SummaryFrom studies of behaviour, chemical communication, genomics and developmental biology, among many others, honey bees have long been a key organism for fundamental breakthroughs in biology. With a genome sequence in hand, and much improved genetic tools, honey bees are now an even more appealing target for answering the major questions of evolutionary biology, population structure, and social organization.At the same time, agricultural incentives to understand how honey bees fall prey to disease, or evade and survive their many pests and pathogens, have pushed for a genetic understanding of individual and social immunity in this species. Below we describe and reference tools for using modern molecular-biology techniques to understand bee behaviour, health, and other aspects of their biology. We focus on DNA and RNA techniques, largely because techniques for assessing bee proteins are covered in detail in Hartfelder et al. (2013). We cover practical needs for bee sampling, transport, and storage, and then discuss a range of current techniques for genetic analysis. We then provide a roadmap for genomic resources and methods for studying bees, followed by specific statistical protocols for population genetics, quantitative genetics, and phylogenetics. Finally, we end with three important tools for predicting gene regulation and function in honey bees: Métodos estándar para la investigación molecular en Apis mellifera ResumenLas abejas de miel han sido durante mucho tiempo un organismo clave para avances fundamentales en biología a partir de estudios de su comportamiento, comunicación química, genómica y de biología del desarrollo, entre otros muchos. Con la secuencia del genoma en la mano y herramientas genéticas mucho mejores, las abejas son ahora un blanco aún más atractivo para responder a las preguntas más importantes de la biología evolutiva, la estructura de las poblaciones y la organización social. Al mismo tiempo, los incentivos agrícolas para entender cómo las abejas caen enfermas, o evadir y sobrevivir a sus muchas plagas y patógenos, han presionado para comprender genéticamente la inmunidad individual y social en esta especie. A continuación se describen y se hace referencia a herramientas que hacen uso de modernas técnicas de biología molecular para entender el comportamiento de las abejas, su salud y otros aspectos de su biología. Nos centramos en las técnicas de ADN y ARN, en gran parte debido a que las técnicas de evaluación de las proteínas de la abeja se tratan en detalle en Hartfelder et al. (2013). Cubrimos las necesidades prácticas de toma de muestras de abejas, su transporte y almacenamiento, y luego se discuten una serie de técnicas actuales de análisis genético. A continuación, se proporciona una hoja de ruta para los recursos genómicos y métodos para estudiar las abejas, seguido de protocolos estadísticos específicos de la genética de poblaciones, la genética cuantitativa y la filogenia.Finalmente, se termina con tres herramientas importantes para predecir la regulación génica y la fu...

In eusocial insects the production of daughters is generally restricted to mated queens, and unmated workers are functionally sterile. The evolution of this worker sterility has been plausibly explained by kin selection theory [Hamilton W (1964) J Theor Biol 7:1-52], and many traits have evolved to prevent conflict over reproduction among the females in an insect colony. In honeybees (Apis mellifera), worker reproduction is regulated by the queen, brood pheromones, and worker policing. However, workers of the Cape honeybee, Apis mellifera capensis, can evade this control and establish themselves as social parasites by activating their ovaries, parthenogenetically producing diploid female offspring (thelytoky) and producing queen-like amounts of queen pheromones. All these traits have been shown to be strongly influenced by a single locus on chromosome 13 [Lattorff HMG, et al. (2007) Biol Lett 3:292-295]. We screened this region for candidate genes and found that alternative splicing of a gene homologous to the gemini transcription factor of Drosophila controls worker sterility. Knocking out the critical exon in a series of RNAi experiments resulted in rapid worker ovary activation-one of the traits characteristic of the social parasites. This genetic switch may be controlled by a short intronic splice enhancer motif of nine nucleotides attached to the alternative splice site. The lack of this motif in parasitic Cape honeybee clones suggests that the removal of nine nucleotides from the altruistic worker genome may be sufficient to turn a honeybee from an altruistic worker into a parasite.caste determination | cuticular protein 2-family | gene expression T he evolution of a sterile worker caste in eusocial hymenoptera has been plausibly explained by inclusive fitness theory (1). Generally, workers refrain from reproduction as a result of intracolonial reproductive hierarchies. Ovary activation in honeybee workers (Apis mellifera) is inhibited by the pheromones from the queen and the brood (2). In addition, the multiple mating of the queen and the coexistence of many half-sibling subfamilies in the colony facilitates worker policing, where workers remove eggs laid by other workers (3), leading to <1% of worker-laid offspring (4).These mechanisms often fail, however, to control worker reproduction of the Cape honeybee (Apis mellifera capensis). In this subspecies, laying workers can function as social parasites invading foreign colonies, killing the resident queen and establishing themselves as pseudoqueens (5-7). The proximate mechanisms for this parasitic life history strategy are well understood (8). Parasitic pseudoqueen workers produce a queen-like pheromonal bouquet indicating the presence of a queen to the host workers (9-13). Moreover, the diploid offspring of the parasitizing workers produce brood pheromones, suggesting the presence of a laying queen to the host workers and preventing these workers from activating their ovaries. Finally, the diploid eggs laid by the parasitic workers are not policed as pr...

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-Deformed wing virus (DWV) represents an ideal model to study the interaction between mode of transmission and virulence in honey bees since it exhibits both horizontal and vertical transmissions. However, it is not yet clear if venereal-vertical transmission represents a regular mode of transmission for this virus in natural honey bee populations. Here, we provide clear evidence for the occurrence of high DWV titres in the endophallus of sexually mature drones collected from drone congregation areas (DCAs). Furthermore, the endophallus DWV titres of drones collected at their maternal hives were no different from drones collected at nearby DCAs, suggesting that high-titre DWV infection of the endophallus does not hinder the ability of drones to reach the mating area. The results are discussed within the context of the dispersal of DWV between colonies and the definition of DWV virulence with respect to the transmission route and the types of tissues infected.honey bee / DWV / drone congregation areas / vertical transmission

RNA interference involves the targeted knockdown of mRNA triggered by complementary dsRNA molecules applied to an experimental organism. Although this technique has been successfully used in honeybees (Apis mellifera), it remains unclear whether the application of dsRNA leads to unintended expression knockdown in unspecific, non-targeted genes. Therefore, we studied the gene expression of four non-target genes coding for proteins that are involved in different physiological processes after treatment with three dsRNAs in two abdominal tissues. We found unspecific gene downregulation depending on both the dsRNA used and the different tissues. Hence, RNAi experiments in the honeybee require rigid controls and carefully selected dsRNA sequences to avoid misinterpretation of RNAi-derived phenotypes.