Novel microsatellite DNA loci for Bombus terrestris (Linnaeus, 1758)

We present details and characteristics of 123 novel polymorphic microsatellite DNA loci for Bombus terrestris. Thirty-four of these loci have been tested in nine other Bombus species and 25 of them showed polymorphisms in at least one species. These microsatellite DNA loci together with the already established 60 loci will be useful for characterizing wild and managed populations of B. terrestris and other Bombus species as well as for detailed genetic studies in including mapping studies and genome annotations.     

SERINE PROTEASE FROM MIDGUT OF Bombus terrestris MALES

A serine protease was isolated from midguts of the bumblebee male Bombus terrestris by a combination of precipitation procedures with column chromatography. The purified enzyme exhibited two bands with molecular masses of 25 and 26 kDa as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. These bands showed a proteolytic activity in zymography assay. Midgut enzymes showed optimum proteolytic activity at pH 9 and 35°C using N‐succinyl‐L‐alanyl‐L‐alanyl‐L‐prolyl‐L‐phenyl‐alanine 4‐nitroanilide as a substrate. The Michaelis constant (Km) and maximum reaction rate (Vmax) were 0.55 ± 0.042 mM and 0.714 ± 0.056 μmol p‐nitroalanine produced min^?1 mg protein^?1, respectively. Inhibition was affected by trypsin inhibitor, but not by phenylmethylsulfonyl fluoride and N‐tosyl‐L‐phenylalanine chloromethyl ketone, which indicated the trypsin‐like but not chymotrypsin‐like specificity. The identity of the serine protease was confirmed by nanoliquid‐tandem mass spectrometry. Eleven unique peptides of the B. terrestris serine protease were found. It shows high homology to a previously reported B. ignitus serine protease covering more than 65% of the protein amino acid sequence.     

A method for year-round rearing of cuckoo bumblebees (Hymenoptera: Apoidea: Bombus subgenus Psithyrus)

Summary. The study of the social interactions between host bumblebees and their inquiline species requires rearing them. Here we provide a simple method for rearing of cuckoo bumblebees (Bombus subgenus Psithyrus) in the laboratory. Two bumblebee cuckoo-host systems are used: Bombus (Psithyrus) vestalis hosted by B. (Bombus) terrestris and B. (Psithyrus) sylvestris hosted by B. (Pyrobombus) pratorum. First, the invasion of the Psithyrus female is performed in species-specific host colonies containing approximately 10 young workers. On average, a parasitized colony of B. terrestris produced 90 ± 9 young males and 21 ± 3 young females of B. vestalis, whereas a parasitized colony of B. pratorum produced 16 ± 2 young males and 5 ± 1 young females of B. sylvestris. One week after emergence, Psithyrus virgin females are placed in a flight cage exposed to natural light, with Psithyrus males of other colonies. After successful mating, the females are isolated and overwintered in a cold room at 4°C for 2–4 months. This method enables a year-round and mass breeding of Psithyrus species to facilitate studies of these rare species.     

红光熊蜂卵黄原蛋白基因的cDNA全长序列克隆和表达分析

红光熊蜂Bombus ignitus Smith是许多经济作物和野生植物的重要授粉昆虫之一。卵黄原蛋白基因(vitellogenin,Vg)在昆虫的生殖调控中和行为方面起到重要的作用,本试验对Vg基因全长cDNA的克隆和测序及在蜂王、工蜂和雄性蜂三型蜂中的表达分析得出:Vg基因的全长cDNA为5 481 bp,GenBank中的登录号为FJ913883,有一个完整的开放阅读框(ORF),编码1 772个氨基酸,N-末端的前16个氨基酸为一个信号肽。接近C-末端区域存在保守的GL/ICG基元,其后含有9个半胱氨酸,而且DGXR位于GL/ICG基元上游18个氨基酸残基处。其氨基酸序列与韩国的熊蜂B.ignitus和B.hypocrita相似性高达95%,与西方蜜蜂Apis mellifera的相似性达到51%。Vg的mRNA首先在蜂王蛹期的白眼蛹(Pw)时期出现,其表达量在蜂王整个蛹期发育过程中呈上升趋势,且在黑眼蛹(Pbd)时期达到最高,在成年蜂的脂肪体中的表达量仍在升高,甚至更高。Vg也在工蜂蛹期的白眼蛹(Pw)时期被检测到,然后在整个蛹期发育过程中呈现上升趋势,在刚羽化出房时达到高峰,Vg的mRNA水平随着成年蜂日龄的增加而增加,到15日龄时达到最高,然后呈现下降趋势。对于雄性蜂,Vg的mRNA虽然卵黄原蛋白基因的mRNA水平几乎在整个蛹期发育阶段都表达,但是表达水平非常低,只有在刚羽化出房时期表达水平较高。     

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.     

Sex, horizontal transmission, and multiple hosts prevent local adaptation of Crithidia bombi, a parasite of bumblebees (Bombus spp.)

Local adaptation within host-parasite systems can evolve by several non-exclusive drivers (e.g., host species-genetic adaptation; ecological conditions-ecological adaptation, and time-temporal adaptation). Social insects, especially bumblebees, with an annual colony life history not only provide an ideal system to test parasite transmission within and between different host colonies, but also parasite adaptation to specific host species and environments. Here, we study local adaptation in a multiple-host parasite characterized by high levels of horizontal transmission. Crithidia bombi occurs as a gut parasite in several bumblebee species. Parasites were sampled from five different host species in two subsequent years. Population genetic tools were used to test for the several types of adaptation. Although we found no evidence for local adaptation of the parasite toward host species, there was a slight temporal differentiation of the parasite populations, which might have resulted from severe bottlenecks during queen hibernation. Parasite populations were in Hardy-Weinberg equilibrium and showed no signs of linkage disequilibrium suggesting that sexual reproduction is an alternative strategy in this otherwise clonal parasite. Moreover, high levels of multiple infections were found, which might facilitate sexual genetic exchange. The detection of identical clones in different host species suggested that horizontal transmission occurs between host species and underpins the lack of host-specific adaptation.     

Factors influencing dominance behaviour in queenless bumblebee workers (Bombus terrestris)

ABSTRACT. The establishment of a dominance order within seventy eight groups of four callow workers was investigated. The dominant worker establishes its position by means of overt aggression. Aggression starts 20—40 h after confinement. Dominance order becomes established within 2 days of removal from the colony. Once established, the dominance order is maintained by dominant-subordinate interactions.
The dominant worker starts ovipositing at an age of about 6 days. Subordinates rarely lay eggs. In 25% of the groups the dominant worker is superseded by another worker before any oviposition occurs. Once the dominance order is established, the dominant worker inhibits endocrine activity in subordinates.
Differences in body size and in endocrine activity are thought to be responsible for the ranking of workers in the dominance hierarchy. Ranking seems to depend more on the activity of the corpora allata (CA) than on the activity of the ovaries. Ovariectomized workers do not differ from control workers in the performance of dominance behaviour. Dominant ovariectomized workers do not construct egg cells, but they do defend these if they are present.     

短舌熊蜂及红光熊蜂耐寒力的测定

以过冷却点和冰点为评价指标,对短舌熊蜂Bombus terrestris的幼虫,蛹,成年工蜂,成年雄蜂,处女蜂王,越冬后的蜂王及红光熊蜂Bombus ignitus的成年工蜂,成年雄蜂,处女蜂王的耐寒力进行检测。结果显示,短舌熊蜂幼虫期耐寒力最低,其次是蛹,工蜂和雄蜂,处女王较高。耐寒力最高的是越冬后的蜂王,其含水量也最低。将短舌熊蜂与红光熊蜂的成年工蜂,成年雄蜂和处女蜂王三型蜂分别进行对比,结果显示,红光熊蜂成年雄蜂与处女蜂王的耐寒力要比短舌熊蜂高。样本的湿重和含水量与过冷却点和冰点无显著相关性。     

Microsatellite loci for Cherrie's tanager (Ramphocelus costaricensis)

Cherrie's tanager (Ramphocelus costaricensis) is a common woodland bird of the Pacific lowlands of Costa Rica. Males and females have sexually dimorphic plumage and are socially monogamous. As part of a study of sexual selection in this species, we characterized six species‐specific polymorphic microsatellite loci and one cross‐species locus that works well in Cherrie's tanager to ascertain parentage.     

An interspecific comparison of foraging range and nest density of four bumblebee (Bombus) species

Bumblebees are major pollinators of crops and wildflowers in northern temperate regions. Knowledge of their ecology is vital for the design of effective management and conservation strategies but key aspects remain poorly understood. Here we employed microsatellite markers to estimate and compare foraging range and nest density among four UK species: Bombus terrestris, Bombus pascuorum, Bombus lapidarius, and Bombus pratorum. Workers were sampled along a 1.5-km linear transect across arable farmland. Eight or nine polymorphic microsatellite markers were then used to identify putative sisters. In accordance with previous studies, minimum estimated maximum foraging range was greatest for B. terrestris (758 m) and least for B. pascuorum (449 m). The estimate for B. lapidarius was similar to B. pascuorum (450 m), while that of B. pratorum was intermediate (674 m). Since the area of forage available to bees increases as the square of foraging range, these differences correspond to a threefold variation in the area used by bumblebee nests of different species. Possible explanations for these differences are discussed. Estimates for nest density at the times of sampling were 29, 68, 117, and 26/km2 for B. terrestris, B. pascuorum, B. lapidarius and B. pratorum, respectively. These data suggest that even among the most common British bumblebee species, significant differences in fundamental aspects of their ecology exist, a finding that should be reflected in management and conservation strategies.     

Microsatellite loci for the Chinese bamboo rat Rhizomus sinensis

We describe 19 polymorphic microsatellite loci for the bamboo rat Rhizomus sinensis. When tested with 20 samples from a single population, these loci exhibited a mean of 4.5 alleles per locus and a mean expected heterozygosity of 0.612. All loci were in Hardy-Weinberg equilibrium and no evidence for linkage disequilibrium was detected between any loci. These loci will be useful for studying population genetic diversity and differentiation in the Chinese bamboo rat.     

Microsatellite loci for the Siberian flying squirrel,Pteromys volans

We report the isolation and characterization of polymorphic microsatellite loci for the Siberian flying squirrel Pteromys volans. The seven most useful loci had between six and 11 alleles and expected heterozygosities ranging from 0.477 to 0.866. We also tested the utility of these loci in other squirrel species, northern flying squirrels (Glaucomys sabrinus and G. volans) and the common red squirrel (Sciurus vulgaris). Three of the Siberian flying squirrel loci were polymorphic in other squirrel species, suggesting a limited potential for cross‐species use.     

Microsatellite loci within sixgill sharks, Hexanchus griseus

Novel sixgill shark (Hexanchus griseus) microsatellite loci were developed and tested on five shark species. A suite of microsatellite loci previously developed for lemon sharks (Negaprion brevirostris) was also tested. Data on 15 microsatellites are presented including primer sequences, number of alleles (a), observed (H_O) and expected heterozygosities (H_E), and F_IS values for sixgill sharks (a = 10–69, H_O = 0.24–1.00, H_E = 0.76–0.96 and F_IS = –0.21–0.60), sevengill sharks (Notorynchus cepedianus) (a = 6–40, H_O = 0.20–0.73, H_E = 0.59–0.94 and F_IS = –0.47–0.58), Pacific spiny dogfish (Squalus acanthias) (a = 3–13, H_O = 0.00–0.96, H_E = 0.24–0.93 and F_IS =–0.52–1.00), angle sharks (Squatina californica) (a = 1–4, H_O = 0.00–1.00, H_E = 0.60–1.00 and F_IS =–1.00–0.25), and leopard sharks (Triakis semifasciata) (a = 3–16, H_O = 0.20–1.00, H_E = 0.53–0.92 and F_IS = –0.57–1.00). A final suite of 14 microsatellites (13 developed from sixgill sharks and one from lemon sharks) were found to be polymorphic and conform to Hardy–Weinberg equilibrium within sixgill sharks.     

Microsatellite loci for Ponto-Caspian gobies: markers for assessing exotic invasions

We developed and tested eight polymorphic microsatellite loci for Ponto‐Caspian ‘neogobiin’ gobies, many of which are invasive in Eurasia and North America, whose study will aid understanding of the population genetics underlying their success. We tested samples from one to two locations from 12 taxa in the recently revised genera Babka, Benthophilus, Mesogobius, Neogobius = Apollonia, Ponticola and Proterorhinus; including the bighead, Caspian, knout, monkey, racer, round, tadpole and tubenose gobies; and taxa from introduced vs. native populations, those diverging between fresh and marine waters, and those differentiated between the Black and Caspian Seas. Populations conformed to Hardy–Weinberg equilibrium expectations, averaging five to 15 alleles per locus and 0.11 to 0.67 mean heterozygosity. Allelic variation significantly differentiated among all taxa and populations.     

Microsatellite loci from red sea urchins (Strongylocentrotus franciscanus)

Strongylocentrotus franciscanus is a keystone marine herbivore in the Pacific North West of North America, that is commercially harvested for its roe in wild fisheries and is under development as an aquaculture species. We report the development of 11 novel, highly polymorphic di‐ and trinucleotide loci for use in population studies of this species. In a survey of over 200 individuals from three coastal sites in British Columbia, observed heterozygosities of the loci ranged from 0.39 to 0.85 and an average of 26 alleles per locus were resolved. Seven of the 11 microsatellite loci were in Hardy–Weinburg equiulibruim (HWE).     

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.     

Microsatellite loci for the mayfly Baetis rhodani (Baetidae,Ephemeroptera)

The first microsatellite primers are described for the mayfly family Baetidae (order Ephemeroptera). Seven polymorphic loci were isolated and characterized for the species Baetis rhodani from two enriched genomic libraries. A total of 183 individuals were genotyped using these loci. Allelic diversity was high for all loci, and observed heterozygosities ranged between 0.382 and 0.772. A heterozygote deficiency was detected in some loci, suggesting the presence of ‘null’ alleles.     

Microsatellite evolution at two hypervariable loci revealed by extensive avian pedigrees

Genealogies generated through a long-term study of superb fairy-wrens (Malurus cyaneus) were used to investigate mutation within two hypervariable microsatellite loci. Of 3,230 meioses examined at the tetranucleotide locus (Mcy micro 8), 45 mutations were identified, giving a mutation rate of 1.4%. At the dinucleotide locus (Mcy micro 4) 30 mutations were recorded from 2,750 meioses giving a mutation rate of 1.1%. Mutations at both loci primarily (80%; 60/75) involved the loss or gain of a single repeat unit. Unlike previous studies, there was no significant bias toward additions over deletions. The mutation rate at Mcy micro 8 increased with allele size, and very long alleles (>70 repeats) mutated at a rate of almost 20%. The length of the mutating allele and allele span, however, were strongly correlated so it was not possible to isolate the causative factor. Allele size did not appear to affect mutation rate at Mcy micro 4, but the repeat number was considerably lower at this locus. The gender of the mutating parent was significant only at Mcy micro 8, where mutations occurred more frequently in maternal alleles. However, at both loci we found that alleles inherited from the mother were on average larger than those from the father, and this in part drove the higher mutation rate among maternally inherited alleles at Mcy micro 8.