Osman Kaftanoglu

The Making of a Queen: TOR Pathway Is a Key Player in Diphenic Caste Development

Background Honey bees (Apis mellifera) provide a principal example of diphenic development. Excess feeding of female larvae results in queens (large reproductives). Moderate diet yields workers (small helpers). The signaling pathway that links provisioning to female developmental fate is not understood, yet we reasoned that it could include TOR (target of rapamycin), a nutrient- and energy-sensing kinase that controls organismal growth. Methodology/Principal Findings Here, the role of Apis mellifera TOR (amTOR) in caste determination is examined by rapamycin/FK506 pharmacology and RNA interference (RNAi) gene knockdown. We show that in queen-destined larvae, the TOR inhibitor rapamycin induces the development of worker characters that are blocked by the antagonist FK506. Further, queen fate is associated with elevated activity of the Apis mellifera TOR encoding gene, amTOR, and amTOR gene knockdown blocks queen fate and results in individuals with worker morphology. Conclusions/Significance A much-studied insect dimorphism, thereby, can be governed by the TOR pathway. Our results present the first evidence for a role of TOR in diphenic development, and suggest that adoption of this ancestral nutrient-sensing cascade is one evolutionary pathway for morphological caste differentiation in social insects.

Larval and nurse worker control of developmental plasticity and the evolution of honey bee queen–worker dimorphism

Social evolution in honey bees has produced strong queen–worker dimorphism for plastic traits that depend on larval nutrition. The honey bee developmental programme includes both larval components that determine plastic growth responses to larval nutrition and nurse components that regulate larval nutrition. We studied how these two components contribute to variation in worker and queen body size and ovary size for two pairs of honey bee lineages that show similar differences in worker body–ovary size allometry but have diverged over different evolutionary timescales. Our results indicate that the lineages have diverged for both nurse and larval developmental components, that rapid changes in worker body–ovary size allometry may disrupt queen development and that queen–worker dimorphism arises mainly from discrete nurse‐provided nutritional environments, not from a developmental switch that converts variable nutritional environments into discrete phenotypes. Both larval and nurse components have likely contributed to the evolution of queen–worker dimorphism.

The Genetic Basis of Transgressive Ovary Size in Honeybee Workers

Ovarioles are the functional unit of the female insect reproductive organs and the number of ovarioles per ovary strongly influences egg-laying rate and fecundity. Social evolution in the honeybee (Apis mellifera) has resulted in queens with 200–360 total ovarioles and workers with usually 20 or less. In addition, variation in ovariole number among workers relates to worker sensory tuning, foraging behavior, and the ability to lay unfertilized male-destined eggs. To study the genetic architecture of worker ovariole number, we performed a series of crosses between Africanized and European bees that differ in worker ovariole number. Unexpectedly, these crosses produced transgressive worker phenotypes with extreme ovariole numbers that were sensitive to the social environment. We used a new selective pooled DNA interval mapping approach with two Africanized backcrosses to identify quantitative trait loci (QTL) underlying the transgressive ovary phenotype. We identified one QTL on chromosome 11 and found some evidence for another QTL on chromosome 2. Both QTL regions contain plausible functional candidate genes. The ovariole number of foragers was correlated with the sugar concentration of collected nectar, supporting previous studies showing a link between worker physiology and foraging behavior. We discuss how the phenotype of extreme worker ovariole numbers and the underlying genetic factors we identified could be linked to the development of queen traits.

PDK1 and HR46 Gene Homologs Tie Social Behavior to Ovary Signals.

The genetic basis of division of labor in social insects is a central question in evolutionary and behavioral biology. The honey bee is a model for studying evolutionary behavioral genetics because of its well characterized age-correlated division of labor. After an initial period of within-nest tasks, 2–3 week-old worker bees begin foraging outside the nest. Individuals often specialize by biasing their foraging efforts toward collecting pollen or nectar. Efforts to explain the origins of foraging specialization suggest that division of labor between nectar and pollen foraging specialists is influenced by genes with effects on reproductive physiology. Quantitative trait loci (QTL) mapping of foraging behavior also reveals candidate genes for reproductive traits. Here, we address the linkage of reproductive anatomy to behavior, using backcross QTL analysis, behavioral and anatomical phenotyping, candidate gene expression studies, and backcross confirmation of gene-to-anatomical trait associations. Our data show for the first time that the activity of two positional candidate genes for behavior, PDK1 and HR46, have direct genetic relationships to ovary size, a central reproductive trait that correlates with the nectar and pollen foraging bias of workers. These findings implicate two genes that were not known previously to influence complex social behavior. Also, they outline how selection may have acted on gene networks that affect reproductive resource allocation and behavior to facilitate the evolution of social foraging in honey bees.

Natural product propolis: chemical composition

The chemical composition of propolis from East Mediterranean (Hatay, Adana and Mersin) was studied in order to determine the major compounds by using GC-MS. In this study, the ethanolic extract of propolis prepared by mixing 1900 mL 70% ethanol and 100 g propolis was used. Chemical analysis of propolis extracts indicated that the propolis samples had high concentrations of the aromatic acids, esters and other derivatives which are responsible for the anti-bacterial, anti-fungal, anti-viral, anti-inflammatory and anti-cancer properties of propolis such as benzyl cinnamate, methyl cinnamate, caffeic acid, cinnamyl cinnamate and cinnamoylglcine besides the most common compounds as fatty acid, terpenoids, esters, alcohols hydrocarbons and aromatic acids.

Preservation of Honeybee Spermatozoa in Liquid Nitrogen

SummaryHoneybee (Apis mellifera) semen was stored for 359 days at −196°C in liquid nitrogen. Eleven semen diluents were tested. Use of a mixture of 40% semen and 60% Kiev diluent containing 10% dimethylsulphoxide, combined with freezing of samples at the rate of 3–4°/min produced relatively good post-thaw sperm motility. Fifteen queens were inseminated instrumentally with semen stored for 305–359 days. Thirteen survived and produced a brood area of 345 ± 60 cm2, of which 47·3 ± 6·42% were worker progeny from the frozen spermatozoa. Four queens were superseded by their daughters which mated naturally and built up strong colonies.

Surgically increased ovarian mass in the honey bee confirms link between reproductive physiology and worker behavior.

Honey bee (Apis mellifera L.) workers are essentially sterile females that are used to study how complex social behavior develops. Workers perform nest tasks, like nursing larvae, prior to field tasks, like foraging. Despite worker sterility, this behavioral progression correlates with ovary size: workers with larger ovaries (many ovary filaments) start foraging at younger ages on average. It is untested, however, whether the correlation confers a causal relationship between ovary size and behavioral development. Here, we successfully grafted supernumerary ovaries into worker bees to produce an artificial increase in the amount of ovary tissue. We next measured fat body mRNA levels for the yolk precursor gene vitellogenin, which influences honey bee behavioral development and can correlate with ovary size. Vitellogenin was equally expressed in surgical controls and bees with supernumerary ovaries, leading us to predict that these groups would be characterized by equal behavior. Contrary to our prediction, bees with supernumerary ovaries showed accelerated behavioral development compared to surgical controls, which behaved like reference bees that were not treated surgically. To explore this result we monitored fat body expression levels of a putative ecdysteroid-response gene, HR46, which is genetically linked to ovary size in workers. Our data establish that social insect worker behavior can be directly influenced by ovaries, and that HR46 expression changes with ovary size independent of vitellogenin.

Support for the reproductive ground plan hypothesis of social evolution and major QTL for ovary traits of Africanized worker honey bees (Apis mellifera L.)

BackgroundThe reproductive ground plan hypothesis of social evolution suggests that reproductive controls of a solitary ancestor have been co-opted during social evolution, facilitating the division of labor among social insect workers. Despite substantial empirical support, the generality of this hypothesis is not universally accepted. Thus, we investigated the prediction of particular genes with pleiotropic effects on ovarian traits and social behavior in worker honey bees as a stringent test of the reproductive ground plan hypothesis. We complemented these tests with a comprehensive genome scan for additional quantitative trait loci (QTL) to gain a better understanding of the genetic architecture of the ovary size of honey bee workers, a morphological trait that is significant for understanding social insect caste evolution and general insect biology.ResultsBack-crossing hybrid European x Africanized honey bee queens to the Africanized parent colony generated two study populations with extraordinarily large worker ovaries. Despite the transgressive ovary phenotypes, several previously mapped QTL for social foraging behavior demonstrated ovary size effects, confirming the prediction of pleiotropic genetic effects on reproductive traits and social behavior. One major QTL for ovary size was detected in each backcross, along with several smaller effects and two QTL for ovary asymmetry. One of the main ovary size QTL coincided with a major QTL for ovary activation, explaining 3/4 of the phenotypic variance, although no simple positive correlation between ovary size and activation was observed.ConclusionsOur results provide strong support for the reproductive ground plan hypothesis of evolution in study populations that are independent of the genetic stocks that originally led to the formulation of this hypothesis. As predicted, worker ovary size is genetically linked to multiple correlated traits of the complex division of labor in worker honey bees, known as the pollen hoarding syndrome. The genetic architecture of worker ovary size presumably consists of a combination of trait-specific loci and general regulators that affect the whole behavioral syndrome and may even play a role in caste determination. Several promising candidate genes in the QTL intervals await further study to clarify their potential role in social insect evolution and the regulation of insect fertility in general.

A Washing Technique for Collection of Honeybee Semen

SummaryA simple and fast method for collecting honeybee semen in large quantity is described. Semen and mucus of many drones, everted and ejaculated by hand, were scraped into Kiev diluent in a semen-washing funnel and a collecting tube. The semen-mucus-diluent mixture was centrifuged at 2500 rpm for 10 min to separate semen from mucus and the diluent. The collecting tube containing semen was incorporated into a special large-capacity syringe, and the semen was used to inseminate queen honeybees.No significant differences were found in the amount of brood produced between 31 queens instrumentally inseminated with semen collected by this method, queens inseminated with semen collected by the conventional Mackensen technique, and open-mated queens (P > 0·05 in all cases). However, significant differences were observed in the onset of oviposition (P < 0·05 in all cases). There was a delay before oviposition by all the instrumentally inseminated queens, which was greatest for queens inseminated with semen col...

Rearing Honey Bees, Apis mellifera, in vitro 1: Effects of Sugar Concentrations on Survival and Development

Abstract A new method for rearing honey bees, Apis mellifera L. (Hymenoptera: Apidae), in vitro was developed and the effects of sugar concentrations on survival and development were studied. Seven different glucose (G) and fructose (F) compositions (0%G+0%F, 3%G+3%F, 6%G+6%F, 12%G+12%F, 0%G+12%F, 12%G+0%F, and 4%G+8%F) were tested. Larvae were able to grow to the post defecation stage without addition of sugars (Diet 1), but they were not able to metamorphose and pupate. Adults were reared from diets 2–7. The average larval survival, prepupal larval weights, adult weights, and ovariole numbers were affected significantly due to the sugar compositions in the diets. High sugar concentrations (12%G+12%F) increased the number of queens and intercastes.