Mitochondrial DNA sequence variation of the bumblebee,Bombus ardens (Hymenoptera: Apidae)

The bumblebee, Bombus ardens, is a valuable natural resource, and is most notably utilized for greenhouse pollination. In order to gain a greater understanding of the population genetic structure and the genetic diversity of this species, we sequenced a portion of the mitochondrial COI gene corresponding to the “DNA Barcode” region (658 bp) from 160 individuals collected over 15 Korean localities. Uncorrected pairwise distances among the eight haplotypes suggested low intraspecific genetic diversity, with a maximum sequence divergence of 0.3%. Such a low level of intraspecific genetic diversity was further reflected in local populations, particularly to islet populations, such as those of Youngheungdo, Jakyakdo, and Ulleugdo, which had zero genetic diversity. Geographically, one haplotype (BARBA01) was widespread and dominant, with a frequency of 90.6% (145 among 160 individuals). Other haplotypes were restricted to one to three localities and had low frequency. Overall, a very high rate of per generation female migration ratio (Nm = 4.6 to infinite) and a very low level of genetic fixation (F_ST = 0 to 0.099) were detected between pairs of localities, suggesting that the B. ardens populations on the Korean peninsula are panmictic, which is consistent with our understanding of their dispersal capability.     

小峰熊蜂访花偏爱性

传粉昆虫在访花时,通常会表现出对某一类型花的偏爱性。本研究利用人工制作的大小、颜色、形态和气味不同的9种类型的花来研究小峰熊蜂Bombus hypocrita的访花偏爱性。结果表明：当增加花的大小、形态、气味等附加特征数时,小峰熊蜂的偏爱性程度与花朵附加特征数有显著相关性（P<0.01）。当花朵颜色由2种增加到4种,熊蜂对紫色花的偏爱性程度降低,但花朵颜色的种类与小峰熊蜂的访花偏爱性没有相关性（P>0.05）,花颜色的种类对熊蜂访紫色花的偏爱性影响不大。大小为5 cm的紫花被访次数（108±9次）明显高于大小为3 cm的紫花被访次数（40±4次）（P<0.01）,说明熊蜂明显喜欢访大花瓣的紫花。完全盛开的紫花被访次数（129±13次）显著高于刚绽放的花被访次数（26±3次）（P<0.01）,说明熊蜂喜欢访盛开的紫花。柠檬味的紫花被访次数（63±8次）明显低于草莓味的紫色花被访次数（88±2次）（P<0.05）,说明熊蜂喜欢访草莓味的花朵。     

西方蜜蜂和兰州熊蜂在设施桃园的访花偏好性比较

【目的】为了比较西方蜜蜂 Apis mellifera 和兰州熊蜂 Bombus lantschouensis 在设施桃园内对不同时期桃花的访花偏好性、以及这种偏好性与花粉活力和采集花粉花蜜之间的关系。【方法】记录2种蜂在温室桃园内访问早期花、中期花和晚期花的比例,测定桃花不同时期的花粉活力以及2种蜂携带的花粉活力,观察2种蜂采集花蜜和采集花粉的成功率,统计2种蜂访花过程中桃花所处的枝条数及植株数。【结果】桃花不同时期的花粉活力差异显著,早期花花药未开裂,花粉未释放,中期花花粉活力（58.3%）显著高于晚期花花粉活力（34.2%）（P<0.01）;西方蜜蜂更加偏好访问中期花,对中期花的访问率高达75.3%,显著高于兰州熊蜂对中期花的访问率（49.2%）（P<0.01）;西方蜜蜂携带的花粉活力（92.1%）显著高于兰州熊蜂携带的花粉活力（72.9%）,但是西方蜜蜂采集花粉和采集花蜜的成功率均低于兰州熊蜂（P<0.01）;在访问一定数量的桃花过程中,兰州熊蜂在设施桃园内访问的枝条数和植株数较多,分布范围较广（P<0.01）。【结论】和兰州熊蜂相比,西方蜜蜂对活力花粉的辨别能力更强,更加偏好访问花粉活力较高的花朵,这种偏好性导致其采集花粉花蜜的成功率降低。     

Workers dominate male production in the neotropical bumblebee Bombus wilmattae (Hymenoptera: Apidae)

Background

Cooperation and conflict in social insects are closely linked to the genetic structure of the colony. Kin selection theory predicts conflict over the production of males between the workers and the queen and between the workers themselves, depending on intra-colonial relatedness but also on other factors like colony efficiency, sex ratios, cost of worker reproduction and worker dominance behaviour. In most bumblebee (Bombus) species the queen wins this conflict and often dominates male production. However, most studies in bumblebees have been conducted with only a few selected, mostly single mated species from temperate climate regions. Here we study the genetic colony composition of the facultative polyandrous neotropical bumblebee Bombus wilmattae, to assess the outcome of the queen-worker conflict over male production and to detect potential worker policing.

Results

A total of 120 males from five colonies were genotyped with up to nine microsatellite markers to infer their parentage. Four of the five colonies were queen right at point of time of male sampling, while one had an uncertain queen status. The workers clearly dominated production of males with an average of 84.9% +/- 14.3% of males being worker sons. In the two doubly mated colonies 62.5% and 96.7% of the male offspring originated from workers and both patrilines participated in male production. Inferring the mother genotypes from the male offspring, between four to eight workers participated in the production of males.

Conclusions

In this study we show that the workers clearly win the queen-worker conflict over male production in B. wilmattae, which sets them apart from the temperate bumblebee species studied so far. Workers clearly dominated male production in the singly as well the doubly mated colonies, with up to eight workers producing male offspring in a single colony. Moreover no monopolization of reproduction by single workers occurred.     

Male production by non-natal workers in the bumblebee, Bombus deuteronymus (Hymenoptera: Apidae)

Social insect societies are considered to be composed of many extremely cooperative individuals. While workers are traditionally believed to behave altruistically, recent studies have revealed behaviors that are more selfish. One such example is intraspecific social parasitism, where workers invade conspecific colonies and produce male offspring that are reared by unrelated host workers. Such intraspecific parasitism has been reported in honeybees (Apis cerana, and A. florea) and “semi-wild” bumblebee colonies of Bombus terrestris. Here we report on intraspecific social parasitism by workers in “wild” colonies of the bumblebee B. deuteronymus. Three of the 11 B. deuteronymus colonies studied were invaded by non-natal workers, of which 75% became reproductive and produced 19% of the adult males. The invading non-natal workers produced significantly more males than resident natal workers and the non-natal brood was not discriminated against by the natal workers.     

乙基多杀菌素和联苯肼酯对地熊蜂的毒性及风险评估

【目的】明确乙基多杀菌素和联苯肼酯对地熊蜂Bombus terrestris的毒性, 探讨这两种农药亚致死浓度对地熊蜂体内乙酰胆碱酯酶(AchE)、谷胱甘肽-S-转移酶(GST)和羧酸酯酶(CarE) 3种解毒酶活性的影响。【方法】采用饲喂法测定60 g a.i./L乙基多杀菌素和43%联苯肼酯对地熊蜂采集蜂的急性经口毒性,依据农药对蜜蜂生态风险的危害熵(hazard quotient, HQ)值评估这两种农药对地熊蜂的风险。同时测定了这两种农药亚致死剂量(LD₅₀和LD₈₀)处理后地熊蜂AchE, GST和CarE的活性变化。【结果】60 g a.i./L乙基多杀菌素和43%联苯肼酯对地熊蜂采集蜂的急性经口毒性测定48 h时LD₅₀值分别为3.590和1 447 μg a.i./蜂,其中60 g a.i./L乙基多杀菌素表现为中毒,43%联苯肼酯表现为低毒。两种农药对地熊蜂采集蜂的HQ值均低于50,表现为低风险。LD₅₀和LD₈₀剂量的乙基多杀菌素处理组与对照组相比,3 h时地熊蜂AchE活性被激活,显著高于对照组(P＜0.05),分别为对照组的1.45和1.23倍,24 h后活性受到抑制,两个剂量处理组AchE活性均显著低于对照组(P＜0.05);CarE活性3 h时同样被激活,显著高于对照组(P＜0.05),LD₅₀和LD₈₀剂量处理组CarE活性分别为对照组的1.24和1.53倍, 24 h后活性受到抑制,其中LD₅₀剂量处理组CarE活性显著低于对照组(P＜0.05),LD₈₀剂量处理组CarE活性与对照组差异不显著(P＞0.05);LD₅₀和LD₈₀剂量处理组GST活性3 h被激活,显著高于对照组(P＜0.05),分别为对照组的2.24和2.58倍,24 h后活性降低,但两个剂量处理组GST活性仍显著高于对照组(P＜0.05)。43%联苯肼酯处理后,与对照组相比3 h时LD₅₀和LD₈₀剂量处理组AchE活性与对照组差异不显著(P＞0.05),24 h后AchE活性降低,显著低于对照组(P＜0.05),分别是对照组的75%和80%;CarE活性3 h时被抑制,LD₅₀剂量处理组CarE活性显著低于对照组(P＜0.05),LD₈₀剂量处理组CarE活性低于对照组,但差异不显著(P＞0.05),24 h后CarE活性被激活,其中LD₅₀剂量处理组CarE活性高于对照组,但差异不显著(P＞0.05),LD₈₀剂量处理组CarE活性显著高于对照组(P＜0.05);LD₅₀剂量处理组GST活性3 h时被激活,显著高于对照组(P＜0.05),24 h后活性降低,但仍显著高于对照组(P＜0.05),3 h和24 h的活性分别为对照组的2.04和1.72倍,LD₈₀剂量处理组3 h的GST活性与对照组无显著差异(P＞0.05),24 h后活性降低,显著低于对照组(P＜0.05)。【结论】乙基多杀菌素和联苯肼酯对地熊蜂的HQ 评估均表现为低风险,其中联苯肼酯对地熊蜂的安全性较高,在熊蜂授粉过程中可以按照推荐剂量应用,但过量施用或者长期施用可能会造成熊蜂体内药剂积累引起生理或者行为的变化,乙基多杀菌素在温室及大田授粉期的使用剂量和方法有待进一步研究。     

Hazards of insecticides to the bumble bees Bombus impatiens (Hymenoptera: Apidae) foraging on flowering white clover in turf

Insecticides used on turf are sometimes applied to areas with flowering weeds that attract honey bees and native pollinators. We tested residual effects of such treatments on colony vitality and behavior of the bumble bees Bombus impatiens Cresson foraging on turf containingwhite clover, Trifolium repens L. Imidacloprid, a syst emic chloronicotinyl used for preventive control of root-feeding grubs, was applied as granules, followed by irrigation, or sprayed as a wettable powder, with or without irrigation. Hives were confined on the plots in large field cages after residues had dried and colony vitality (i.e., numbers of brood, workers, and honey pots, and weights of queens, workers, and whole colonies with hives) was evaluated after 28-30 d. Workers' foraging activity and defensive response to an aggressive stimulus also were evaluated. In another test, weedy turf was sprayed with chlorpyrifos, carbaryl, or cyfluthrin at labeled rates for surface-feeding pests. Bee colonies were confined on the plots after residues had dried, with effects on colony vitality evaluated after 14 d. Finally, foraging activity of wild bumble bees was monitored on open plots to determine if insecticide-treated areas were avoided. Imidacloprid granules, and imidacloprid sprays applied with posttreatment irrigation, had no effect on colony vitality or workers' behavior, suggesting that such treatments pose little systemic or residual hazard to bumble bees. In contrast, exposure to dry nonirrigated residues of all of the aforementioned insecticides had severe impact on colony vitality. Foraging workers did not avoid insecticide-treated areas. Means by which turf managers can reduce hazards of insecticide applications to pollinators are discussed.     

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.     

密林熊蜂雄性成蜂生殖系统发育动态

为了研究密林熊蜂Bombus patagiatus Nylander雄蜂出房后生殖系统发育变化过程, 本研究首次对密林熊蜂雄性成蜂（1-15日龄）生殖系统的精液量、 精子数、 性腺大小、 储精囊大小等进行了测定。结果显示： 雄性成蜂在1日龄时精液量最大, 达到5.95±0.39 μL; 镜检精子数在2日龄最多; 而精液中活动精子数出现波浪式的循环趋势, 9日龄达到最大值, 为25.93±1.06个; 精液中精子总数在9日龄达到最大值, 为160.67±17.11万个; 活动精子百分比以8、 9日龄最高, 分别达到86.38%±2.02%及86.45%±2.50%; 性腺长度在1日龄时最小, 为1 522.01±37.93 μm; 储精囊的中部宽度在7日龄最宽, 达86.38±2.96 μm, 但与9日龄相比无显著差异。由此可见, 8-9日龄雄性成蜂个体各指标较为稳定, 该日龄即为雄性成蜂性成熟且可以交配的时期。本研究结果为密林熊蜂人工繁育提供了理论依据。     

The first record of the rare oligolectic bumblebee Bombus gerstaeckeri Morawitz (Hymenoptera: Apidae: Bombini) lrom Ukraine

The oligolectic bumblebee Bombus gerstaeckeri Morawitz has been recorded in the Ukrainian Carpathians for the first time during field research from 2002 to 2007. Queens and workers of the species have been observed foraging on two Aconitum spp. within 540 m and 1400 m altitude. The additional forage resource of this species, that is Salvia glutinosa L., has been recorded at low height. Three competitor bumblebees, not confined only to Aconitum spp. have been observed foraging on monkshoods: B. hortorum (L.), B. pascuorum (Scopoli) and B. wurflenii Radoszkowski.     

Morphometric and molecular identification of the female castes of Bombus ignitus and B. ardens (Apidae: Hymenoptera)

Among Korean bumblebees, Bombus ignitus and B. ardens are relatively abundant and important for pollination of wildflowers and agricultural crops. Although the males are easily distinguishable phenotypically, the female castes are difficult to identify from each other. Here we evaluated the value of some morphometric characters in species identification. Also, we developed a polymerase chain reaction followed by restriction fragment length polymorphism (PCR-RFLP) to discriminate these similar species. In spite of statistically significant differences of some morphological characters between two species, overlapping quantitative traits hindered accurate identification of the species. However, using 435 bp of COI gene and AluI, BspHI and Earl restriction enzymes allowed molecular identifications of these two species with unique profiles from the digestion by these restriction enzymes. This method can also be applied for older specimens with some morphological characters damaged. We also developed species-specific primers for fast and cost-effective identification of these species.     

Cloning and characteristics of the antibacterial peptide gene abaecin in the bumblebee Bombus lantschouensis (Hymenoptera: Apidae)

Among the antimicrobial peptides, abaecin is rich in proline content and plays a vital role in insect innate immune defense. Here, the full-length gene of abaecin from the bumblebee Bombus lantschouensis was cloned, and its expression profiles for different tissues, developmental stages and reproductive statuses were analyzed by RT-qPCR. Meanwhile, the responses of abaecin to a bacterium (Escherichia coli) and a fungus (Beauveria bassiana) were tested. The full length of abaecin cDNA was 470 bp, and the open reading frame (ORF) was 258 bp, encoding a polypeptide of 85 amino acids. The abaecin gene consists of three exons and two introns. Phylogenetic analysis showed that Bombus ignitus was the closest species to B. lantschouensis base on putative Abaecin protein sequence. Expression analysis showed that abaecin was expressed broadly in different tissues, with the highest expression in fat bodies and extremely low expression in antennae. Regarding developmental stage, low expression of abacein was detected in eggs and larvae, and high expression was detected in pupal stages. The highest expression was observed at the Pw pupal stage (pupae with an unpigmented body cuticle and white eyes), and the expression then decreased from the Pp (pupae with pink eyes) to the Pdd (dark-eye pupae with a dark-pigmented cuticle) stages. In addition, the expression of abaecin was higher in egg-laying than in non-egg-laying female bumblebees. Both E. coli and B. bassiana infections induced the expression of abaecin. Our results indicated that the abaecin gene plays important roles in the development, reproduction and immune responses of bumblebees. During the artificial rearing of bumblebees, a good environment should be created to avoid infection with bacteria or fungi.     

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.     

The complete nucleotide sequence and gene organization of the mitochondrial genome of the bumblebee, Bombus ignitus (Hymenoptera: Apidae)

The complete 16,434-bp nucleotide sequence of the mitogenome of the bumble bee, Bombus ignitus (Hymenoptera: Apidae), was determined. The genome contains the base composition and codon usage typical of metazoan mitogenomes. An unusual feature of the B. ignitus mitogenome is the presence of five tRNA-like structures: two each of the tRNALeu(UUR)-like and tRNASer(AGN)-like sequences and one tRNAPhe-like sequence. These tRNA-like sequences have proper folding structures and anticodon sequences, but their functionality in their respective amino acid transfers remained uncertain. Among these sequences, the tRNALeu(UUR)-like sequence and the tRNASer(AGN)-like sequence are seemingly located within the A+T-rich region. This tRNASer(AGN)-like sequence is highly unusual in that its sequence homology is very high compared to the tRNAMet of other insects, including Apis mellifera, but it contains the anticodon ACT, which designates it as tRNASer(AGN). All PCG and rRNAs are conserved in positions observed most frequently in insect mitogenome structures, but the positions of the tRNAs are highly variable, presenting a new arrangement for an insect mitogenome. As a whole, the B. ignitus mitogenome contains the highest A+T content (86.9%) found in any of the complete insects mt sequences determined to date. All protein-coding sequences started with a typical ATN codon. Nine of the 13 PCGs have a complete termination codon (all TAA), but the remaining four genes terminate with the incomplete TA or T. All tRNAs have the typical clover-leaf structures of mt tRNAs, except for tRNASer(AGN), in which the DHU arm forms a simple loop. All anticodons of B. ignitus tRNAs are identical to those of A. mellifera. In the A+T-rich region, a highly conserved sequence block that was previously described in Orthoptera and Diptera was also present. The stem-and-loop structures that may play a role in the initiation of mtDNA replication were also found in this region. Phylogenetic analysis among three corbiculate tribes, represented by Melipona bicolor (Meliponini), A. mellifera (Apini), and B. ignitus (Bombini), showed the closest relationship between M. bicolor and B. ignitus.     

Studies on the feeding of Sitona lineatus L. (Coleoptera: Curculionidae) on white clover (Trifolium repens L.) seedlings

Measurements of feeding damage by sitona weevil (Sitona lineatus L.) adults on differing numbers of seedlings of white clover (Trifolium repens L.) at the first and fourth trifoliate leaf stage were made in the glasshouse at 20°C. S. lineatus consumed more of the trifoliate component of the seedling. Sitona adults caused significant yield reduction at all levels of plant population. Total clover consumption increased with increasing size of sitona population, but consumption per adult weevil was reduced.     

Nest takeover by queen and its positive impact on colony development in the Japanese bumblebee Bombus ignitus (Apidae: Hymenoptera)

Lone Bombus ignitus queens are known to start nests in small underground cavities. To examine the nest‐site preference of post‐hibernating queens, choice tests were carried out by providing queens with orphan colonies and empty cavities as possible nesting sites within an experimental box. Our results showed that the queens had a strong preference for takeover of an orphan colony, suggesting that nest takeover (usurpation) could occur in nature even with the presence of possible empty cavities for nesting. We compared the colony‐growth process and final production of sexuals between non‐takeover and takeover colonies. The increase in the number of egg cups was faster in the takeover colonies, suggesting that orphan broods elicit earlier oviposition by the usurping queen. Reproductives emerged earlier (significant for new queens) and in greater numbers (males) from takeover colonies than from non‐takeover colonies. Thus, post‐hibernating B. ignitus queens would search for and take over small orphan colonies to increase their fitness.     

Reproductive success of a colony of the introduced bumblebee Bombus terrestris (L.) (Hymenoptera: Apidae) in a Tasmanian National Park

Abstract  Ever since a feral population of the bumblebee Bombus terrestris was found in suburban Hobart, on the Australian island of Tasmania, there has been ongoing debate surrounding the capacity of this species to utilise Australian native vegetation. Although several studies have reported B. terrestris foraging on Tasmanian native vegetation, doubts have been raised as to whether this reflects successful breeding in native vegetation. This study documents the success of a colony of B. terrestris in a Tasmanian National Park, isolated from urban and agricultural areas by 10 km of sea. Examination of the larval cocoons revealed that this colony had produced at least 304 new queens and 939 workers and drones. Pollen stores found in the colony were mostly from native plants, particularly Eucalyptus. These results strongly suggest that B. terrestris is able to reproduce successfully in parts of Australia that still support almost exclusively native vegetation.     

Ousting of the Common Redstart (Aves: Turdidae: Phoenicurus phoenicurus) from its nests by the bumblebee Bombus niveatus vorticosus (Hymenoptera: Apidae)

During a study of bird nesting in SW. Anatolia, 125 man-made nest boxes have been installed, 3 m high, in the trees. These nest boxes have been occupied by several bird species (Phoenicurus phoenicurus, Parus major, Parus ater, Sitta krueperi, Certhia brachydactyla). Of the 48 Redstart nests, 19 (40 %) had been successfully invaded by the bumblebee Bombus niveatus vorticosus. This invasion occurred at different times in the nest building cycle: - during the building of the nest, - during the incubation or - after the hatching of the young. Once installed in the nest of the Redstart, the bumblebee does not directly attack the bird, but disturbs it by continuously rearranging the nesting material and by covering the bird’s brood. Eventually, the bird deserts its nest and brood. The Redstart does not display any agonistic behaviour towards the bumblebee. Nests of other bird species are never invaded.