Mating frequency and genetic relatedness of workers in the hornet Vespa analis (Hymenoptera: Vespidae)

Mating frequency of Vespa analis queens and the genetic relatedness of their workers was analyzed by DNA microsatellite genotyping. Of 20 colonies studied, 18 had a queen inseminated by a single male and two had queens each inseminated by two males. The estimated effective number of matings was 1.05 ± 0.037 (mean ± SE), with 75–85% of the offspring of the two multiply mated queens sired by a single male. The pedigree relatedness between nestmate workers averaged over the 20 colonies was estimated to be 0.74 ± 0.008, almost identical to the predicted value of 0.75 for colonies headed by a singly mated queen. Multiple matrilines; that is, the presence of workers not related to the current queens, were detected in six colonies, suggesting that queen replacement occurred via usurpation of the founding queens in these six colonies. These results demonstrate that the kin structure of V. analis is similar to that reported in other vespid species.     

Kin structure and colony male reproduction in the hornet<Emphasis Type="Italic"> Vespa crabro</Emphasis> (Hymenoptera: Vespidae)

We estimated queen mating frequency, genetic relatedness among workers, and worker reproduction in Vespa crabro flavofasciata using microsatellite DNA markers. Of 20 colonies examined, 15 contained queens inseminated by a single male, 3 colonies contained queens inseminated by two males, and 2 colonies contained queens inseminated by three males. The genetic relatedness among workers was estimated to be 0.73±0.003 (mean±SE). For this high relatedness, kin selection theory predicts a potential conflict between queens and workers over male production. To verify whether males are derived from queens or workers, 260 males from 13 colonies were genotyped at four microsatellite loci. We found that all of the males were derived from the queens. This finding was further supported by the fact that only 33 of 2,990 workers dissected had developed ovaries. These workers belonged to 2 of the 20 colonies. There was no relationship between queen mating frequency and worker reproduction, and no workers produced male offspring in any of the colonies. These results suggest that male production dominated by queens in V. crabro flavofasciata is possibly due to worker policing.     

Do hornets have zombie workers?

Colonies of the European hornet, Vespa crabro, are typically founded by a single queen mated to a single male. From the resulting colony relatedness pattern we predicted strong worker-queen conflict over male production where both the workers and the queen attempt to produce the colony's males. To test for this conflict, male production was studied in 15 hornet nests using a combination of DNA microsatellite analysis (282 males), worker ovary dissections (500 workers from eight nests) and 50 h of observation (four nests). In contrast to our prediction, the data show that hornet males are queens' sons, that workers never attempt to lay eggs, rarely have activated ovaries, and that there is no direct aggression between the queen and the workers. This contrasts with other data for vespine wasps, which support relatedness predictions. Dolichovespula arenaria has the same kin structure as V. crabro and workers produce males in many colonies. The similarity between these two species makes it difficult to explain why workers do not reproduce in V. crabro. Self-restraint is expected if worker reproduction significantly reduces colony productivity but there is no obvious reason why this should be important to V. crabro but not to D. arenaria. Alternatively, queen control may be important. The absence of expressed queen-worker conflict rules out physical control. Indirect pheromonal control is a possibility and is supported by the occurrence of royal courts and queen pheromone in Vespa but not Dolichovespula. Pheromonal queen control is considered evolutionarily unstable, but could result from a queen-worker arms race over reproductive control in which the queen is ahead. The genetic data also revealed diploid males in one colony, the first example in the vespine wasps, and two colonies with double matrilines, suggesting that occasional usurpation by spring queens occurs.     

REPRODUCTIVE CONFLICT IN BUMBLEBEES AND THE EVOLUTION OF WORKER POLICING

Worker policing (mutual repression of reproduction) in the eusocial Hymenoptera represents a leading example of how coercion can facilitate cooperation. The occurrence of worker policing in “primitively” eusocial species with low mating frequencies, which lack relatedness differences conducive to policing, suggests that separate factors may underlie the origin and maintenance of worker policing. We tested this hypothesis by investigating conflict over male parentage in the primitively eusocial, monandrous bumblebee, Bombus terrestris. Using observations, experiments, and microsatellite genotyping, we found that: (a) worker‐ but not queen‐laid male eggs are nearly all eaten (by queens, reproductive, and nonreproductive workers) soon after being laid, so accounting for low observed frequencies of larval and adult worker‐produced males; (b) queen‐ and worker‐laid male eggs have equal viabilities; (c) workers discriminate between queen‐ and worker‐laid eggs using cues on eggs and egg cells that almost certainly originate from queens. The cooccurrence in B. terrestris of these three key elements of “classical” worker policing as found in the highly eusocial, polyandrous honeybees provides novel support for the hypothesis that worker policing can originate in the absence of relatedness differences maintaining it. Worker policing in B. terrestris almost certainly arose via reproductive competition among workers, that is, as “selfish” policing.     

No evidence that reproductive bumblebee workers reduce the production of new queens

Kin selection theory predicts potential conflict between queen and workers over male parentage in hymenopteran societies headed by one, singly mated queen, because each party is more closely related to its own male offspring. In ‘late-switching’ colonies of the bumblebee Bombus terrestris, i.e. colonies whose queens lay haploid eggs relatively late in the colony cycle, workers start to lay male eggs shortly after the queen lays the female eggs that will develop into new queens. It has been hypothesized that this occurs because workers recognize, via a signal given by the queen instructing female larvae to commence development as queens, that egg laying is now in their kin-selected interest. This hypothesis assumes that aggressive behaviour in egg-laying workers does not substantially reduce the production of new queens, which would decrease the workers' fitness payoff from producing males. We tested the hypothesis that reproductive activity inB. terrestris workers does not reduce the production of new queens. We used microsatellite genotyping to sex eggs and hence to select eight size-matched pairs of ‘late-switching’ colonies from a set of commercial colonies. From one colony of each pair we removed every egg-laying or aggressive worker observed. From the other colony, we simultaneously removed a nonegg-laying, nonaggressive worker. Removed workers were replaced with young workers from separate colonies at equal frequencies within the pair. There was no significant difference in queen productivity between colonies with reduced or normal levels of egg-laying or aggressive workers. Therefore, as predicted, reproductive B. terrestris workers did not significantly reduce the production of new queens.     

Polyandry and colony genetic structure in the primitive ant Nothomyrmecia macrops

The Australian endemic ant Nothomyrmecia macrops is considered one of the most ‘primitive’ among living ants. We investigated the genetic structure of colonies to determine queen mating frequencies and nestmate relatedness. An average of 18.8 individuals from each of 32 colonies, and sperm extracted from 34 foraging queens, were genotyped using five highly variable microsatellite markers. Queens were typically singly (65%) or doubly mated (30%), but triple mating (5%) also occurred. The mean effective number of male mates for queens was 1.37. No relationship between colony size and queen mate number was found. Nestmate workers were related by b=0.61 ± 0.03, significantly above the threshold under Hamilton’s rule over which, all else being equal, altruistic behaviour persists, but queens and their mates were unrelated. In 25% of the colonies we detected a few workers that could not have been produced by the resident queen, although there was no evidence for worker reproduction. Polyandry is for the first time recorded in a species with very small mature colonies, which is inconsistent with the sperm‐limitation hypothesis for the mediation of polyandry levels. Facultative polyandry is therefore not confined to the highly advanced ant genera, but may have arisen at an early stage in ant social evolution.     

Diploid male production in a rare and locally distributed bumblebee, Bombus florilegus (Hymenoptera, Apidae)

The European bumblebee B. terrestris was recently introduced in Japan for agricultural purposes and has now become naturalized. The naturalization of this exotic species may have great detrimental effects on closely related native Japanese bumblebees. The Japanese bumblebee Bombus florilegus is a rare and locally distributed species found in the Nemuro Peninsula of Hokkaido, Japan. In order to assess its population genetics, we estimated the genetic structure of B. florilegus in 16 breeding colonies (queen, workers, and males) and 20 foraging queens by analyzing microsatellite DNA markers. Of the 36 queens analyzed by genotyping and dissection, 32 had been inseminated by a male. The remaining 4 had not been inseminated at all. Of the 4 nonmated queens, one was triploid. Diploid males were found in 4 breeding colonies. Based on the microsatellite data, it appears that B. florilegus has low reproductive success. Since matched mating and nonmating within local populations are high, the extinction risk is correspondingly higher. Our results suggest that conservation of the Japanese B. florilegus is required in order to protect it from both habitat destruction and the naturalization of alien species. Received 19 July 2007; revised 13 October 2007; accepted 15 October 2007.     

Worker interests and male production in Polistes gallicus, a Mediterranean social wasp

The resolution of social conflict in colonies may accord with the interests of the most numerous party. In social insect colonies with single once-mated queens, workers are more closely related to the workers' sons than they are to the queens' sons. Therefore, they should prefer workers to produce males, against the queen's interests. Workers are capable of producing males as they arise from unfertilized eggs. We found Polistes gallicus to have colonies of single, once-mated queens, as determined by microsatellite genotyping of the workers, so worker interests predict worker male production. In colonies lacking queens, workers produced the males, but not in colonies with original queens. Thus worker interests were expressed only when the queen was gone. The high fraction of missing queens and early end to the colony cycle relative to climate so early in the season is surprising and may indicate a forceful elimination of the queen.     

Male parentage and queen mating frequency in the bumblebee <Emphasis Type="Italic">Bombus ignitus</Emphasis> (Hymenoptera: bombinae)

Kin selection theory predicts conflict between queens and workers in the social insect colony with respect to male production. This conflict arises from the haplodiploid system of sex determination in Hymenoptera that creates relatedness asymmetries in which workers are more closely related to the sons of other workers than to those of the queen. In annual hymenopteran societies that are headed by a single queen, the mating frequency of the queen is the only factor that affects the colony kin structure. Therefore, we examined the mating structure of queens and the parentage of males in a monogynous bumblebee, Bombus ignitus, using DNA microsatellites. In the seven colonies that were studied, B. ignitus queens mated once, thereby leading to the prediction of conflict between the queen and workers regarding male production. In each of the five queen-right colonies, the majority of the males (95%) were produced by the colony’s queen. In contrast, workers produced approximately 47% of all the males in two queenless colonies. These results suggest that male production in B. ignitus is a conflict between queen and workers.     

Early collapse of Vespa simillima (Hymenoptera, Vespidae) colonies in central Japan

During the past 15 years in the Ina Valley, Japan, mature fifth instar larvae of the yellow hornet, Vespa simillima, have been occasionally ejected from colonies during September. During 2005, this unusual behavior was particularly widespread, and collection of several V. simillima colonies confirmed that very few fifth instar larvae were present. When compared with an average colony, constructed from 41 colonies collected 20 years previously, colonies in 2005 had 80% fewer fifth instar larvae, despite queen egg‐laying rates remaining similar. It may be that the amount of food provided to the larvae by workers is less than the amount of larval secretions received by workers, which is causing the larvae to become emaciated and preventing them from pupating. This phenomenon normally occurs naturally at the end of the colony cycle in November, when prey abundance decreases and larval secretions or other sources of carbohydrates, for example from ripe fruit, tree sap, or aphid secretions, are needed by sexuals to build up fat reserves. However, another possibility is that the unexplained appearance of this phenomenon in September, which is causing colonies to collapse without producing any or very few sexuals, is due to foragers feeding the larvae prey that are contaminated with insect growth regulators, via pesticides, which are known to prevent successful pupation.     

Ant workers selfishly bias sex ratios by manipulating female development.

Kin selection theory predicts that social insects should perform selfish manipulations as a function of colony genetic structure. We describe a novel mechanism by which this occurs. First, we use microsatellite analyses to show that, in a population of the ant Leptothorax acervorum, workers' relatedness asymmetry (ratio of relatedness to females and relatedness to males) is significantly higher in monogynous (single-queen) colonies than in polygynous (multiple-queen) colonies. Workers rear mainly queens in monogynous colonies and males in polygynous colonies. Therefore, split sex ratios in this population are correlated with workers' relatedness asymmetry. Together with significant female bias in the population numerical and investment sex ratios, this finding strongly supports kin-selection theory. Second, by determining the primary sex ratio using microsatellite markers to sex eggs, we show that the ratio of male to female eggs is the same in both monogynous and polygynous colonies and equals the overall ratio of haploids (males) to diploids (queens and workers) among adults. In contrast to workers of species with selective destruction of male brood, L. acervorum workers therefore rear eggs randomly with respect to sex and must achieve their favoured sex ratios by selectively biasing the final caste (queen or worker) of developing females.     

Colony structure of a slavemaking ant. I. Intracolony relatedness, worker reproduction, and polydomy

Colony and population structure of the obligate slavemaker ant Protomognathus americanus was analyzed via four nuclear microsatellite loci and mitochondrial DNA (mtDNA) markers. Colonies of P. americanus usually contain a single queen, and here we show that she is singly inseminated. Nestmate workers are generally full sisters and their relatedness does not deviate from the expected value of 0.75. Even though colonies were strictly monogynous, we were able to infer that colony takeover by related queens was common and queen replacement by unrelated queens was rare. Polydomy is widespread, with neighboring nests having the same genetic composition. Although we found no evidence of population viscosity or inbreeding from nuclear markers, mtDNA markers provided evidence for small-scale genetic structuring. Haplotype structuring and takeover by related queens suggest philopatry of newly mated queens. In this species, workers reproduce in queenright and queenless nests and worker reproduction accounts for more than 70% of all males. Although sex-ratio theory points to slavemaking ants as important systems for studying queen-worker conflict, our results indicate no basis for such conflict in P. americanus, because extensive worker reproduction generates shifts in relatedness values. Rather, the dual effects of independent polydomous nest units and local resource competition among queens produce male-biased allocation ratios in this species.     

The evolution of worker sterility in honeybees: the genetic basis of failure of worker policing

Worker honeybees (Apis mellifera) usually only lay eggs when their colony is queenless. However, an extremely rare ''anarchistic'' phenotype occurs, in which workers develop functional ovaries and lay large numbers of haploid eggs which develop into adult drones despite the presence of the queen. Studies of such colonies can give important insights into the mechanisms by which worker sterility is maintained in normal colonies. Here we report on the results of a breeding programme which enhanced the frequency of the anarchistic phenotype. Colonies derived from queens inseminated only by worker-laid males showed up to 9% of workers with highly developed ovaries. In these colonies a large proportion of males arose from worker-laid eggs. Colonies headed by queens inseminated with 50% worker-laid drones and 50% queen-laid drones showed variable phenotypes. In most such colonies there was no worker reproduction. In some, many workers had highly developed ovaries, but no worker-laid eggs were reared. In one colony, many worker-laid eggs were reared to maturity. The results suggest that the anarchy phenotype results from a complex interaction of queen genotype, the worker genotype of subfamilies that successfully reproduce and of those that do not, and the external environment.     

Queen mating frequency and maternity of males in the stingless bee Trigona carbonaria Smith

Summary. Aspects of the sociobiology of the native Australian stingless bee Trigona carbonaria (Hymenoptera, Meliponini) were investigated using highly variable microsatellite markers. The queen mating frequency was estimated by examining genotypes of samples of workers from five colonies across three to five microsatellite loci. In each case, results were consistent with the workers being progeny of a queen mated with a single male. Microsatellite analysis of haploid males from two of the colonies suggested they predominantly arose from queen-laid eggs. As workers are more related to sons-of-workers than to sons-of-queens in monandrous colonies, this is somewhat surprising, and suggests that there may be colony-level costs associated with worker reproduction, or that queens are able to regulate worker reproduction via the ritualized cell provisioning and oviposition process. Males from another T. carbonaria colony were found to be diploid.     

Investigation of the population genetic structure and mating system in the ant Pheidole pallidula

The origin of eusociality in haplo-diploid organisms such as Hymenoptera has been mostly explained by kin selection. However, several studies have uncovered decreased relatedness values within colonies, resulting primarily from multiple queen matings (polyandry) and/or from the presence of more than one functional queen (polygyny). Here, we report on the use of microsatellite data for the investigation of sociogenetic parameters, such as relatedness, and levels of polygyny and polyandry, in the ant Pheidole pallidula. We demonstrate, through analysis of mother-offspring combinations and the use of direct sperm typing, that each queen is inseminated by a single male. The inbreeding coefficient within colonies and the levels of relatedness between the queens and their mate are not significantly different from zero, indicating that matings occur between unrelated individuals. Analyses of worker genotypes demonstrate that 38% of the colonies are polygynous with 2-4 functional queens, and suggest the existence of reproductive skew, i.e. unequal respective contribution of queens to reproduction. Finally, our analyses indicate that colonies are genetically differentiated and form a population exhibiting significant isolation-by-distance, suggesting that some colonies originate through budding.     

Kin selection, relatedness, and worker control of reproduction in a large-colony epiponine wasp, Brachygastra mellifica

Hamilton's kin selection theory predicts conflicts of interestamong relatives, even within highly cooperative social insectsocieties. Because workers are the most numerous caste, collectiveworker interests may be an important force in determining theoutcome of conflicts. In this study, we used genotypes fromtwo DNA microsatellite loci to show that two kinds of collectiveworker interests are satisfied in Brachygastra mellifica, amember of the multiqueen epiponine wasps. First, from the highrelatedness of queens (0.66) and the fact that queens are singlymated (shown by genotyping their stored sperm), we calculatedthat new queens are reared in colonies with a harmonic meanof 1.2 old queens, whereas males are reared in colonies withmuch higher queen numbers. This split sex ratio result is predictedunder worker control. It matches other studies of epiponines,but B. mellifica has much larger mature colonies (averaging7951 adults) with many more queens (averaging 398), showingthe pattern holds for large-colony species. Second, we reportthe first genetic data on parentage of males in epiponines andshow that these are also consistent with collective worker interests.Workers are on average significantly more related to queensthan to other workers (r = .37 versus .23) and should thereforesuppress each other and allow the queens to lay haploid (male)eggs. Though many workers have developed ovaries and could layeggs, the genetic analyses showed that most or all males comefrom queens.     

Queen condition,mating frequency,queen loss,and levels of worker reproduction in the hornets Vespa affinis and V. simillima

Abstract 1. Mature queens of the hornet Vespa affinis are unusual in that they lose all their body hair and wings. This has been used as evidence for matricide in hornets. Factors favouring matricide such as low paternity and long annual lifecycle are present in most hornets. 2. We determined paternity in V. affinis and compared queen condition, levels of queen loss, and worker reproduction in 163 V. affinis and 63 V. simillima colonies. These two species have very similar lifecycles and colony sizes, but different paternities (1.3 vs 2.4). Furthermore, in queen‐right V. affinis colonies small proportions (4%) of reproductive workers were present, whereas in queen‐right V. simillima colonies no reproductive workers were detected. 3. Despite these differences, queen damage in both species was influenced predominantly by her age and in V. affinis was not influenced by presence of reproductive workers. 4. The number of queen‐less V. simillima (9/63 = 14%) and V. affinis (28/163 = 17%) colonies were also not significantly different (P = 0.8). Therefore, queen damage and proportions of queen‐less colonies were not linked making it a poor indicator of matricide. 5. It is proposed that graded control of sexual and worker production in wasps and hornets prevents a situation where kin‐selected matricide is ever favoured.     

Differential time allocation of foraging workers in the subterranean termite

Background

Worker reproduction has an important influence on the social cohesion and efficiency of social insect colonies, but its role in the success of invasive ants has been neglected. We used observations of 233 captive colonies, laboratory experiments, and genetic analyses to investigate the conditions for worker reproduction in the invasive Anoplolepis gracilipes (yellow crazy ant) and its potential cost on interspecific defence. We determined the prevalence of worker production of males and whether it is triggered by queen absence; whether physogastric workers with enlarged abdomens are more likely to be reproductive, how normal workers and physogastric workers compare in their contributions to foraging and defence; and whether worker-produced males and males that could have been queen- or worker-produced differ in their size and heterozygosity.

Results

Sixty-six of our 233 captive colonies produced males, and in 25 of these, some males could only have been produced by workers. Colonies with more workers were more likely to produce males, especially for queenless colonies. The average number of days between the first appearance of eggs and adult males in our colonies was 54.1 ± 10.2 (mean ± SD, n = 20). In our laboratory experiment, queen removal triggered an increase in the proportion of physogastric workers. Physogastric workers were more likely to have yolky oocytes (37–54.9%) than normal workers (2–25.6%), which is an indicator of fertile or trophic egg production. Physogastric workers were less aggressive during interspecific aggression tests and foraged less than normal workers. The head width and wing length of worker-produced males were on average 4.0 and 4.3% greater respectively than those of males of undetermined source. Our microsatellite DNA analyses indicate that 5.5% of worker-produced males and 14.3% of males of undetermined source were heterozygous, which suggests the presence of diploid males and/or genetic mosaics in A. gracilipes.

Conclusions

Our experimental work provides crucial information on worker reproduction in A. gracilipes and its potential cost to colony defence. The ability of A. gracilipes workers to produce males in the absence of queens may also contribute to its success as an invasive species if intranidal mating can take place between virgin queens and worker-produced males.

     

It's good to be queen: classically eusocial colony structure and low worker fitness in an obligately social sweat bee

Lasioglossum malachurum, a bee species common across much of Europe, is obligately eusocial across its range but exhibits clear geographic variation in demography and social behaviour. This variation suggests that social interactions between queens and workers, opportunities for worker oviposition, and patterns of relatedness among nest mates may vary considerably, both within and among regions. In this study, we used three microsatellite loci with 12-18 alleles each to examine the sociogenetic structure of colonies from a population at Agios Nikolaos Monemvasias in southern Greece. These analyses reveal that the majority of colonies exhibit classical eusocial colony structure in which a single queen mated to a single male monopolizes oviposition. Nevertheless, we also detect low rates of multiqueen nest founding, occasional caste switching by worker-destined females, and worker oviposition of both gyne and male-producing eggs in the final brood. Previous evidence that the majority of workers show some ovarian development and a minority (17%) have at least one large oocyte contrasts with the observation that only 2-3% of gynes and males (the so-called reproductive brood) are produced by workers. An evaluation of the parameters of Hamilton's Rule suggests that queens benefit greatly from the help provided by workers but that workers achieve greater fitness by provisioning and laying their own eggs rather than by tending to the queen's eggs. This conflict of interest between the queen and her workers suggests that the discrepancy between potential and achieved worker oviposition is due to queen interference. Comparison of relatedness and maternity patterns in the Agios Nikolaos Monemvasias population with those from a northern population near Tübingen, Germany, points to a north-south cline of increasingly effective queen control of worker behaviour.     

Colony structure and reproductive sharing among queens in a tropical paper wasp, <Emphasis Type="Italic">Polistes olivaceus</Emphasis>

Reproductive partitioning among group members is a key feature in social Hymenoptera. We investigated the genetic colony structure of a tropical paper wasp Polistes olivaceus, with an emphasis on variation in the number of queens and reproductive sharing among queens. Among 22 P. olivaceus colonies, 6 were monogynous, 9 polygynous, and 7 were queenless. Adults and brood (eggs and larvae) were genotyped based on six polymorphic microsatellite loci. In each of the polygynous colonies, progenies were assigned to their mothers using maximum-likelihood methods. Nestmate queens were full sisters. The vast majority of reproduction appeared to be monopolized by the dominant queen (α), and the overall reproductive skews were 0.63 ± 0.04 (B index) and 0.97 ± 0.02 (S _c index). Although all nestmate queens had equal reproductive potential, the high magnitude of reproductive skew was enigmatic in this species. Although 9.55 ± 2.07 workers contained developed ovaries in 11 of 15 queen-right colonies, they were unrelated to the nestmate queens but related to each other as full sisters, suggesting that they were the remaining offspring of superseded queens. In 2 of the 11 colonies, we detected male eggs produced by reproductive workers. On average, 7.27 % of the total genotyped male eggs were derived from reproductive workers among the colonies. These results suggest three possibilities regarding the presence of reproductive workers in the P. olivaceus colonies: drifting between colonies, putative remaining offspring from superseded queens, and the offspring of unrelated females who joined the colonies and reproduced there. We found no worker-derived larvae or adult males, suggesting that male eggs were removed by nestmates at some point between oviposition and hatching.