This diploma thesis is focused on study of antimutagenic properties in selected kinds of honey and propolis. In honey extracts compounds with antimutagenic and antioxidant effect were analysed by spectrophotometry, RP/HPLC/UV-VIS, HPLC/PDA and on-line LC/MS. Further, effect of long-term storage on active compound levels was studied. Antimutagenic activity was tested by simple eukaryotic system - yeast strain Saccharomyces cerevisiae D7. The highest antimutagenic effect and simultaneously the highest stability of antimutagenity values was found in several kinds of multi-floral honey, rape seed honey, honey from eucalypt and orange flowers and in honeydew honey. Samples obtained from trade network exhibited at general lower antimutagenity values when compared with samples from bee-keeper. The highest content of total phenolics was detected in honey with royal yelly, the lowest content was measured in acacia honey. The highest values of total flavonoids exhibited buckwheat and eucalypt flower honey. Total phenolic content was substantially changed during long-term storage (decrease about 50 -70 %), while total flavonoid content was stable and no significant changes during storage were observed. Honey with high antimutagenity values exhibited also higher phenolic and predominantly flavonoid content and high antioxidant activity. Because of relative stability and low changes in most of honey samples during storage it can be concluded that honey belong to foodstuffs which are able to conserve their positive nutritive properties for a long time.

No description available.

Over 270 million people play soccer worldwide, and its popularity grows every day. In team sport exercise, fatigue may result from numerous factors including limited fuel, depleted energy stores and production of compounds that promote an inflammatory response. While inflammation is an essential mechanism for repairing damaged muscle tissue with exercise, prolonged inflammation leads to increased production of reactive oxygen species that can damage cell membranes, muscle, and signaling proteins. To prevent this response and improve performance, athletes are increasingly looking to nutritional interventions. Carbohydrate and antioxidant supplementation have both shown evidence of producing an ergogenic effect and attenuating inflammation and oxidative stress with prolonged endurance exercise. Less is known about how these interventions may influence intermittent, high-intensity exercise characteristic of soccer. In particular, this exercise presents a unique challenge in that opportunities for nutrient intake are limited to pre-game and half-time. In our first study, we had 10 male collegiate soccer players perform a 90-min. soccer-simulation test, that we developed, which was followed by a progressive shuttle run (PSR) test to exhaustion. They consumed a honey-sweetened beverage (H), a sports drink (S), or a placebo (P) before and half-way through the protocol. Both H and S provided 1.0 g·kg⁻¹ carbohydrate and ~17.6 mL·kg⁻¹ total volume for each trial. Overall, the test resulted in increased fatigue and production of inflammatory markers and antioxidant capacity. There was no significant difference between treatments for any performance measure. Mean times for a high intensity run and rating of perceived exertion increased with time, and there was an overall decrease in PSR time compared to baseline (-22.9%). There was a rise in glucose (15.6%), IL-6 (548%), IL-1ra, IL-10 (514%) and ORAC (15%) post-test but no change in cortisol. Insulin was significantly lower by 1 h-post. IL-1ra levels increased post-test for H (25.8%), S (65.5%), and P (63.9%), but the change for H was less than the other treatments. No treatment effects for the other blood measures were observed. The lack of an ergogenic effect of carbohydrate on soccer performance calls into question the benefit of supplementation at a frequency typical of a regulation soccer match in highly trained athletes with adequate energy stores. Since acute carbohydrate ingestion in the first study did not attenuate some markers of inflammation (e.g. IL-6), we chose to focus on an alternative theory for the rise in inflammatory markers with strenuous exercise in our second study. One aspect of soccer, repeated sprinting, results in increased ROS production partially through the activation of the enzyme xanthine oxidase (XO). Quercetin, a flavonol in plants that has shown some ergogenic effects with endurance exercise, inhibits XO in vitro. The effect of quercetin on team sport exercise had not been studied. We gave recreationally active males a commercial sports drink (S) or S + 500 mg of quercetin (Q) 2x/d for 1 wk prior to a repeated sprint test (RST). Sprint times increased (5.9%) for both treatments as did plasma XO activity (47%), IL-6 (77%), and uric acid (25%) from pre-test to post-test. Q supplementation did not attenuate plasma XO activity or IL-6 and actually increased one calculated index of fatigue, percent fatigue decrement (5.1%- Q and 3.8%- P). These findings add to the growing body of literature that quercetin supplementation does not attenuate exercise-induced inflammation and oxidative stress in vivo. Collectively, this research has practical implications for sports drink companies who are exploring the use of flavonoid compounds in product formulation. Specifically, they should reconsider adding quercetin to their beverages if they are marketing to team sport athletes. Also, soccer players should be made aware that, at ingestion frequencies typical of a soccer match, they may not expect a significant performance benefit from acute carbohydrate supplementation. / Ph. D.

No description available.

The loss of managed honey bee colonies, resulting from their unintentional exposure to pesticides, is a topic of concern for the agricultural and apicultural industry. Current methods for reducing pesticide exposure to bees involve the application of pesticides before crop bloom or in the evening when foraging bees are less likely to be exposed to these applications. There is an urgent need for additional protection procedures to reduce the annual losses of managed bee colonies. Another method for protecting these pollinators is the use of chemical deterrents to reduce the interaction times of foraging bees with pesticide-treated crops. Historically, insect repellents (IRs) have been used to prevent the spread of deadly human diseases by arthropod vectors. However, it has been shown that bees can be repelled from pesticide-treated crops using DEET and bee pheromonal compounds. Here, I report the toxicological and deterrent effects of bee pheromonal compounds, as well as the deterrent effects of heterocyclic amines (HCAs) on bees. The results of this study indicate that the bee pheromonal compounds, at 8, 20, 60 and 100% concentrations, are toxic to bees and inhibit the feeding of bees within a confined space. Additionally, the pheromonal compounds and the HCAs are as efficacious as DEET in deterring bees from treated food sources. The HCA piperidine was observed to effectively deter bee foragers from a sugar feeder in a high-tunnel experiment as well as from melon flowers and knapweed in field experiments. Electroantennogram recordings were conducted to verify an olfactory response of the bees to the tested compounds. Pheromonal compounds were readily detected by bee antennae; whereas, the HCAs did not elicit significant responses in the bee antennae. These data suggest that bee pheromonal compounds, as well as HCAs, may serve as candidates for the further investigation as repellents to protect bees from unintentional pesticide exposures. / Ph. D. / The loss of managed honey bee colonies, resulting from their unintentional exposure to pesticides, is a topic of concern for the agricultural and apicultural industry. Current methods for reducing pesticide exposures to bees involve the application of pesticides before crop bloom or in the evening when foraging bees are less likely to be exposed to these applications. There is an urgent need for additional protection procedures to reduce the annual losses of managed bee colonies. One method for protecting these pollinators is use of chemical deterrents to reduce the interaction times of foraging bees with pesticide-treated crops. Insect repellents (IRs) primarily have been used for biting and blood-feeding arthropod pests. However, it has been shown that bees can be repelled from pesticide-treated crops using DEET and bee pheromonal compounds. Here, I report the toxicological and deterrent effects of bee pheromonal compounds as well as the deterrent effects of heterocyclic amines (HCAs) on bees. The goals of this study were to: 1) examine the toxicological effects of the pheromonal compounds on bees, 2) develop a laboratory testing protocol for evaluating the deterrent effects of pheromonal bee compounds and heterocyclic amines (HCAs) to bees, 3) evaluate the deterrent effects of HCAs to bees using high-tunnel and semi-field experimentations, and 4) characterize the olfactory responses of bees to the above compounds.

The waggle dance of the honey bee (Apis mellifera L.) is perhaps the most celebrated animal communication behavior. With a waggle dance, a forager bee who has discovered a profitable resource on the landscape, usually floral nectar or pollen, can inform her nestmates of its location and recruit them to exploit it by communicating both a distance and a direction. Since Karl von Frisch described the waggle dance in 1942, scientific exploration of the dance has exploded into the realms of its structure, function, role in the regulation of collective foraging in the context of the hive as a super-organism, and even its utility as a study system for understanding sublethal behavioral effects of pesticide exposure. This dissertation presents three novel studies of the waggle dance. In the first, we asked whether consistent inter-bee differences (i.e., individuality) in a waggle dance distance - duration calibrations could affect communication success. In the second, we characterized the networks of recruitment arising from waggle dance communications and explored the role of the aforementioned individuality in network formation. In the third, we tested whether sublethal exposure to glyphosate (GLY), the most-applied herbicide in the world, could affect foraging, recruitment, or the levels and balance of biogenic amines in the bee brain. In each of these experiments, we housed bees in clear-walled observation colonies and trained cohorts of bees to visit artificial feeders to record both foraging and recruitment data. In our first experiment, we found that individuality in waggle dance behavior does shape communication outcomes, indicating that individual-level behavioral differences should not be discounted as factors at work in eusocial insect societies. In the second, we present the first network density and dance burstiness data from in vivo bee networks, revealing that recruitment networks are sparse, and waggle dancers are partitioned into bursty and non-bursty behavioral types. In the third, we show that not only can sublethal GLY exposure reduce foraging, but it can also produce significant correlations between levels of the important insect neurotransmitter octopamine and its two biosynthetic precursors, tyramine and tyrosine, where levels in control bees were unrelated. The results of this dissertation research, while distinct by experiment, together emphasize the continuing usefulness and tractability of the honey bee colony as a system in which to study the role of individuality in animal communication and to better understand the threat posed by non-insecticidal pesticide chemistries to the planet's most economically impactful pollinator. / Doctor of Philosophy / One of the most famous and well-studied animal behaviors is the waggle dance of the honey bee. A honey bee's waggle dance works similarly to a Yelp review for a restaurant: a bee who has found a good food source, like a flower patch offering especially sweet nectar or high-quality pollen, can come back to the colony and recommend it to her nestmates with a dance. The waggle dance is even more specific than a Yelp review, however, in that it also gives instructions to find the food source, communicating both a distance and a direction so that dance followers can go out into the landscape and look for the food source themselves. Even though the waggle dance has been studied extensively since it was first described by Karl von Frisch in the 1940s, there are still unknowns about how it works, and how it might be impacted by certain stressors. This dissertation presents three different experiments aimed at shedding light on these unknowns. First, it has recently been shown that there are consistent differences between bees in the way they communicate distance in the dance, and we tested whether that between-bee individuality can affect the likelihood that two bees will communicate successfully. Second, we studied how information about a food source moves from bee to bee via the waggle dance to form a communication network. Specifically, we described how efficiently information moved from bee to bee, patterns of dancing behavior, and the role of that individuality in its formation. Third and lastly, we looked to see whether exposure to a weedkiller called glyphosate (GLY) could affect either honey bees' waggle dance or food-collecting behavior, as well as levels of certain neurotransmitters in their brains that are involved in those behaviors. In all three experiments, we collected our data by housing bees in a clear-walled observation hives that let us view and film their waggle dance behavior, and then training groups of bees to collect artificial nectar from a feeder station that we provided, so we could also observe them as they collected food. We found that individuality in waggle dance communication can indeed affect the likelihood of communication success between two given bees, where the likelihood of communication success is greater when the dancer communicates a farther distance to the food source than the follower would. In the second experiment, our study of the waggle dance communication network showed that (1) information does not flow from bee to bee very efficiently, and (2) bees either dance quite regularly or sporadically. As far as we know, we are the first to describe these aspects of the waggle dance communication network, which may be useful in the field of computing algorithms inspired by living organisms. Finally, our third experiment showed that mild GLY exposure not only reduced how frequently bees collected food from our feeder, but also changed the relative amounts of certain neurotransmitters in their brains. This result emphasizes the importance of understanding how weedkillers that are not meant to target beneficial insects like honey bees are actually affecting them, so that we can make better-informed decisions to protect honey bees and other good insects.

The decline of honey bee (Apis mellifera) colony numbers in recent years presents an economic and ecological threat to agriculture. One outstanding threat to honey bees is the unintended exposure to agricultural pesticides. Previous studies report that acute exposures to the common-use herbicide atrazine elicit oxidative stress in non-target insects; however, little information is currently available on the exposure risk of atrazine to honey bees. This project examined biochemical and molecular oxidative stress response markers of honey bees following laboratory and field treatments of atrazine. Laboratory experiments were conducted with honey bees exposed to increasing concentrations of atrazine for 24 h whereas hive experiments were conducted with bees exposed to one sub-lethal concentration of atrazine for 28 d. The overall antioxidant enzyme activities of atrazine-treated honey bees were decreased compared to the untreated honey bees in both the laboratory and hive experiments. After exposure to atrazine in the laboratory and field, semi-quantitative RT-PCR analysis of antioxidant-encoding genes reveals the differential expression of genes in atrazine-treated bees that are important for oxidative stress tolerance in the laboratory and field experiments. Here, we provide evidence that the laboratory and hive exposure of honey bees to the common-use herbicide atrazine results in oxidative stress responses that can compromise the health of bee colonies. The data will be discussed with regard to the protection of these pollinators against the untended exposure of agricultural pesticides. / Master of Science in Life Sciences / The pollination service provided by insects, primarily honey bees, is estimated to contribute approximately one-third of the diet consumed by the average American. Honey bees are vitally important pollinators due to their broad range of foraging activities and ease of husbandry within a managed colony. In recent decades, colony numbers have decreased in the developed areas of the planet and pesticide usage has been implicated in these losses. Atrazine is the second most commonly used agricultural herbicide in the country and has been linked to oxidative stress in beneficial insects in the past. Oxidative stress is the result of an uncontrolled build-up of reactive oxygen species in an aerobic organism. These reactive oxygen species are dangerous because they are capable of damaging proteins, DNA, and cell walls. Every aerobic organism also possesses antioxidant function which serves to prevent or counteract damage caused by reactive oxygen species. This study examined antioxidant enzyme activities and antioxidant-encoding gene expression levels, which were used as indicators of oxidative stress biomarkers, in honey bees exposed to atrazine in the laboratory and in the hive environment. Honey bees were exposed to atrazine at increasing concentrations in the laboratory for 24 h and at one environmentally relevant dose for 28 d in the hive. After exposure to atrazine in the laboratory and the hive, four out of five antioxidant enzyme levels of honey bees decreased which implied an increase in oxidative stress and a decrease in antioxidant defenses. Activity of one enzyme, lipid peroxidase, increased in honey bees after exposure to atrazine. Lipid peroxidase is the most common measure of cellular injury during oxidative stress, once again signifying an increase in reactive oxygen species production and oxidative stress. Expression levels of seven antioxidantencoding genes were examined in honey bees after atrazine exposure and expression levels of some genes changed compared to the untreated control and expression levels in some genes remained the same compared to the untreated control. These changes in antioxidant-encoding gene expression levels may imply an increase in oxidative stress due to exposure of the honey bees to atrazine. This study aimed to examine biomarkers of oxidative stress in honey bees exposed to the commonly used herbicide atrazine with the hope of raising awareness of harmful effects caused by atrazine and protecting these important pollinators from unintended exposure to agricultural pesticides.

The honey bee, Apis mellifera, provides about $15 billion USD in crop value each year in the U.S. alone in the form of pollination services. Since 2006, commercial beekeepers have reported an average annual overwintering loss of about 28.6% of all managed colonies. There are many factors that are thought to contribute to colony loss including bee-specific pests (e.g. the Varroa destructor mite), bee-specific pathogens (e.g. Nosema fungus), modern beekeeping practices, diminished genetic variability, poor queens, climate change, and exposure to agricultural pesticides. While not the single cause of colony loss, the neonicotinoid insecticides elicit sublethal effects to honey bees that could increase their sensitivities to other stressors that affect colony health. Previous studies found that honey bees have differential sensitivities to the neonicotinoid insecticide imidacloprid, which suggest a mechanism of tolerance to the insecticide in certain colonies. In this study, I examined the imidacloprid sensitivity of honey bees collected from different colonies. After determining a range of LC50 values in the tested colonies, I examined the metabolic detoxification activities of honey bees collected from two colonies that represented the highest and lowest LC50 values, between which there was a 36-fold difference in their LC50 values. I discovered that of the three main families of metabolic detoxification enzymes, general esterases, cytochrome P450 monooxygenases, and glutathione S-transferases (GSTs), a reduction of GST activity with diethyl maleate (DEM) significantly increased imidacloprid-mediated mortality to the honey bees. A comparative analysis of GST kinetic activity from imidacloprid-susceptible and -insensitive honey bees revealed a lower bimolecular inhibition rate constant (ki) for imidacloprid-insensitive individuals (5.07 ± 0.098 nmol/min/mg protein) compared to the imidacloprid-sensitive honey bees (17.23 ± 1.235 nmol/min/mg protein). The IC50 of DEM estimated for bees from each colony showed that the imidacloprid-susceptible honey bees possess a higher IC50 (10 μM) than that of the tolerant honey bees (3 μM). These data suggest that the GSTs in the imidacloprid-tolerant honey bees might be a more efficient detoxification mechanism for the conjugation and elimination of imidacloprid, or imidacloprid metabolites, compared to that of imidacloprid-susceptible honey bees. Therefore, I hypothesize that the differences in metabolic detoxification enzyme activities of honey bees collected from different colonies can result in the differential toxicities of honey bees exposed to neonicotinoid insecticides, such as imidacloprid. However, a thorough examination of imidacloprid detoxification in honey bees is warranted to confirm this hypothesis. / Master of Science in Life Sciences / Honey bees are the most important crop pollinator known to humans. The domestication and use of these insects constitutes a multi-billion dollar industry. Their pollination services alone are a necessary part of modern day agriculture. One of the concerns raised today with regard to honey bee health is their exposure to insecticides used widely in modern agriculture to manage crop pests and protect our food supply from devastating crop loss. One insecticide family that has gained much attention lately are the neonicotinoids. These insecticides are reported to elicit sublethal effects to honey bees that can affect colony health. Some of the more widely used neonicotinoids include, but are not limited to, imidacloprid, thiacloprid, and acetamiprid. The goal of this study was to examine the acute toxicity of imidacloprid to honey bees collected from different colonies and to compare the metabolic detoxification enzyme activities of the honey bees to understand the mechanism(s) of imidacloprid sensitivity in the honey bees. Here, I report a 36-fold difference in the acute toxicity of imidacloprid to the honey bees collected from different colonies. A comparison of glutathione <i>S</i>-transferases activities in imidacloprid-susceptible and -tolerant honey bees suggest that these metabolic detoxification enzymes may assist in the conjugation of imidacloprid, or associated metabolites, and thus facilitate the removal of the insecticide from the honey bees.

BACKGROUND: The honey bee is an important model system for increasing understanding of molecular and neural mechanisms underlying social behaviors relevant to the agricultural industry and basic science. The western honey bee, Apis mellifera, has served as a model species, and its genome sequence has been published. In contrast, the genome of the Asian honey bee, Apis cerana, has not yet been sequenced. A. cerana has been raised in Asian countries for thousands of years and has brought considerable economic benefits to the apicultural industry. A cerana has divergent biological traits compared to A. mellifera and it has played a key role in maintaining biodiversity in eastern and southern Asia. Here we report the first whole genome sequence of A. cerana. RESULTS: Using de novo assembly methods, we produced a 238 Mbp draft of the A. cerana genome and generated 10,651 genes. A.cerana-specific genes were analyzed to better understand the novel characteristics of this honey bee species. Seventy-two percent of the A. cerana-specific genes had more than one GO term, and 1,696 enzymes were categorized into 125 pathways. Genes involved in chemoreception and immunity were carefully identified and compared to those from other sequenced insect models. These included 10 gustatory receptors, 119 odorant receptors, 10 ionotropic receptors, and 160 immune-related genes. CONCLUSIONS: This first report of the whole genome sequence of A. cerana provides resources for comparative sociogenomics, especially in the field of social insect communication. These important tools will contribute to a better understanding of the complex behaviors and natural biology of the Asian honey bee and to anticipate its future evolutionary trajectory.

Thesis (MSc)--University of Stellenbosch, 2006. / ENGLISH ABSTRACT: The aim of this study was to investigate the phylogeographic structuring of the honey badger, Mellivora capensis, a highly mobile medium sized carnivore with an extensive distribution throughout sub-Saharan Africa extending into the Middle East and India. Particular focus was placed on providing preliminary data potentially useful for the development of translocation policies for this species in southern Africa. Where possible, genetic results were also compared with current trinomial designations to determine whether subspecies status given to geographical groupings was supported by the genetic data. Mitochondrial control region sequence data was obtained for most a selection of specimen’s available while nuclear microsatellite variation was determined for a subset of individuals where there were sufficient sample sizes available. Phylogeographic structuring of the maternal mitochondrial lineage was initially obscured by the coamplification of a closely related numt. To overcome co-amplification, the numt was identified and mtDNA specific primers were designed. Mitochondrial DNA results are based on the most variable 230 bp of the control region (sequenced for 78 individuals) while five polymorphic nuclear microsatellite markers were scored (for 55 individuals). Analysis, employing both nuclear and mitochondrial data, showed that although a pattern of isolation by distance can be detected, there was evidence for the presence of phylogeographic structuring between eastern and southern Africa. This could be interpreted as due to vicariance, probably associated by rifting and climatic occilations during the Pleistocene. Analyses support the identification of distinct management units for eastern and southern African populations although some evidence exists for secondary introgression between these two regions. Following this, we recommend that translocations between these broad geographic areas should be avoided. Within these geographic areas, because of a general pattern of isolation by distance, we recommend that individuals for translocations come from geographically proximate populations. In some instances, phylogeographic structuring is concordant with subspecies designations but additional sampling will be needed to make any firm taxonomic conclusions. / AFRIKAANSE OPSOMMING: Die doelwit van hierdie studie was om die filogeografiese struktuur van die ratel, Mellivora capensis, ‘n hoogs bewegende medium groot karnivoor met ‘n wydverspreide distribusie deur sub-Sahara Afrika wat strek tot in die Midde Ooste en India, te bepaal. Spesifieke fokus is geplaas op die voorsiening van preliminêre data potensieel bruikbaar vir dir ontwikkeling van verplasing strategieë vir hierdie spesie in suidelike Afrika. Genetiese resultate is, waar moontlik, vergelyk met huidige drieledige kategorieë om te bepaal of subspesies ondersteun word deur die genetiese data. Mitochondriale ‘control region’ DNS volgorde data was verkry vir die meeste van die monsters beskikbaar en kern mikrosatelliet variasie was bepaal vir ‘n gedeelte van individue waar voldoende monster groottes beskikbaar was. Filogeografiese strukturering van die materne mitochondriale merker was oorspronklik versteek deur die ko-amplifikasie van ‘n naby verwante ‘numt’. Om die ko-amplifikasie te oorkom is die ‘numt’ geïdentifiseer en mtDNS spesifieke voorvoerders is ontwerp. Mitochondriale DNS resultate is gebaseer op die mees veranderlike 230 bp van die ‘control region’ (waar die DNS volgorde vir 78 individue bepaal is) en vyf polimorfiese kern mikrosatelliet merkers (in 55 individue). Analises, wat gebruik maak van kern en mitochondriale data, toon wel ‘n patroon van isolasie deur afstand, maar ook ‘n duidelike sigbare filogeograpfiese strukturering tussen oostelike en suidelike Afrika. Hierdie is geïnterpreteer as vikariansie, heel waarskynlik ge-assosieer deur berg verskuiwings en klimaatsveranderinge deur die Pleistocene. Analises ondersteun die identifikasie van definitiewe verkillende bestuurseenhede vir oostelike en suidelike Afrika maar sekere bewyse bestaan dat sekondêre introgressie tussen streke bestaan. Dit word aanbeveel dat translokasies tussen hierdie geografiese areas voorkom moet word. Binne geografiese areas, as gevolg van ‘n algemene patroon van isolasie deur afstand, is dit aanbeveel dat individue vir verplasing van nabygeleë populasies moet wees. In sommige gevalle het filogeografiese strukturering ooreen gestem met subspesies kategorieë, maar verdere materiaal is nodig voor definitiewe taksonomiese besluite geneem kan word.