This dissertation examines some important physiological and pheromonal factors regulating foraging and colony growth in honey bee colonies. The first study analyzed effects of soybean trypsin inhibitor (SBTI) on the development of hypopharyngeal gland, midgut enzyme activity and survival of the honey bee. In this study newly emerged caged bees were fed pollen diets containing three different concentrations of SBTI. Bees fed 1% SBTI had significantly reduced hypopharyngeal gland protein content. This study indicated that nurse bees fed a pollen diet containing at least 1% SBTI would be poor producers of larval food. In the second study nurse bee biosynthesis of brood food was manipulated using SBTI, and the resulting effects on pollen foraging were measured. Experimental colonies were given equal amounts of SBTI treated and untreated pollen. SBTI treatments had significantly lower hypopharyngeal gland protein content than controls. There was no significant difference in the ratio of pollen to non-pollen foragers and pollen load weights collected between the treatments. These results supported the pollen foraging effort predictions generated from the direct independent effects hypothesis. In the third study we tested whether brood pheromone (BP) regulated queen egg laying via modulation of worker-queen interactions and nurse bee rearing behaviors. This experiment had BP and control treatments. Queens in the BP treatment laid greater number of eggs, were fed for a greater amount of time and were less idle. Significantly more time was spent in cell cleaning by the bees in BP treatments. The results suggest that brood pheromone regulated queen egg-laying rate by modulating worker-queen interactions and nurse bee rearing behavior. The final study of this dissertation focused on how dose-dependent BP-mediated division of labor affected the partitioning of non-foraging and foraging work forces and the amount of brood reared. Triple cohort colonies were used and there were three treatments, Low BP, High BP and Control. Low BP treatments had significantly higher ratio of pollen to non-pollen foragers and greater pollen load weights. Low BP treatment bees foraged at a significantly younger age. This study has shown that BP elicits dose-dependent modulation of foraging and brood rearing behaviors.

This dissertation examines some important physiological and pheromonal factors regulating foraging and colony growth in honey bee colonies. The first study analyzed effects of soybean trypsin inhibitor (SBTI) on the development of hypopharyngeal gland, midgut enzyme activity and survival of the honey bee. In this study newly emerged caged bees were fed pollen diets containing three different concentrations of SBTI. Bees fed 1% SBTI had significantly reduced hypopharyngeal gland protein content. This study indicated that nurse bees fed a pollen diet containing at least 1% SBTI would be poor producers of larval food. In the second study nurse bee biosynthesis of brood food was manipulated using SBTI, and the resulting effects on pollen foraging were measured. Experimental colonies were given equal amounts of SBTI treated and untreated pollen. SBTI treatments had significantly lower hypopharyngeal gland protein content than controls. There was no significant difference in the ratio of pollen to non-pollen foragers and pollen load weights collected between the treatments. These results supported the pollen foraging effort predictions generated from the direct independent effects hypothesis. In the third study we tested whether brood pheromone (BP) regulated queen egg laying via modulation of worker-queen interactions and nurse bee rearing behaviors. This experiment had BP and control treatments. Queens in the BP treatment laid greater number of eggs, were fed for a greater amount of time and were less idle. Significantly more time was spent in cell cleaning by the bees in BP treatments. The results suggest that brood pheromone regulated queen egg-laying rate by modulating worker-queen interactions and nurse bee rearing behavior. The final study of this dissertation focused on how dose-dependent BP-mediated division of labor affected the partitioning of non-foraging and foraging work forces and the amount of brood reared. Triple cohort colonies were used and there were three treatments, Low BP, High BP and Control. Low BP treatments had significantly higher ratio of pollen to non-pollen foragers and greater pollen load weights. Low BP treatment bees foraged at a significantly younger age. This study has shown that BP elicits dose-dependent modulation of foraging and brood rearing behaviors.

Brood pheromone of the honey bee, (Apis mellifera) has been shown to elicit a wide array of primer and releaser effects on non‐foragers and foragers leading to the regulation of nursing, pollen foraging, and behavioral development such that the behavior of the colony may be regulated by the amount and condition of the larvae. To date, all studies into the effects of brood pheromone have either used uncharacterized whole extracts or a single blend of brood pheromone characterized from a population of honey bees in France. The variation in the relative proportions of the ten fatty‐acid ester components that characterize brood pheromone and some effects of this variation on pollen foraging and sucrose response thresholds were therefore observed. The objectives met in this dissertation were to determine whether changes in brood pheromone component proportions (blend) or amount communicates larval nutritional status and reports the results of observations of nurses and foragers in response to blends of brood pheromone from deprived and‐non deprived larvae, to measure how changes in brood pheromone blend changed pollen foraging behavior and if such changes could account for the pollen foraging differences between Africanized and European bees, and finally to observe the effects of exposure time on brood pheromone blend and to observe whether non‐foragers made contact with the pheromone. Brood pheromone was found to vary by larval rearing environment, but did not elicit the expected behaviors that would support a cue of nutritional status. Brood pheromone also varied significantly by mitochondrial lineage/population source and responses to brood pheromone appeared to be coadapted to blend, suggesting that brood pheromone may be important in race recognition. Finally, brood pheromone varied significantly over time and was found to be removed from sources by bees, suggesting possible mechanisms for loss of effect. Combined the results of this research indicate that brood pheromone blend differences lead to profound changes in colony behavior related to pollen foraging and food provisioning, providing novel tools for colony manipulation and mechanisms for understanding brood rearing division of labor and chemical communication.

Honey bees (Apis mellifera) are the most economically important insect pollinator of agricultural crops in the United States. Honey bee colonies are required for pollination of approximately one-third of the nation’s fruit, vegetable, nut, and forage crops, with an estimated annual value in the billions of dollars. The economic value of a honey bee colony comes from its population size, as large colonies provide the necessary foraging force required for large-scale crop pollination services. A major component of colony strength is its genetic diversity, a consequence of the reproductive mating strategy of the queen known as polyandry. Despite some inherent risks of multiple mating, several studies have demonstrated significant advantages of intracolonial genetic diversity for honey bee colony productivity. Colony-level benefits include better disease resistance, more stable brood nest thermoregulation, and greater colony growth. Instrumental insemination of honey bee queens is a technique to precisely control queen mating, and thereby creates the opportunity to investigate the effects of intracolonial genetic diversity on colony performance. In this dissertation, I first consider the effects of intracolonial genetic diversity on pollen foraging using colonies headed by queens which were instrumentally inseminated with either one or twenty drones to generate colonies of very high or very low intracolonial genetic diversity, respectively. I found that colonies with high intracolonial genetic diversity amass significantly more pollen and rear more brood than colonies with low intracolonial genetic diversity. Of particular interest, colonies with low intracolonial genetic diversity collected a significantly greater variety of pollen types. I discuss these results in the context of scouting and recruiting, and suggest a more efficient foraging strategy exists among genetically diverse colonies. While intracolonial genetic diversity is positively correlated with collected pollen, its effect on the colony’s ability to process and distribute inbound protein resources is unknown. Again using colonies headed by queens instrumentally inseminated with either one or twenty drones, I studied the effects of intracolonial genetic diversity on pollen consumption and digestion by nurse bees, as well as protein allocation among nestmates by assessing total soluble protein concentration of late instar larvae, and total soluble hemolymph protein concentration in both nurses and pollen foragers. I found that nurse bees from colonies with high intracolonial genetic diversity consume and process more protein than nurses from colonies with low intracolonial genetic diversity, even when given equal access to protein resources. Further, both forager hemolymph protein concentrations and larval total protein concentrations were higher among the colonies with high intracolonial genetic diversity. My findings suggest that protein processing and distribution within a honey bee colony is affected by the social context of the hive. I discuss “worker policing”, and the role of nurse bees in modulating the foraging effort. Finally, I assess the standing genetic variability among several colonies sourced from different genetic and geographic locations. Using microsatellite DNA from workers sampled from each colony, I determined allelic richness, gene diversity, and effective mating frequency for each genetic line. I found differences in all three metrics between lines, and for one line in particular, there was no correlation with genetic variation and effective mating frequency, suggesting non-random mating. My results showed very different levels of intracolonial genetic diversity among naturally mated queens. Because of its impact on colony performance, the importance of maintaining genetic diversity in breeding populations is discussed.

A fundamental question in biology is how animals efficiently locate and use diverse resources. Pollinators foraging on flowers are one of our most thoroughly studied examples of generalist foraging behavior and cognition. Individual pollinators typically specialize on a subset of flowering species available to them. Specialization by nectar-foraging pollinators is often the consequence of learned or innate preferences for floral display traits such as color, pattern, and scent. Pollinators must also typically learn to extract nectar from each floral type. By specializing, pollinators reduce costs associated with learning and forgetting nectar extraction routines. Specialization also benefits the plant by enhancing conspecific pollen transfer. Yet nectar is not the only floral reward. The pollen of hundreds of thousands of plant species is collected by pollinators such as bees, beetles, and flies. In fact, solitary and social bees must collect both pollen and nectar to survive. However, much of the vast literature on bee foraging behavior concerns the collection of nectar. This research investigated mechanisms by which generalist bumblebees (Bombus impatiens) specialize on diverse floral resources. Most foragers in a colony were reward generalists over their lifetime, but specialized daily on either pollen or nectar collection. Lifetime patterns of pollen collection were associated with interindividual differences in sensory morphology. Pollen-foraging bumblebees had weak innate preferences, but learned strong preferences for pollen-only plant species, with preferences mediated primarily by anther properties. The anthers provided indirect cues of concealed pollen, and bees learned to prefer properties of the anthers to select potentially rewarding flowers. While learning was involved in the formation of floral preferences by pollen foragers, pollen extraction behavior relied little on learning. Specifically, floral sonication, which is used by bees to extract concealed pollen, was modified only modestly with experience. Furthermore, bees foraged efficiently for pollen from diverse floral resources without relying on instrumental (associative) learning. Efficient foraging involved switching between two distinct motor routines: floral sonication and scrabbling. Switching was regulated by two ubiquitous floral cues: chemical anther cues eliciting sonication and mechanical pollen cues suppressing it (and eliciting scrabbling). I discuss how mechanisms of floral specialization by generalist pollen-foraging bees could drive floral trait evolution.

abstract: How a colony regulates the division of labor to forage for nutritional resources while accommodating for changes in colony demography is a fundamental question in the sociobiology of social insects. In honey bee, Apis mellifera, brood composition impacts the division of labor, but it is unknown if colonies adjust the allocation of foragers to carbohydrate and protein resources based on changes in the age demography of larvae and the pheromones they produce. Young and old larvae produce pheromones that differ in composition and volatility. In turn, nurses differentially provision larvae, feeding developing young worker larvae a surplus diet that is more queen-like in protein composition and food availability, while old larvae receive a diet that mimics the sugar composition of the queen larval diet but is restrictively fed instead of provided ad lib. This research investigated how larval age and the larval pheromone e-β ocimene (eβ) impact foraging activity and foraging load. Additional cage studies were conducted to determine if eβ interacts synergistically with queen mandibular pheromone (QMP) to suppress ovary activation and prime worker physiology for nursing behavior. Lastly, the priming effects of larval age and eβ on worker physiology and the transition from in-hive nursing tasks to outside foraging were examined. Results indicate that workers differentially respond to larvae of different ages, likely by detecting changes in the composition of the pheromones they emit. This resulted in adjustments to the foraging division of labor (pollen vs. nectar) to ensure that the nutritional needs of the colony's brood were met. For younger larvae and eβ, this resulted in a bias favoring pollen collection. The cage studies reveal that both eβ and QMP suppressed ovary activation, but the larval pheromone was more effective. Maturing in an environment of young or old larvae primed bees for nursing and impacted important endocrine titers involved in the transition to foraging, so bees maturing in the presence of larvae foraged earlier than control bees reared with no brood. / Dissertation/Thesis / Ph.D. Biology 2014

Os estudos que visam avaliar a influência dos fatores bióticos (disponibilidade de recursos florais e competição) e abióticos (variáveis climáticas) nas atividades externas (comportamento de forrageamento de pólen) e internas (taxa de construção de células de cria) das colônias de abelhas sem ferrão são importantes para compreender as adaptações comportamentais das espécies. A Caatinga, região semiárida do nordeste brasileiro, é caracterizada por temperaturas anuais elevadas e longos períodos de seca. Nesse estudo, nós investigamos respostas comportamentais de Melipona subnitida Ducke (Apidae, Meliponini) às condições ambientais desse bioma. Os experimentos foram realizados no município de Mossoró/RN, Brasil, esta região foi considerada como uma das 26 áreas de extrema prioridade para a conservação do bioma Caatinga. Na área de estudo nós registramos fontes importantes de pólen e néctar para as abelhas, incluindo espécies fundamentais para a manutenção das colônias durante a estação seca. As abelhas forrageiras de M. subnitida coletaram em um período curto do dia e do ano grandes quantidades de recursos disponíveis, concentrando a força forrageira nas fontes mais lucrativas. Visto que a quantidade de pólen estocado dentro do ninho (oferta) controla a produção de cria, a observada redução da taxa de construção de células de cria durante a estação quente e seca sugere um mecanismo importante para economizar recursos polínicos estocados e consequentemente permiti a sobrevivência das colônias durante períodos prolongados de escassez. Através da modelagem de distribuição de espécies, verificamos um potencial déficit de recursos forrageiros para espécie M. subnitida nas próximas décadas. Diante dessas observações foram discutidas as principais estratégias para a conservação dessa espécie de abelha sem ferrão na região semiárida do nordeste brasileiro. / For a profound understanding of behavioural adaptations of stingless bee species to certain environmental conditions, studies are necessary that aim at evaluating the influence of biotic (availability of floral resources and competition) and abiotic factors (climatic variables) on colony-external activities (pollen foraging) as well as colony-internal activities (brood cell-construction). We investigated the behavioural responses of Melipona subnitida Ducke (Apidae, Meliponini) to the environmental conditions of the Caatinga, a biome in the Brazilian North-East characterized by elevated ambient temperatures and extended dry periods. The experiments were performed in the surroundings of Mossoró/RN, Brazil, which is considered one of the 26 priority areas concerning the conservation of the Caatinga. In the study area, we found important pollen and nectar sources for bees, including tree species of fundamental importance for colony maintenance during the dry season. The foragers of M. subnitida collected large quantities of pollen during a short period during the day and a short period in the course of the year, concentrating their foraging efforts predominantly at highly rewarding resources. Given that the offer of pollen within the nest directly influences brood production, the observed reduction of brood cell-construction during the hot and dry season suggests an important mechanism for economizing the stored pollen, which guarantees the survival of the colony during extended periods of dearth. Modeling future distribution of the species, we identified a potential deficit of food resources for M. subnitida in the next decades. Using the results of our study, we discussed the main strategies for the conservation of this stingless bee species in the semiarid region of the Brazilian Northeast.

Os estudos que visam avaliar a influência dos fatores bióticos (disponibilidade de recursos florais e competição) e abióticos (variáveis climáticas) nas atividades externas (comportamento de forrageamento de pólen) e internas (taxa de construção de células de cria) das colônias de abelhas sem ferrão são importantes para compreender as adaptações comportamentais das espécies. A Caatinga, região semiárida do nordeste brasileiro, é caracterizada por temperaturas anuais elevadas e longos períodos de seca. Nesse estudo, nós investigamos respostas comportamentais de Melipona subnitida Ducke (Apidae, Meliponini) às condições ambientais desse bioma. Os experimentos foram realizados no município de Mossoró/RN, Brasil, esta região foi considerada como uma das 26 áreas de extrema prioridade para a conservação do bioma Caatinga. Na área de estudo nós registramos fontes importantes de pólen e néctar para as abelhas, incluindo espécies fundamentais para a manutenção das colônias durante a estação seca. As abelhas forrageiras de M. subnitida coletaram em um período curto do dia e do ano grandes quantidades de recursos disponíveis, concentrando a força forrageira nas fontes mais lucrativas. Visto que a quantidade de pólen estocado dentro do ninho (oferta) controla a produção de cria, a observada redução da taxa de construção de células de cria durante a estação quente e seca sugere um mecanismo importante para economizar recursos polínicos estocados e consequentemente permiti a sobrevivência das colônias durante períodos prolongados de escassez. Através da modelagem de distribuição de espécies, verificamos um potencial déficit de recursos forrageiros para espécie M. subnitida nas próximas décadas. Diante dessas observações foram discutidas as principais estratégias para a conservação dessa espécie de abelha sem ferrão na região semiárida do nordeste brasileiro. / For a profound understanding of behavioural adaptations of stingless bee species to certain environmental conditions, studies are necessary that aim at evaluating the influence of biotic (availability of floral resources and competition) and abiotic factors (climatic variables) on colony-external activities (pollen foraging) as well as colony-internal activities (brood cell-construction). We investigated the behavioural responses of Melipona subnitida Ducke (Apidae, Meliponini) to the environmental conditions of the Caatinga, a biome in the Brazilian North-East characterized by elevated ambient temperatures and extended dry periods. The experiments were performed in the surroundings of Mossoró/RN, Brazil, which is considered one of the 26 priority areas concerning the conservation of the Caatinga. In the study area, we found important pollen and nectar sources for bees, including tree species of fundamental importance for colony maintenance during the dry season. The foragers of M. subnitida collected large quantities of pollen during a short period during the day and a short period in the course of the year, concentrating their foraging efforts predominantly at highly rewarding resources. Given that the offer of pollen within the nest directly influences brood production, the observed reduction of brood cell-construction during the hot and dry season suggests an important mechanism for economizing the stored pollen, which guarantees the survival of the colony during extended periods of dearth. Modeling future distribution of the species, we identified a potential deficit of food resources for M. subnitida in the next decades. Using the results of our study, we discussed the main strategies for the conservation of this stingless bee species in the semiarid region of the Brazilian Northeast.