Y. Le Conte

Differential gene expression of the honey bee Apis mellifera associated with Varroa destructor infection

BackgroundThe parasitic mite, Varroa destructor, is the most serious pest of the western honey bee, Apis mellifera, and has caused the death of millions of colonies worldwide. This mite reproduces in brood cells and parasitizes immature and adult bees. We investigated whether Varroa infestation induces changes in Apis mellifera gene expression, and whether there are genotypic differences that affect gene expression relevant to the bees tolerance, as first steps toward unravelling mechanisms of host response and differences in susceptibility to Varroa parasitism.ResultsWe explored the transcriptional response to mite parasitism in two genetic stocks of A. mellifera which differ in susceptibility to Varroa, comparing parasitized and non-parasitized full-sister pupae from both stocks. Bee expression profiles were analyzed using microarrays derived from honey bee ESTs whose annotation has recently been enhanced by results from the honey bee genome sequence. We measured differences in gene expression in two colonies of Varroa-susceptible and two colonies of Varroa-tolerant bees. We identified a set of 148 genes with significantly different patterns of expression: 32 varied with the presence of Varroa, 116 varied with bee genotype, and 2 with both. Varroa parasitism caused changes in the expression of genes related to embryonic development, cell metabolism and immunity. Bees tolerant to Varroa were mainly characterized by differences in the expression of genes regulating neuronal development, neuronal sensitivity and olfaction. Differences in olfaction and sensitivity to stimuli are two parameters that could, at least in part, account for bee tolerance to Varroa; differences in olfaction may be related to increased grooming and hygienic behavior, important behaviors known to be involved in Varroa tolerance.ConclusionThese results suggest that differences in behavior, rather than in the immune system, underlie Varroa tolerance in honey bees, and give an indication of the specific physiological changes found in parasitized bees. They provide a first step toward better understanding molecular pathways involved in this important host-parasite relationship.

Primer effects of a brood pheromone on honeybee behavioural development

Primer pheromones are thought to act in a variety of vertebrates and invertebrates but only a few have been chemically identified. We report that a blend of ten fatty–acid esters found on the cuticles of honeybee larvae, already known as a kairomone, releaser pheromone and primer pheromone, also act as a primer pheromone in the regulation of division of labour among adult workers. Bees in colonies receiving brood pheromone initiated foraging at significantly older ages than did bees in control colonies in five out of five trials. Laboratory and additional field tests also showed that exposure to brood pheromone significantly depressed blood titres of juvenile hormone. Brood pheromone exerted more consistent effects on age at first foraging than on juvenile hormone, suggesting that the primer effects of this pheromone may occur via other, unknown, mechanisms besides juvenile hormone. These results bring the number of social factors known to influence honeybee division of labour to three: worker–worker interactions, queen mandibular pheromone and brood pheromone.

Regulation of brain gene expression in honey bees by brood pheromone

Pheromones are very important in animal communication. To learn more about the molecular basis of pheromone action, we studied the effects of a potent honey bee pheromone on brain gene expression. Brood pheromone (BP) caused changes in the expression of hundreds of genes in the bee brain in a manner consistent with its known effects on behavioral maturation. Brood pheromone exposure in young bees causes a delay in the transition from working in the hive to foraging, and we found that BP treatment tended to upregulate genes in the brain that are upregulated in bees specialized on brood care but downregulate genes that are upregulated in foragers. However, the effects of BP were age dependent; this pattern was reversed when older bees were tested, consistent with the stimulation of foraging by BP in older bees already competent to forage. These results support the idea that one way that pheromones influence behavior is by orchestrating large‐scale changes in brain gene expression. We also found evidence for a relationship between cis and BP regulation of brain gene expression, with several cis‐regulatory motifs statistically overrepresented in the promoter regions of genes regulated by BP. Transcription factors that target a few of these motifs have already been implicated in the regulation of bee behavior. Together these results demonstrate strong connections between pheromone effects, behavior, and regulation of brain gene expression.

Effect of Aliphatic Esters on Ovary Development of Queenless Bees (Apis mellifera L.)

In honeybee queenright colonies, ovary development in workers is inhibited by queen-produced pheromones [1–4] and by the presence of the brood [5]. It seems that the presence of the unsealed brood provides an inhibitory signal stronger than the queen’s pheromone [6]. When administered orally with honey, the ethyl alcohol or acetone extracts prepared from unsealed honey bee brood show inhibitory effects on worker ovary development [7]. More recent results show that a mixture of ten fatty acid esters (methyl and ethyl esters of palmitic, linoleic, linolenic, stearic, and oleic acids) present naturally on the larval cuticle [8] induce a strong inhibitory effect on ovary development of caged bees in controlled conditions. This inhibition occurs by contact, diffusion, or ingestion of the blend [9]. Based on this preliminary work, the aim of this paper was to determine which esters are involved in the inhibition of ovary development of caged bees and to test whether these compounds maintain their effect under natural conditions. First we screened the efficiency of different esters. For this purpose each ester was mixed individually in food (bee candy: mixture of honey and powdered sugar) at a concentration of 10 (wt/wt) and was tested under controlled conditions on newly emerged bees (Apis mellifera mellifera). Each group of 120 bees was placed in a 12!10!4 cm cage, provided with a vial of tap water, pollen,

Social immunity in honeybees (Apis mellifera): transcriptome analysis of varroa‐hygienic behaviour

Honeybees have evolved a social immunity consisting of the cooperation of individuals to decrease disease in the hive. We identified a set of genes involved in this social immunity by analysing the brain transcriptome of highly varroa‐hygienic bees, who efficiently detect and remove brood infected with the Varroa destructor mite. The function of these candidate genes does not seem to support a higher olfactory sensitivity in hygienic bees, as previously hypothesized. However, comparing their genomic profile with those from other behaviours suggests a link with brood care and the highly varroa‐hygienic Africanized honeybees. These results represent a first step toward the identification of genes involved in social immunity and thus provide first insights into the evolution of social immunity.

Modifications of the cuticular hydrocarbon profile of Apis mellifera worker bees in the presence of the ectoparasitic mite Varroa jacobsoni in brood cells.

Varroa jacobsoni is an ectoparasite of Apis mellifera which invades brood cells, on 8-day-old larvae several hours before cell capping. Reproduction of the parasite takes place in the capped brood cells during the nymphose of the bee. Cuticular hydrocarbons of unparasitized bees and of bees parasitized by Varroa jacobsoni were extracted and analysed by gas chromatography (GC) coupled with mass spectrometry (GC-MS). Three developmental stages of worker honey bees were studied: larvae, pupae and emergent adults. The comparison between unparasitized and parasitized hosts was performed with Principal Components Analysis coupled with a multivariate variance analysis. The cuticular hydrocarbon profiles of honey bees were qualitatively similar, for the 3 developmental stages and regardless of the presence of Varroa in the cells. Nevertheless, comparison of the relative proportions of hydrocarbons showed that the cuticular profiles of pupae and emergent adults parasitized by 1 mite and of larvae parasitized by 2 mites were significantly different from the corresponding unparasitized individuals. Such modifications could be regarded (i) as a cause of the multi-infestation in larvae during invasion of brood and (ii) as a consequence of stress and/or removal of proteins contained in the haemolymph of the host during its development.

Microsatellite analysis of sperm admixture in honeybee.

Sperm usage was investigated in an instrumentally inseminated honeybee queen. Her progeny were examined in the first 3 months of the egg‐laying period using a microsatellite marker. Frequencies of different subfamilies differed significantly from one month to another. However, there was no evidence for sperm displacement or sperm precedence of a specific male in the worker progeny. The variance of subfamily proportions decreased over time suggesting that sperm admixture in the spermatheca was incomplete at the beginning of the egg‐laying period of the queen and improved progressively during the first months after mating.

Worker-worker inhibition of honey bee behavioural development independent of queen and brood

Summary.Foragers inhibit the behavioural development of young adult worker honey bees, delaying the age at onset of foraging. But the similar effect caused by pheromones produced by both the queen and brood raised the possibility that some of the previously attributed forager effects might be due to queen, brood, or both. Here we studied whether physical contacts between young bees and old foragers can inhibit behavioural development while controlling for queen and brood effects. Results demonstrated that foragers inhibit the behavioural development of young adult worker bees independent of the queen and brood, via a mechanism that requires physical contact.

Unity in defence: honeybee workers exhibit conserved molecular responses to diverse pathogens

BackgroundOrganisms typically face infection by diverse pathogens, and hosts are thought to have developed specific responses to each type of pathogen they encounter. The advent of transcriptomics now makes it possible to test this hypothesis and compare host gene expression responses to multiple pathogens at a genome-wide scale. Here, we performed a meta-analysis of multiple published and new transcriptomes using a newly developed bioinformatics approach that filters genes based on their expression profile across datasets. Thereby, we identified common and unique molecular responses of a model host species, the honey bee (Apis mellifera), to its major pathogens and parasites: the Microsporidia Nosema apis and Nosema ceranae, RNA viruses, and the ectoparasitic mite Varroa destructor, which transmits viruses.ResultsWe identified a common suite of genes and conserved molecular pathways that respond to all investigated pathogens, a result that suggests a commonality in response mechanisms to diverse pathogens. We found that genes differentially expressed after infection exhibit a higher evolutionary rate than non-differentially expressed genes. Using our new bioinformatics approach, we unveiled additional pathogen-specific responses of honey bees; we found that apoptosis appeared to be an important response following microsporidian infection, while genes from the immune signalling pathways, Toll and Imd, were differentially expressed after Varroa/virus infection. Finally, we applied our bioinformatics approach and generated a gene co-expression network to identify highly connected (hub) genes that may represent important mediators and regulators of anti-pathogen responses.ConclusionsOur meta-analysis generated a comprehensive overview of the host metabolic and other biological processes that mediate interactions between insects and their pathogens. We identified key host genes and pathways that respond to phylogenetically diverse pathogens, representing an important source for future functional studies as well as offering new routes to identify or generate pathogen resilient honey bee stocks. The statistical and bioinformatics approaches that were developed for this study are broadly applicable to synthesize information across transcriptomic datasets. These approaches will likely have utility in addressing a variety of biological questions.

Propolis chemical composition and honeybee resistance against Varroa destructor

Propolis is known as honeybee chemical defence against infections and parasites. Its chemical composition is variable and depends on the specificity of the local flora. However, there are no data concerning the relationship between propolis chemical composition and honeybee colony health. We tried to answer this question, studying the chemical composition of propolis of bee colonies from an apiary near Avignon, which are tolerant to Varroa destructor, comparing it with colonies from the same apiary which are non-tolerant to the mites. The results indicated that non-tolerant colonies collected more resin than the tolerant ones. The percentage of four biologically active compounds – caffeic acid and pentenyl caffeates – was higher in propolis from tolerant colonies. The results of this study pave the way to understanding the effect of propolis in individual and social immunity of the honeybees. Further studies are needed to clarify the relationship between propolis chemical composition and honeybee colony health.