The switch from within-hive activities to foraging behavior is a major transition in the life cycle of a honeybee (Apis mellifera) worker. A prominent regulatory role in this switch has long been attributed to juvenile hormone (JH), but recent evidence also points to the yolk precursor protein vitellogenin as a major player in behavioral development. In the present study, we injected vitellogenin double-stranded RNA (dsVg) into newly emerged worker bees of Africanized genetic origin and introduced them together with controls into observation hives to record flight behavior. RNA interference-mediated silencing of vitellogenin gene function shifted the onset of long-duration flights (>10 min) to earlier in life (by 3-4 days) when compared with sham and untreated control bees. In fact, dsVg bees were observed conducting such flights extremely precociously, when only 3 days old. Short-duration flights (<10 min), which bees usually perform for orientation and cleaning, were not affected. Additionally, we found that the JH titer in dsVg bees collected after 7 days was not significantly different from the controls. The finding that depletion of the vitellogenin titer can drive young bees to become extremely precocious foragers could imply that vitellogenin is the primary switch signal. At this young age, downregulation of vitellogenin gene activity apparently had little effect on the JH titer. As this unexpected finding stands in contrast with previous results on the vitellogenin/JH interaction at a later age, when bees normally become foragers, we propose a three-step sequence in the constellation of physiological parameters underlying behavioral development.

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SummaryDespite their tremendous economic importance, and apart from certain topics in the field of neurophysiology such as vision, olfaction, learning and memory, honey bees are not a typical model system for studying general questions of insect physiology. The reason is their social lifestyle, which sets them apart from a "typical insect" and, during social evolution, has resulted in the restructuring of certain physiological pathways and biochemical characteristics in this insect. Not surprisingly, the questions that have attracted most attention by researchers working on honey bee physiology and biochemistry in general are core topics specifically related to social organization, such as caste development, reproductive division of labour and polyethism within the worker caste. With certain proteins playing key roles in these processes, such as the major royal jelly proteins (MRJPs), including royalactin and hexamerins in caste development, and vitellogenin in reproductive division of labour and age polyethism, a major section herein will present and discuss basic laboratory protocols for protein analyses established and standardized to address such questions in bees. A second major topic concerns endocrine mechanisms underlying processes of queen and worker development, as well as reproduction and polyethism, especially the roles of juvenile hormone and ecdysteroids. Sensitive techniques for the quantification of juvenile hormone levels circulating in haemolymph, as well as its synthesis by the corpora allata are described. Although these require certain instrumentation and a considerable degree of sophistication in the analysis procedures, we considered that presenting these techniques would be of interest to laboratories planning to specialize in such analyses. Since biogenic amines are both neurotransmitters and regulators of endocrine glands, we also present a standard method for the detection and analysis of certain biogenic amines of interest. Further questions that cross borders between individual and social physiology are related to energy metabolism and thermoregulation. Thus a further three sections are dedicated to protocols on carbohydrate quantification in body fluid, body temperature measurement and respirometry. Métodos estándar para la investigación de la fisiología y bioquímica de Apis mellifera ResumenA pesar de su enorme importancia económica, y aparte de ciertos temas en el campo de la neurofisiología, tales como la visión, el olfato, el aprendizaje y la memoria, las abejas no son un sistema modelo típico para el estudio de cuestiones generales sobre la fisiología de los insectos. La razón de ello es su forma de vida social, lo que las diferencia de un "insecto típico" y que durante la evolución social, se ha traducido en la reestructuración de ciertas vías fisiológicas y bioquímicas propias de este insecto. Como era de esperar, las preguntas que han atraído mayor atención por parte de los investigadores que trabajan en la fisiología y la bioquímica de la abeja melífera, son en general tema...

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ABSTRACT. Pollen substitute diets are a valuable resource for maintaining strong and health honey bee colonies. Specific diets may be useful in one region or country and inadequate or economically unviable in others. We compared two artificial protein diets that had been formulated from locally-available ingredients in Brazil with bee bread and a nonprotein sucrose diet. Groups of 100 newly-emerged, adult workers of Africanized honey bees in Brazil and European honey bees in the USA were confined in small cages and fed on one of four diets for seven days. The artificial diets included a high protein diet made of soy milk powder and albumin, and a lower protein level diet consisting of soy milk powder, brewer's yeast and rice bran. The initial protein levels in newly emerged bees were approximately 18-21 µg/µL hemolymph. After feeding on the diets for seven days, the protein levels in the hemolymph were similar among the protein diet groups (~37-49 µg/µL after seven days), although Africanized bees acquired higher protein levels, increasing 145 and 100% on diets D1 and D2, respectively, versus 83 and 60% in the European bees. All the protein diets resulted in significantly higher levels of protein than sucrose solution alone. In the field, the two pollen substitute diets were tested during periods of low pollen availability in the field in two regions of Brazil. Food consumption, population development, colony weight, and honey production were evaluated to determine the impact of the diets on colony strength parameters. The colonies fed artificial diets had a significant improvement in all parameters, while control colonies dwindled during the dearth period. We conclude that these two artificial protein diets have good potential as pollen substitutes during dearth periods and that Africanized bees more efficiently utilize artificial protein diets than do European honey bees.

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