Michelle M. Elekonich

Age and natural metabolically-intensive behavior affect oxidative stress and antioxidant mechanisms

Flying honey bees have among the highest mass-specific metabolic rates ever measured, suggesting that their flight muscles may experience high levels of oxidative stress during normal daily activities. We measured parameters of oxidative stress and antioxidant capacity in highly metabolic flight muscle and less active head tissue in cohorts of age-matched nurse bees, which rarely fly, and foragers, which fly several hours per a day. Naturally occurring foraging flight elicited an increase in flight muscle Hsp70 content in both young and old foragers; however catalase and total antioxidant capacity increased only in young flight muscle. Surprisingly, young nurse bees also showed a modest daily increase in Hsp70, catalase levels and antioxidant capacity, and these effects were likely due to collecting the young nurses soon after orientation flights. There were no differences in flight muscle carbonyl content over the course of daily activity and few differences in Hsp70, catalase, total antioxidant capacity and protein carbonyl levels in head tissue regardless of age or activity. In summary, honey bee flight likely produces high levels of reactive oxygen species in flight muscle that, when coupled with age-related decreases in antioxidant activity may be responsible for behavioral senescence and reduced longevity.

The effects of age and behavioral development on honey bee (Apis mellifera) flight performance

SUMMARY A critical but seldom-studied component of life history theory is how behavior and age affect whole-organism performance. To address this issue we compared the flight performance of honey bees (whose behavioral development and age can be assessed independently via simple manipulations of colony demographics) between distinct behavioral castes (in-hive nurse bees vs out-of-hive foragers) and across lifespan. Variable-density gases and high-speed video were used to determine the maximum hovering flight capacity and wing kinematics of age-matched nurse bees and foragers sampled from a single-cohort colony over a period of 34 days. The transition from hive work to foraging was accompanied by a 42% decrease in body mass and a proportional increase in flight capacity (defined as the minimum gas density allowing hovering flight). The lower flight capacity of hive bees was primarily due to the fact that in air they were functioning at a near-maximal wing angular velocity due to their high body masses. Foragers were lighter and when hovering in air required a much lower wing angular velocity, which they were able to increase by 32% during maximal flight performance. Flight performance of hive bees was independent of age, but in foragers the maximal wingbeat frequency and maximal average angular velocity were lowest in precocious (7–14 day old) foragers, highest in normal-aged (15–28 day old) foragers and intermediate in foragers older than 29 days. This pattern coincides with previously described age-dependent biochemical and metabolic properties of honey bee flight muscle.

Extreme thermotolerance and behavioral induction of 70-kDa heat shock proteins and their encoding genes in honey bees

Foraging honey bees frequently leave the hive to gather pollen and nectar for the colony. This period of their lives is marked by periodic extremes of body temperature, metabolic expenditure, and flight muscle activity. Following ecologically relevant episodes of hyperthermia between 33°C and 50°C, heat shock protein 70 (Hsp70) expression and hsp70/hsc70-4 activity in brains of nonflying laboratory-held bees increased by only two to three times baseline at temperatures 46–50°C. Induction was undetectable in thoracic–flight muscles. Yet, thorax hsp70 mRNA (but not hsc70-4 mRNA) levels were up to ten times higher in flight-capable hive bees and foraging bees compared to 1-day-old, flight-incapable bees, while brain hsp70/hsc70-4 mRNA levels were low and varied little among behavioral groups. These data suggest honey bee tissues, especially flight muscles, are extremely thermotolerant. Furthermore, Hsp70 expression in the thoraces of flight-capable bees is probably flight-induced by oxidative and mechanical damage to flight muscle proteins rather than temperature.

Muscle biochemistry and the ontogeny of flight capacity during behavioral development in the honey bee, Apis mellifera

SUMMARY A fundamental issue in physiology and behavior is understanding the functional and genetic mechanisms that underlie major behavioral shifts in organisms as they adopt new environments or life history tactics. Such transitions are common in nature and include the age-related switch from nest/hive work to foraging in social insects such as honey bees (Apis mellifera). Because of their experimental tractability, recently sequenced genome and well understood biology, honey bees are an ideal model system for integrating molecular, genetic, physiological and sociobiological perspectives to advance understanding of behavioral and life history transitions. When honey bees (Apis mellifera) transition from hive work to foraging, their flight muscles undergo changes that allow these insects to attain the highest rates of flight muscle metabolism and power output ever recorded in the animal kingdom. Here, we review research to date showing that honey bee flight muscles undergo significant changes in biochemistry and gene expression and that these changes accompany a significant increase in the capacity to generate metabolic and aerodynamic power during flight. It is likely that changes in muscle gene expression, biochemistry, metabolism and functional capacity may be driven primarily by behavior as opposed to age, as is the case for changes in honey bee brains.

Requirements for In Vitro Germination of Paenibacillus larvae Spores

Paenibacillus larvae is the causative agent of American foulbrood (AFB), a disease affecting honey bee larvae. First- and second-instar larvae become infected when they ingest food contaminated with P. larvae spores. The spores then germinate into vegetative cells that proliferate in the midgut of the honey bee. Although AFB affects honey bees only in the larval stage, P. larvae spores can be distributed throughout the hive. Because spore germination is critical for AFB establishment, we analyzed the requirements for P. larvae spore germination in vitro. We found that P. larvae spores germinated only in response to l-tyrosine plus uric acid under physiologic pH and temperature conditions. This suggests that the simultaneous presence of these signals is necessary for spore germination in vivo. Furthermore, the germination profiles of environmentally derived spores were identical to those of spores from a biochemically typed strain. Because l-tyrosine and uric acid are the only required germinants in vitro, we screened amino acid and purine analogs for their ability to act as antagonists of P. larvae spore germination. Indole and phenol, the side chains of tyrosine and tryptophan, strongly inhibited P. larvae spore germination. Methylation of the N-1 (but not the C-3) position of indole eliminated its ability to inhibit germination. Identification of the activators and inhibitors of P. larvae spore germination provides a basis for developing new tools to control AFB.

Effects of Flight on Gene Expression and Aging in the Honey Bee Brain and Flight Muscle

Honey bees move through a series of in-hive tasks (e.g., “nursing”) to outside tasks (e.g., “foraging”) that are coincident with physiological changes and higher levels of metabolic activity. Social context can cause worker bees to speed up or slow down this process, and foragers may revert back to their earlier in-hive tasks accompanied by reversion to earlier physiological states. To investigate the effects of flight, behavioral state and age on gene expression, we used whole-genome microarrays and real-time PCR. Brain tissue and flight muscle exhibited different patterns of expression during behavioral transitions, with expression patterns in the brain reflecting both age and behavior, and expression patterns in flight muscle being primarily determined by age. Our data suggest that the transition from behaviors requiring little to no flight (nursing) to those requiring prolonged flight bouts (foraging), rather than the amount of previous flight per se, has a major effect on gene expression. Following behavioral reversion there was a partial reversion in gene expression but some aspects of forager expression patterns, such as those for genes involved in immune function, remained. Combined with our real-time PCR data, these data suggest an epigenetic control and energy balance role in honey bee functional senescence.

Comparison of in vitro methods for the production of Paenibacillus larvae endospores

Paenibacillus larvae endospores are the infectious particles of the honey bee brood disease, American Foulbrood. We demonstrate that our previously published protocol (Alvarado et al., 2013) consistently yields higher numbers and purer preparations of P. larvae endospores, than previously described protocols, regardless of the strain tested (B-3650, B-3554 or B-3685).

Biomedical Research with Honey Bees

In addition to the economic benefits honey bees provide through pollination and honey production, this species is an important model system that provides insight into many areas of biomedical science. With a long history as a subject of studies of social behavior, learning and memory, and immunology (due to the allergens in their venom), the honey bee is emerging as a major model for phenotypic plasticity, development and aging, circadian rhythms, muscle metabolism and behavioral genomics. Sequencing of the honey bee genome revealed that honey bees share many genes and biochemical pathways with humans, thus opening new avenues of research. Biomedical research with honey bees benefits from this species’ well known physiology, the tractability of its natural behaviors both in and out of the laboratory, a fully sequenced genome with accompanying suite of cellular and molecular tools, and a large interactive community of basic and applied researchers. Only Drosophila rivals the honey bee as an insect model system for biomedical research.