Does the diapause experience of bumblebee queens Bombus terrestris affect colony characteristics?

1. Bumblebee colonies show much variation in the number of workers, drones, and queens produced. Because this variation prevails even when colonies are kept under identical conditions, it does not seem to be caused by extrinsic factors but rather by differences between founding queens. 2. The most likely factor that could cause differences between queens is diapause. Although colonies are raised under standardised conditions, the queens often experience diapause of different length. If there are costs associated with diapause that influence post‐diapause reproduction, the diapause history of the queens could affect colony characteristics. 3. Here, several colony characteristics are compared: number of first and second brood workers; total number of workers, drones, and queens; energy spent on sexuals; sex ratio; rate of worker production; time to emergence of first reproductive; and colony lifetime. Colonies were used where the queens experienced a diapause treatment of 0 (nondiapause queens), 2, and 4 months. 4. Although no proof was found for the existence of costs associated with diapause, the colony characteristics of nondiapause queens were significantly different from those of diapause queens. Colonies of nondiapause queens produced the lowest number of workers but the highest number of young queens. 5. It is argued that these nondiapause colonies are more time‐constrained than diapause colonies because nondiapause colonies produce two generations within the same season and should therefore be more efficient in producing sexual offspring. 6. Moreover, nondiapause colonies should rear a more female‐biased sex ratio because they can be certain of the presence of males produced by other (diapause) colonies.     

Sex ratio variation in the bumblebee Bombus terrestris

Patterns of sex allocation in bumblebees have been enigmaticand difficult to interpret in either a Fisherian context orin a kin-selection perspective. We gathered data on severalhundred laboratory-reared colonies of the bumblebee Bombus terrestrisand analyzed sex allocation as a function of diapause durationand a series of variables describing colony development. Ouranalyses addressed both sex allocation patterns across differentcohorts of laboratory colonies reared at different times andsex allocation patterns across individual colonies within thesecohorts. We used path analysis to test a hypothetical modellinking a sequence of colony-development variables to the crucialreproductive parameters at the end of the colony life cycle.We show that (1) population-wide patterns of sex allocationshow equal investment in the sexes and are thus consistent withqueen control, but not with worker control. (2) A significantpart of the colony-level and cohort-specific variation in sexallocation is related to the hibernation conditions of foundingqueens: Queens with longer than average winter diapause producelarger cohorts of first and second brood workers, switch tohaploid eggs early, and produce colonies that raise mostly malesand few new queens and vice versa. (3) Colony-level sex allocationis significantly related to the time span between the switchpoint (date of first haploid egg laid by the queen) and thecompetition point (date of first haploid egg laid by one ofthe workers): the longer this period, the more male biased thesex ratio. (4) The breeding constraints of an annual life cycle,the short reproductive season, and the presumably high premiumon early produced males imply that bumblebee workers have norealistic options to capitalize on their relatedness asymmetrytoward the different kinds of reproductive brood by biasingthe sex ratio.     

Impact of artificial photoperiodism on the colony development of the bumblebee Bombus terrestris

This study investigated the effect of the photoperiodic regimes 0 h light : 24 h dark (LD 0:24), LD 8:16, LD 16:8 and LD 24:0 at 28°C and 50% Relative Humidity (RH) on the colony development of hibernated (2.0, 2.5, 3.0, 3.5 and 4.0 months) bumblebee queens. The queens which had hibernated for 3.0 months and which were reared in a LD 8:16 photoperiod showed the highest rate of colony initiation (88.2%), colony foundation (67.6%) and progeny queen production (38.2%). The photoperiod LD 8:16 also produced the shortest period of colony initiation and colony foundation. The highest number of sexual males (171.2 ± 12.2) and queens (91.2 ± 9.9) were produced in the colonies when 3.5 and 3.0 month hibernated queens were kept in an LD 8:16 photoperiod. The results show that light regime and hibernation duration affect colony characteristics of Bombus terrestris.     

Photoperiodic influence on the body mass of bumblebee, Bombus terrestris and its copulation duration

Abstract:  The body mass of Bombus terrestris individuals is an important trait for their behavioural performance and colony organization. In this study, colonies were reared under four different photoperiodic regimes, viz. 0 : 24, 8 : 16, 16 : 8 and 24 : 0 h light : darkness (L : D) at 28°C and 50% relative humidity. The changes in body mass were observed at the stages of larvae, pupae and on the day of adult eclosion. Both the wet and dry mass of sexuals gradually decreased with increasing day length. The relationship between body mass and copulation duration revealed that copulation duration was negatively correlated with male body mass, but positively with queen body mass. Higher number of matings by males resulted in significantly higher duration of copulation.     

Behavioural interactions, kin and disease susceptibility in the bumblebee Bombus terrestris

Behavioural interactions are often analysed in terms of their costs and benefits to the actors [Hamilton, (1964) J. Theor. Biol.7 1-16; Gadagkar, (1993) Trends Ecol. Evol.8 232-234; Foster et al., (2001) Ann. Zool. Fenn.38 229-238]. Using the bumblebee Bombus terrestris, we wish to distinguish between two possible determinants of interaction behaviour between conspecifics, namely kin-directed behaviour that reflects genetic distance between individuals, or, alternatively, interactions guided by a functional distance between individuals, specifically, with respect to disease susceptibility. We find no relationship between contact rate of individuals and the genetic distance of their respective colonies. Interestingly, we do find a significant negative correlation between contact rate and the distance between the two colonies in susceptibility to a spectrum of parasite strains. This cannot be explained by either of the a priori alternatives so we propose two further testable hypotheses to explain our results.     

Age- and task-dependent foraging gene expression in the bumblebee Bombus terrestris

In eusocial insects, the division of labor within a colony, based on either age or size, is correlated with a differential foraging (for) gene expression and PKG activity. This article presents in the first part a study on the for gene, encoding a cGMP-dependent protein kinase (PKG) in the bumblebee Bombus terrestris. Cloning of the open reading frame allowed phylogenetic tracing, which showed conservation of PKGs among social insects. Our results confirm the proposed role for PKGs in division of labor. Btfor gene expression is significantly higher in the larger foragers compared with the smaller sized nurses. More importantly, we discovered an age-related decrease in Btfor expression in both nursing and foraging bumblebees. We therefore speculate that the presence of BtFOR is required for correct adaptation to new external stimuli and rapid learning for foraging. In a second series of experiments, worker bumblebees of B. terrestris were treated with two insecticides imidacloprid and kinoprene, which have shown to cause impaired foraging behavior. Compared with controls, only the latter treatment resulted in a decreased Btfor expression, which concurs with a stimulation of ovarian growth and a shift in labor toward nest-related tasks. The data are discussed in relation to Btfor expression in the complex physiological event of foraging and side-effects by pesticides.     

Induction of colony initiation by Japanese native bumble bees using cocoons of the exotic bumblebee Bombus terrestris

The colony initiation rates of Bombus hypocrita (a native Japanese bumblebee) and Bombus terrestris (a European species) foundresses were compared after 4 weeks of exposure to B. terrestris cocoons. The B. terrestris cocoons, when replaced weekly, were effective for inducing oviposition by foundresses of both species. There were no significant differences in the colony initiation rates of B. terrestris and B. hypocrita, either with the control treatment or with the cocoons. The cocoon method was also tested for five species and two subspecies of native Japanese bumblebees. The colony initiation rate was higher for foundresses of the subgenus Bombus s. str. than for foundresses of the subgenera Pyrobombus, Diversobombus, and Thoracobombus. When replaced weekly, the cocoons of B. terrestris are effective inducers of colony foundation in three Japanese native species, namely B. ignitus, B. hypocrita hypocrita, and B. h. sapporoensis.     

Morphological features,development and reproduction of Melittobia acasta on Bombus terrestris

Morphological features, development and reproduction behavior of the parasite Melittobia acasta (Walker) were studied when reared on the pupae of the bumblebee Bombus terrestris L. in the laboratory under 23°C, 50% relative humidity and 12 h light : 12 h dark conditions. The parasites laid transparent white and elongated eggs. Newly hatched larval size and shape were very similar to eggs but they were identified by their body segments. Larvae increased their body size through moulting and transformed into a vermiform shape. Male pupae were shiny brown with dots. The female pupae were distinguished by their black shiny color, shorter size and the presence of compound eyes. Adult male pupae were dark brown and dwarf‐winged, whereas female pupae were macropterous and brachypterous. Reproduction took place by fertilization and also parthenogenetically. Mean fecundity within 5 days by mated (47.9 ± 30.5 female^?1) and virgin (7.4 ± 6.8 female^?1) females were statistically different. Mated females laid fertilized eggs that produced adult males or females, whereas virgin females laid unfertilized eggs that produced males. Development durations of the virgin female originated eggs, larvae, pupae and adults were statistically identical with those of mated females. The parasites were female‐biased and foundress number did not affect offspring sex ratio. This study shows that both mated and virgin females of M. acasta can produce many offspring on B. terrestris pupae within a short period, indicating that they are dangerous parasites of the bumblebee in a mass rearing system.     

Heritability of sperm length in the bumblebee Bombus terrestris

Sperm length is highly variable, both between and within species, but the evolutionary significance of this variation is poorly understood. Sexual selection on sperm length requires a significant additive genetic variance, but few studies have actually measured this. Here we present the first estimates of narrow sense heritability of sperm length in a social insect, the bumblebee Bombus terrestris. In spite of a balanced and straightforward rearing design of colonies, and the possibility to replicate measurements of sperm within single males nested within colonies, the analysis proved to be complex. Several appropriate statistical models were derived, each depending on different assumptions. The heritability estimates obtained ranged from h ² = 0.197 ± 0.091 to h ² = 0.429 ± 0.154. All our estimates were substantially lower than previous estimates of sperm length heritability in non-social insects and vertebrates.     

Effects of photoperiod and hibernation duration on the lifespan of Bombus terrestris

In this study the effects of photoperiod and hibernation duration on the lifespan of Bombus terrestris queens were examined. Hibernation durations of 2.0, 2.5, 3.0, 3.5 and 4.0 months were studied, as were photoperiods of 0 h light : 24 h dark (LD 0:24), LD 8:16, LD 16:8 and LD 24:0. The queens that hibernated for 2.5 months and were exposed to 1 week of LD 8:16 had the highest survival rate (89.3%); the lowest survival rate was found in queens that hibernated for 4.0 months and were reared at LD 24:0. Photoperiod and hibernation duration had significant effects on egg predation by founding queens, competition between queens and workers, and emergence of sexual queens. Hibernation durations of 2.5 and 3.0 months and a photoperiod of LD 8:16 resulted in a significantly longer lifespan of B. terrestris.     

Population structure and inbreeding in a rare and declining bumblebee, Bombus muscorum (Hymenoptera: Apidae)

Owing to habitat loss populations of many organisms have declined and become fragmented. Vertebrate conservation strategies routinely consider genetic factors, but their importance in invertebrate populations is poorly understood. Bumblebees are important pollinators, and many species have undergone dramatic declines. As monoandrous social hymenopterans they may be particularly susceptible to inbreeding due to low effective population sizes. We study fragmented populations of a bumblebee species, on a model island system, and on mainland Great Britain where it is rare and declining. We use microsatellites to study: population genetic structuring and gene flow; the relationships between genetic diversity, population size and isolation; and frequencies of (sterile) diploid males - an indicator of inbreeding. We find significant genetic structuring (theta = 0.12) and isolation by distance. Populations > 10 km apart are all significantly differentiated, both on oceanic islands and on the mainland. Genetic diversity is reduced relative to closely related common species, and isolated populations exhibit further reductions. Of 16 populations, 10 show recent bottlenecking, and 3 show diploid male production. These results suggest that surviving populations of this rare insect suffer from inbreeding as a result of geographical isolation. Implications for the conservation of social hymenopterans are discussed.     

Expressed sequence tag analysis of the diapausing queen of the bumblebee Bombus ignitus

We constructed a full‐length cDNA library from diapausing queens of the bumblebee Bombus ignitus. A total of 480 randomly selected clones was sequenced by single‐run 5′‐end sequencing. Of these, there were 437 high quality clones, 23 poor quality clones and 20 read‐fail clones. Each high quality clone sequence was searched against a public protein database. The most frequently found matching genes were ribosomal proteins (12.5%), p10 (3.58%), cytochrome P450 monooxygenase (3.13%) and sensory appendage protein (2.9%). Sequence similarity analysis between bumblebees and other insect species showed that 72 out of 437 (16.5%) bumblebee expressed sequence tags (EST) matched sequences of Apis mellifera, with matches to Drosophila melanogaster (6.6%), Caenorhabditis briggsae (6.2%), Lysiphlebus testaceipes (4.8%), Periplaneta americana (3.7%) and Anopheles gambiae (3.4%) following, suggesting that sequence similarity of bumblebee EST is closest to that of A. mellifera. Functional classification of EST based on Gene Ontology showed that most genes found by sequencing are associated with physiological processes in the bumblebee. The results of sequencing and analysis of our 437 cDNA demonstrated that high‐throughput EST sequencing and data analysis are powerful means for identifying novel genes and for expression profiling. Our bumblebee EST collection could be a useful platform for further studies of gene expression in diapausing bumblebees.     

Perception,regulation, and fitness effects of pollen phytosterols in the bumble bee,Bombus terrestris

Premise

Many flowering plants depend on insects for pollination and thus attract pollinators by offering rewards, mostly nectar and pollen. Bee pollinators rely on pollen as their main nutrient source. Pollen provides all essential micro- and macronutrients including substances that cannot be synthesized by bees themselves, such as sterols, which bees need for processes such as hormone production. Variations in sterol concentrations may consequently affect bee health and reproductive fitness. We therefore hypothesized that (1) these variations in pollen sterols affect longevity and reproduction in bumble bees and (2) can thus be perceived via the bees' antennae before consumption.

Methods

We studied the effect of sterols on longevity and reproduction of Bombus terrestris workers in feeding experiments and investigated sterol perception using chemotactile proboscis extension response (PER) conditioning.

Results

Workers could perceive several sterols (cholesterol, cholestenone, desmosterol, stigmasterol, β-sitosterol) via their antennae but not differentiate between them. However, when sterols were presented in pollen, and not as a single compound, the bees were unable to differentiate between pollen differing in sterol content. Additionally, different sterol concentrations in pollen neither affected pollen consumption nor brood development or worker longevity.

Conclusions

Since we used both natural concentrations and concentrations higher than those found in pollen, our results indicate that bumble bees may not need to pay specific attention to pollen sterol content beyond a specific threshold. Naturally encountered concentrations might fully support their sterol requirements and higher concentrations do not seem to have negative effects.

     

Influence of inbreeding in the early stages of artificially reared colonies of Bombus terrestris

In the present study on the bumblebee Bombus terrestris, we investigated the influence of inbreeding on queen fitness by comparing diapause survival and egg-laying success of queens mated with nestmate and non-nestmate males. We then compared the early stage of colonies with or without diploid males and analysed colony characteristics to identify a factor predictive of colony outcome. Diapause survival was no different between queens mated with nestmates and non-nestmates, but in the latter case, egg-laying success was significantly higher. Queens mated with nestmates gave rise to a percentage of diploid male colonies (52.6%) compatible with brother–sister coupling. We obtained 18.6% of colonies with diploid males even from queens mated with non-nestmates, indicating that the colonies of origin were in some way related or homozygous at the sex determination loci. There was no difference in the early growth stage between colonies with or without diploid males, except in the number of workers emerging in the first brood, which was significantly higher in the latter. Among diploid male colonies, the number of workers and the male/worker ratio in the first brood was highly variable and was not a good predictor of subsequent colony growth. Out of 49 colonies with diploid males that reached full development, only 11 had a sufficient size to assume that they could survive in the field or, in a commercial breeding, to be suitable for pollination purposes.     

Intraspecific Variation of the Cephalic Labial Gland Secretions in Bombus terrestris (L.) (Hymenoptera: Apidae)

Variations of secretions of the cephalic part of the labial glands from four different subspecies of Bombus terrestris, B. t. terrestris, B. t. lusitanicus, B. t. sassaricus, and B. t. dalmatinus, were investigated. 95 compounds were detected in the whole data set: 54 in B. t. terrestris, 54 in B. t. lusitanicus, 48 in B. t. sassaricus, and 44 in B. t. dalmatinus. The (E)‐2,3‐dihydrofarnesol is the main compound in B. t. dalmatinus and B. t. sassaricus, while it is dihydrofarnesyl dodecanoate in B. t. terrestris and B. t. lusitanicus. A principal component analysis produced a pattern showing three well distinct groups corresponding to dalmatinus, sassaricus, and terrestris+lusitanicus.     

Effects of social conditions on Juvenile Hormone mediated reproductive development in Bombus terrestris workers

Abstract. During the annual life cycle of the bumble bee Bombus terrestris (L.) colony, there is a stage characterized by worker reproduction in the presence of the queen. It has been proposed that this is a result of a decrease in queen inhibition. This hypothesis was examined by studying the effects of queens taken from colonies at different stages of development on several aspects of worker physiology and behaviour: rates of Juvenile Hormone (JH) release in vitro , ovary development, and behaviour associated with reproduction. After optimizing and validating the radiochemical assay for JH release for bumble bee workers, we found that queenless workers had significantly more developed ovaries and higher rates of release of JH than did queenright workers, confirming and extending previous findings that suggest that bumblebee ovarian development is under JH control. Mated queens, separated from their colony and brood, can have the same inhibitory effect on the reproductive development of callow workers. In contrast, workers confined with virgin queens or in queenless groups demonstrated a significantly higher rate of release of JH, overt aggression and threatening behaviours. However, there were no differences in rates of release of JH between workers confined in groups in the laboratory with queens taken from colonies either before or after the onset of worker reproduction. Furthermore, overt aggression and threatening behaviours were similar and low in both types of groups. These results gave no support to the hypothesis that a decrease in queen inhibition is associated with the onset of worker reproduction. We also show that young workers reared in colonies either before or after worker reproduction occurs, or in queenless colonies, all demonstrated similar, low rates of release of JH. These results suggest that older workers may inhibit the corpora allata of younger workers in queenless colonies.     

New method to stimulate the onset of Bombus terrestris (Hymenoptera: Apidae) rearing: Using worker helpers in the presence of frozen pupae

The failure of bumblebee queens to enthusiastically start a colony under laboratory conditions may be due to lack of oviposition during an experiment, a long delay in oviposition from the termination of hibernation, and failure to rear a large first brood. In the present study, the use of frozen male pupae to start the colony with the assistance of bumblebee workers rather than fresh, young, male pupae was investigated under controlled room temperature and humidity conditions. The period of initiation of the colony decreased with an increase in the number of worker helpers from one to six. The period was as short as 3.9 days in the presence of six worker helpers. Second and third broods also started earlier with the help of workers. The rate of first worker production per egg cup was double that of the normal method (i.e. without worker helpers). Egg eating behavior of the queen was not found in the first brood if more than two workers helped the development of the larvae. Four worker helpers were found to be sufficient, as they could produce 100% colony foundation and 91.46% first worker production colonies. This study showed that using frozen old pupae is a good stimulator for colony foundation in the presence of four bumblebee worker helpers.     

Respiration in bumblebee queens: effect of life phase on the discontinuous ventilation cycle

Evidence for discontinuous ventilation cycles (DVC) has been obtained for many insects under various experimental conditions. Here, we show that DVC's exist for queens of the bumblebee Bombus terrestris in different phases of their life cycle: before entering diapause (pre-diapause queens) and when a colony has been established (post-diapause queens). Pre- and post-diapause queens experience distinct environmental conditions (ambient O₂ and CO₂ concentrations) which is reflected in their DVC: both ventilation frequency and amount of carbon dioxide emitted change with differences in ambient CO₂ concentration.