem 2 reaction centre and uncoupled ("dead") chlorophyll, respectively, were not affected by changing pH from 6.5 to 4.0. The two intermediate components, indicative of electron transfer processes within the reaction centre of photosystem 2, were affected by the pH change. Results indicate that the decrease in the steady state fluorescence yield at low pH was primarily due to the decrease in lifetime and amplitude of the slower of the intermediate components. These results imply that the decrease in steady state fluorescence yield at low pH is not due to changes in energy transfer to and energy capture by the photosystem 2 reaction centre, but is related to changes in charge stabilization and charge recombination in the photosystem 2 reaction centre.

Biological effects of resveratrol on skeletal muscle cells

Resveratrol, a polyphenol found in red wine, has been reported to have antithrombotic, antiatherogenic, and anticancer properties both in vitro and III VIVO. However, possible antidiabetic properties of resveratrol have not been examined. The objective of this study was to investigate the direct effects of resveratrol on basal and insulin-stimulated glucose uptake and to elucidate its mechanism of action in skeletal muscle cells. In addition, the effects of resveratrol on basal and insulin- stimulated amino acid transport and mitogenesis were also examined. Fully differentiated L6 rat skeletal muscle cells were incubated with resveratrol concentrations ranging from 1 to 250 IlM for 15 to 120 min. Maximum stimulation, 201 ± 8.90% of untreated control, (p<0.001), of2eH] deoxy- D- glucose (2DG) uptake was seen with 100 IlM resveratrol after 120 min. Acute, 30 min, exposure of the cells to 100 nM insulin stimulated 2DG uptake to 226 ± 12.52% of untreated control (p<0.001). This appears to be a specific property of resveratrol that is not shared by structurally similar antioxidants such as quercetin and rutin, both of which did not have any stimulatory effect. Resveratrol increased the response of the cells to submaximal insulin concentrations but did not alter the maximum insulin response. Resveratrol action did not require insulin and was not blocked by the protein synthesis inhibitor cycloheximide. L Y294002 and wortmannin, inhibitors of PI3K, abolished both insulin and resveratrolstimulated glucose uptake while phosphorylation of AktlPKB, ERK1I2, JNK1I2, and p38 MAPK were not increased by resveratrol. Resveratrol did not stimulate GLUT4 transporter translocation in GLUT4cmyc overexpressing cells, in contrast to the significant translocation observed with insulin. Furthermore, resveratrol- stimulated glucose transport was not blocked by the presence of the protein kinase C (PKC) inhibitors BIMI and G06983. Despite that, resveratrol- induced glucose transport required an intact actin network, similar to insulin. In contrast to the stimulatory effect seen with resveratrol for glucose transport, e4C]methylaminoisobutyric acid (MeAIB) transport was inhibited. Significant reduction of MeAIB uptake was seen only with 100uM resveratrol (74.2 ± 6.55% of untreated control, p<0.05), which appeared to be maximum. In parallel experiments, insulin (100 nM, 30 min) increased MeAIB transport by 147 ± 5.77% (p<0.00l) compared to untreated control. In addition, resveratrol (100 JlM, 120 min) completely abolished insulin- stimulated amino acid transport (103 ± 7.35% of untreated control,p>0.05). Resveratrol also inhibited cell proliferation in L6 myoblasts with maximal inhibition of eH]thymidine incorporation observed with resveratrol at 50 J.LM after 24 hours (8 ± 1.59% of untreated control, p

Investigations on the production of long chain fatty acids in Choanephora cucurbitarum (Berk.

The fatty acid composition of the total cellular lipids of Choanephora cucurbitarum incubated for 96 hrs on either glucose-ammonium sulfate or malt-weast extract media was determined. The major fatty acids were palmitic, palmitoleic, stearic and linoleic acids. The saturated fatty acid possessing the longest acyl chain was stearate (C 18:0). The presence of glutamic acid (2.0 x 10-1% or 1.36 x la-2M) in either of the above growth media resulted in increase in percent of 1f-linolenic acid, decrease in percent of linoleic ~iCid and appearance of a new series of fatty acid> C ~8 e.g. C ",,,,'V' C2k:O, C26,O. The addition of glutamic acid had no effect on the lipid yield but slightly decreased the degree of unsaturation. Compounds which duplicated the effect of glutamic acid were acetate, malate, citrate, succinate, 0( -ketoglutarate, prOline, -y -aminobutyric acid and glucose (3%) but not aspartic acid or alanine. ~o correlation was found between glutamic acid pool concentration and the presence in the growth medium of those compounds which stimulate long chain fatty acid production. Four hours of incubation with 27 JJ 1-1 glutamate supported the production of long chain fatty acids. This stimulation is inhibited if 272 .u M isophthalic acid is added with 27 AJ M glutamate. But, long chain fatty acids were detected when 27 JJ M eX -ketoglutarate is also present in the incubation mixture. Five hours of incubation with 100 ,Mg/ml of cycloheximide resulted in over 9CY/o inhibition of cytoplasmic :protein synthesise Glutamate (27 .uM) enhanced the synthesis of long chain fatty acids under these conditions. These findings are discussed in an attempt to provide a plausible explanation COmmon to compounds that support the production of long chain fatty acids.

The effect of lysobisphosphatidic acid (LBPA) on α-tocopherol transfer protein (α-TTP) binding to lipid membranes

The a-tocopherol transfer protein (a-TTP) is responsible for the retention of the atocopherol form of vitamin E in living organisms. The detailed ligand transfer mechanism by a-TTP is still yet to be fully elucidated. To date, studies show that a-TTP transfers a-tocopherol from late endosomes in liver cells to the plasma membrane where it is repackaged into very low density lipoprotein (VLDL) and released into the circulation. Late endosomes have been shown to contain a lipid known as lysobisphosphatidic acid (LBP A) that is unique to this cellular compartment. LBPA plays a role in intracellular trafficking and controlling membrane curvature. Taking these observations into account plus the fact that certain proteins are recruited to membranes based on membrane curvature, the specific aim of this project was to examine the effect of LBP A on a-TTP binding to lipid membranes. To achieve this objective, dual polarization interferometry (DPI) and a vesicle binding assay were employed. Whilst DPI allows protein binding affinity to be measured on a flat lipid surface, the vesicle binding assay determines protein binding affinity to lipid vesicles mimicking curved membranes. DPI analysis revealed that the amount of a-TTP bound to lipid membranes is higher when LBPA is present. Using the vesicle binding assay, a similar result was seen where a greater amount of protein is bound to large unilamellar vesicles (LUV s) containing LBP A. However, the effect of LBP A was attenuated when small unilamellar vesicles (SUVs) were replaced with LUVs. The outcome of this project suggests that aTTP binding to membranes is influenced by membrane curvature, which in turn is induced by the presence of LBP A.

Adenovirus-based exogenous gene expression in mammalian cells

Thesis (Ph.D.)--Brock University, 2010.

Elucidation of odour-potent compounds and sensory profiles of Vidal blanc and Riesling icewines from the Niagara Peninsula : effect of harvest date and crop level

I t was hypothesized that the freeze/thaw cycles endured by icewine grapes would change their chemical composition, resulting in unique chemical fingerprint and sensory properties, and would be affected by harvest date (HD) and crop level (CL). The objectives were: 1) to identify odour-active compounds using gas chromatographic and sensory analysis; 2) to determine the effect of CL and HD on these compounds; 3) to determine the icewine sensory profiles; 4) to correlate analytical and sensory results for an overall icewine profile. CharmAnalysis™ determined the Top 15 odour-potent compounds in Vidal and Riesling icewine and table wines; 24 and 23 compounds, respectively. The majority of the compounds had the highest concentrations in the icewines compared to table wines. These compounds were used as the foundation for assessing differences in icewine chemical profiles from different HD and CL. Vidal and Riesling icewine were made from grapes picked at different HD; HI : 19 December; H2: 29 December; H3: 18 January; H4: 11 February (Vidal only). HI wines differed from H3 and H4 wines in both Vidal and Riesling for aroma compounds and sensory profiles. - Three·CL [control (fully cropped), cluster thin at fruit set to one basal cluster/shoot (TFS), and cluster thin at veraison to one basal cluster/shoot (TV)] were evaluated for Riesling and Vidal cultivars over two seasons. Vidal icewines had the highest concentration of aroma compounds in the control and TV icewines in 2003 and in TFS icewines in 2004. In Riesling, most aroma compounds had the highest concentration in the TV icewines and the lowest concentration in the TFS wine for both years. The thinned treatments were associated with almost all of the sensory attributes in both cultivars, both years. HD and CL affected the chemical variables, aroma compounds and sensory properties of Vidal and Riesling icewines and freeze/thaw events changed their sensory profile. The most odour-potent compounds were p-damascenone, cis-rose oxide, 1- octen-3-ol, 4-vinylguaiacol, ethyl octanoate, and ethyl hexanoate. The role of Pdamascenone as a marker compound for icewine requires further investigation. This research provides a strong foundation for the understanding the odour-active volatiles and sensory profiles important to icewine.

The Effect of Skate Blade Radius of Contour and Radius of Hollow on Skating Performance in Male Ice Hockey Players

The primary purpose of this study was to investigate the effect of skate blade shape on skating performance. A secondary purpose was to evaluate if a change in hollow shape can create additional effects on skating performance. Thirty-seven male ice hockey players (age=18 years, SD=3.4) participated. The intervention consisted of four sharpening trials assessed using three on-ice tests. Participant feedback was also assessed using a Likert scale questionnaire. Statistical analysis included within-subject repeated measures MANOVA of trial by skating variables (p≤0.05). Results revealed Contour 1 enhanced performance compared to baseline on six variables at varsity level and five variables at midget level. Contour 1 enhanced performance compared to Contour 2 on six variables at the varsity and midget levels. Contour 1 also scored highest on the feedback questionnaire. Findings of this study indicate that contouring is a necessary practice to achieve optimal skating performance.

A Multi-Objective Genetic Algorithm with Side Effect Machines for Motif Discovery

Understanding the machinery of gene regulation to control gene expression has been one of the main focuses of bioinformaticians for years. We use a multi-objective genetic algorithm to evolve a specialized version of side effect machines for degenerate motif discovery. We compare some suggested objectives for the motifs they find, test different multi-objective scoring schemes and probabilistic models for the background sequence models and report our results on a synthetic dataset and some biological benchmarking suites. We conclude with a comparison of our algorithm with some widely used motif discovery algorithms in the literature and suggest future directions for research in this area.

Investigating the role of apoptosis regulator EndoG on exogenous DNA uptake, stability, replication and recombination

Endonuclease G (EndoG) is a well conserved mitochondrial nuclease with dual lethal and vital roles in the cell. It non-specifically cleaves endogenous DNA following apoptosis induction, but is also active in non-apoptotic cells for mitochondrial DNA (mtDNA) replication and may also be important for replication, repair and recombination of genomic DNA. The aim of our study was to examine whether EndoG exerts similar activities on exogenous DNA substrates such as plasmid DNA (pDNA) and viral DNA vectors, considering their importance in gene therapy applications. The effects of EndoG knockdown on pDNA stability and levels of encoded reporter gene expression were evaluated in the cervical carcinoma HeLa cells. Transfection of pDNA vectors encoding short-hairpin RNAs (shRNAs) reduced levels of EndoG mRNA and nuclease activity in HeLa cells. In physiological circumstances, EndoG knockdown did not have an effect on the stability of pDNA or the levels of encoded transgene expression as measured over a four day time-course. However, when endogenous expression of EndoG was induced by an extrinsic stimulus (a cationic liposome transfection reagent), targeting of EndoG by shRNA improved the perceived stability and transgene expression of pDNA vectors. Therefore, EndoG is not a mediator of exogenous DNA clearance, but in non-physiological circumstances it may non-specifically cleave intracellular DNA regardless of its origin. To investigate possible effects of EndoG on viral DNA vectors, we constructed and evaluated AdsiEndoG, a first generation adenovirus (Ad5 ΔE1) vector encoding a shRNA directed against EndoG mRNA, along with appropriate Ad5 ΔE1 controls. Infection of HeLa cells with AdsiEndoG at a multiplicity of infection (MOI) of 10 p.f.u./cell resulted in an early cell proliferation defect, absent from cells infected at equivalent MOI with control Ad5 ΔE1 vectors. Replication of Ad5 ΔE1 DNA was detected for all vectors, but AdsiEndoG DNA accumulated to levels that were 50 fold higher than initially, four days after infection, compared to 14 fold for the next highest control Ad5 ΔE1 vector. Deregulation of the cell cycle by EndoG depletion, which is characterized by an accumulation of cells in the G2/M transition, is the most likely reason for the observed cell proliferation defect. The enhanced replication of AdsiEndoG is consistent with this conclusion, as Ad5 ΔE1 DNA replication is intimately related to cell cycling and prolongation or delay in G2/M greatly enhances this process. Furthermore, infection of HeLa with AdsiEndoG at MOI of 50 p.f.u./cell resulted in an almost complete disappearance of viable, adherent tumour cells from culture, whereas almost a third of the cells were still adherent after infection with control Ad5 ΔE1 vectors, relative to the non-infected control. Therefore, targeting of EndoG by RNAi is a viable strategy for improving the oncolytic properties of first generation adenovirus vectors. In addition, AdsiEndoG-mediated knockdown of EndoG reduced homologous recombination between pDNA substrates in HeLa cells. The effect was modest but, nevertheless demonstrated that the proposed role of EndoG in homologous recombination of cellular DNA also extends to exogenous DNA substrates.

Investigating the Biological Effects of Rosemary (Rosmarinus officinalis L.) Extract on Skeletal Muscle Glucose Uptake

Skeletal muscle (SKM) is the most important tissue in maintaining glucose homeostasis and impairments in this tissue leads to insulin resistance (IR). Activation of 5’ AMP-activated kinase (AMPK) is viewed as a targeted approach to counteract IR. Rosemary extract (RE) has been reported to decrease blood glucose levels but its effects on SKM are not known. We hypothesized that RE acts directly on SKM to increase glucose uptake (GU). We found an increase in GU (184±5.07% of control, p<0.001) in L6 myotubes by RE to levels similar to insulin and metformin. Carnosic acid (CA) and rosmarinic acid (RA), major polyphenols found in RE, increased GU. RE, CA, and RA significantly increased AMPK phosphorylation and their effects on GU was reduced by an AMPK inhibitor. Our study is the first to show a direct effect of RE, CA and RA on SKM GU by a mechanism that involves AMPK activation.

The role of retinoic acid in long-term memory formation and synaptic plasticity in the mollusc Lymnaea stagnalis

The active metabolite of vitamin A, retinoic acid (RA), is involved in memory formation and hippocampal plasticity in vertebrates. A similar role for retinoid signaling in learning and memory formation has not previously been examined in an invertebrate species. However, the conservation of retinoid signaling between vertebrates and invertebrates is supported by the presence of retinoid signaling machinery in invertebrates. For example, in the mollusc Lymnaea stagnalis the metabolic enzymes and retinoid receptors have been cloned from the CNS. In this study I demonstrated that impairing retinoid signaling in Lymnaea by either inhibiting RALDH activity or using retinoid receptor antagonists, prevented the formation of long-term memory (LTM). However, learning and intermediate-term memory were not affected. An additional finding was that exposure to constant darkness (due to the light-sensitive nature of RA) itself enhanced memory formation. This memory-promoting effect of darkness was sufficient to overcome the inhibitory effects of RALDH inhibition, but not that of a retinoid receptor antagonist, suggesting that environmental light conditions may influence retinoid signaling. Since RA also influences synaptic plasticity underlying hippocampal-dependent memory formation, I also examined whether RA would act in a trophic manner to influence synapse formation and/or synaptic transmission between invertebrate neurons. However, I found no evidence to support an effect of RA on post-tetanic potentiation of a chemical synapse. Retinoic acid did, however, reduce transmission at electrical synapses in a cell-specific manner. Overall, these studies provide the first evidence for a role of RA in the formation of implicit long-term memories in an invertebrate species and suggest that the role of retinoid signaling in memory formation has an ancient origin.

Investigation of the Biological Effects of Rosemary (Rosmarinus Officinalis L.) Extract in Human Lung Cancer Cells

Cancer cells display enhanced growth rates and a resistance to apoptosis. Lung cancer accounts for the most cancer related deaths and non-small cell lung cancer (NSCLC) represents an aggressive form of lung cancer, accounting for almost 80% of all lung cancer cases. The phytochemical rosemary extract (RE) has been reported to have anticancer effects in vitro and in vivo however, limited evidence exists regarding the effects of RE and its polyphenolic constituents carnosic acid (CA) and rosmarinic acid (RA) in lung cancer. The present study shows RE, CA and RA inhibit lung cancer cell proliferation and survival in various NSCLC cell lines and that CA and RA interact synergistically to inhibit cell proliferation. Moreover RE, CA and RA are capable of altering activation and/or expression of Akt, ERK and AMPK, signaling molecules which regulate cell proliferation and survival. RE shows potential as an anticancer agent and should be further investigated.