Abaecin is a proline-rich anti-bacterial peptide which is known to have high activity against Gram-negative bacteria and it is originally isolated from honeybee Apis mellifera. Since the anti-bacterial peptides including abaecin were identified in the hemolymph of A. mellifera infected with Gram-negative bacteria Escherichia coli, this result implies that honeybees express abaecin for defense against natural infection. Anti-microbial peptides including abaecin were also identified in the adult hemolymph of bumblebees. To investigate pharmacological and toxicological properties of abaecin from Bombus ussurensis, biological activities were evaluated by conducting anti-tumor, anti-microbial and cytotoxic activity assays. Abaecin exhibited significantly high anti-tumor activity against ovarian tumor cells SK-OV-3 and NIH; OVCAR-3 at 100 μM after 72 h treatment. Interestingly, abaecin showed none of anti-microbial activity against Gram-negative and Gram-positive bacteria. Since abaecin exhibited extremely low level of hemolytic activity against human erythrocytes, it may serve as a good model peptide for studying its potential as a selective anti-tumor drug.

We investigated pollination and foraging activities of honeybee (Apis mellifera L.) and bumblebee (Bombus terrestris L.) during flowering season of the asian pear (Pyrus pyrifolia N.) under different weather conditions. There was no significant statistical difference about the pollination activities of two species. However, the pollination activities of bumblebee were more active than those of honeybee under low temperature and rainfall period. The activities of honeybee and bumblebee were more influenced by temperature than other factors (i.e. illumination and wind velocity). Honeybee was more sensitive to temperature and illumination than bumblebee. At low temperatures (<20℃) on cloudy days (<30,000 lux) with a certain wind velocity (>4.0 m/s), the pollination activity of the honeybee was lower twice than that of bumblebee.

Bombolitin is a venom peptide originally isolated from bumblebees and possesses various biological activities, including hemolytic activity. Bombolitins exhibit amphipathic α-helical structure in lipid-membrane-mimicking environments. To investigate their pharmacological and toxicological properties, anti-tumor, anti-microbial and cytotoxic activities of bombolitins from Bombus ardens and Bombus ussurensis were evaluated. Bombolitins of the two species exhibited extremely high anti-tumor activity against ovarian tumor cells SK-OV-3 and NIH; OVCAR-3 at 25-50 μM, which is 2-fold more potent than other wasp venom peptides studied to date (Yoon et al., 2015; Yoon et al., 2016). The two bombolitins showed significantly high antimicrobial activity. However, bombolotin of B. ussurensis showed no antimicrobial activity against Escherichia coli. In addition to their high levels of anti-tumor activity, bombolitins showed considerable levels of hemolytic activity. Thus, to utilize bombolitins as a potential candidate for anti-tumor peptide drugs, further studies for reducing cytotoxic properties of bombolitns is essential.

비벡터링(Bee-vectoring)은 벌통의 외부에 약제를 담은 분배장치를 설치하고 뒤영벌(Bombus terrestris)이 외부로 나갈 때 선발된 약제가 들어있는 분배장치를 통과하여 나가게 함으로써 꽃이나 잎 부위에 선발된 약제를 운반하여 병해충 방제에 이용할 수 있는 기술이다. 본 연구에서는 보베리아(Beauveria bassiana, 기적)와 아세타미프리드 수화제(모스피란)를 비벡터링 선발 약제로 선정하고 약제에 대한 뒤영벌 출입 행동을 비교 분석하였다. 출입 행동 조사는 뒤영벌이 가장 많이 활동하는 오전 6시부터 오후 1시까지의 출입 회수를 7일간 조사하였다. 대조구인 무처리 분배장치를 부착한 뒤영벌 벌통에서는 7일간 약 1,500회의 출입을 한 것으로 조사되었으나 보베리아를 설치한 벌통에서는 7일간 약 600회, 아세타미프리 드를 설치한 벌통에서는 1,000회 출입이 관찰되었다. 보베리아 비벡터링 포장효과 조사는 토마토 온실에서 온실가루이에 대한 방제 효과를 조사하였다. 비벡터링 후 처리구의 거리별 온실가루이 밀도는 50m 지점에서 평균 1,339 마리로 높았으나 무처리구의 경우 30m 지점에서 평균 2,295 마리로 높게 조사되었으며 처리구에서 온실가루이의 밀도는 낮은 추세를 보였다.

The honeybee inhibitor cysteine knot (ICK) peptide acts as an antifungal peptide and insecticidal venom toxin. However, the ICK peptide from bumblebees has not been characterized. Here, we report the molecular cloning and antifungal activity of a bumblebee (Bombus ignitus) ICK peptide (BiICK). We identified a BiICK that contains an ICK fold. The BiICK was expressed in the epidermis, fat body, and venom gland of B. ignitus worker bees. A 6.7-kDa recombinant BiICK peptide was expressed in baculovirus-infected insect cells. Recombinant BiICK peptides directly bound to Beauveria bassiana, Ascosphaera apis, and Fusarium graminearum, but they did not bind to Escherichia coli, Paenibacillus larvae, or Bacillus thuringiensis. Consistent with this finding, BiICK exhibited antifungal activity against fungi. These results demonstrate that BiICK acts as an antifungal peptide.

The post-hibernated queens of Korean native bumblebee, Bombus hypocrita saporoensis were collected in Jeong-Sun, Korea to investigate their ecological characteristics and reared under the laboratory conditions; 27℃, 65% R.H. and darkness. The average weight of collected queens was 0.87±0.14g and 0.8g class was most abundant (34.7%) among eight weight classes. The rates of colony initiation, colony foundation and progeny-queen production were 76.3%, 60.0% and 42.5%, respectively. The durations up to colony initiation and colony foundation were 12.0±6.7 days and 49.3±2.4 days, respectively. The duration up to first worker, male and queen emergence were 22.2±3.0 days, 63.4±6.5 days and 63.5±8.7 days, respectively. The number of progenies was 108.5±26.0 workers, 186.7±70.7 males and 39.1±30.3 queens. The longevity of colony founded queen was 90.2±19.9 days whereas that of non-colonized queen was 35.9±30.3 days. Copulation time was 29.5±9.6 min. The number of eggs per egg cell laid new queen was 9.3±2.3 grains. An egg shaped as banana and it’s size was 1.12±0.10 mm in a width, 3.54± 0.16 mm in a length and 2.26±0.34 mg in a weight.

The bumblebee, Bombus terrestris, plays an important role as one of alternative pollinators since the outbreak of honeybee colony collapse disorder. Recently, pathogens and parasites such as viruses, bacteria and mites affecting the life span and fecundity of their host have been discovered in B. terrestris. In this study, in order to detect viral infection in B. terrestris, we collected B. terrestris adults and isolated total RNA for diagnostic PCR. The PCR primers specific for pathogenic viruses were newly designed and applied to gene amplification for cloning and detection. Capsid protein gene of black queen cell virus (BQCV) among examined viral genes was only successfully amplified from collected bumble bee adults and sequenced. To optimize the detection of capsid protein gene of BQCV, 4 regions in the capsid protein gene were selected and further analyzed in quantitative real-time PCR (qRT-PCR). The qRT-PCR analysis revealed that capsid protein gene was directly detected with not more than 200 ng total RNA. This result suggests that an optimized detection via qRT-PCR can be applied for the rapid and sensitive diagnosis of BQCV infection in the field population as well as risk assessment of B. terrestris.

Bumblebees are important pollinators of crops and wildflowers. Bumblebees generally produce one generation per year. One of the key stages for year-round rearing of bumblebees is breaking diapause. To evaluate the effects of a combination method of CO2-narcosis and cold treatment to break the diapause of B. ignitus and B. terrestris queens, we determined whether this method affected their ability to establish a colony after the diapause break. The diapause treatment regimes that were utilized were CO2(CO2– narcosis), CT-1M(cold treatment at 5°C for 1 month), CT-1M-CO2(CO2–narcosis after cold treatment for 1 month), CT-2M-CO2(CO2–narcosis after cold treatment for 2 months), CT-2M (cold treatment for 2 months), CT-2.5M-CO2(CO2–narcosis after cold treatment for 2.5 months) and CT-2.5M (cold treatment at 5°C for 2.5 months). In view of the effects on the colony developmental characteristics of B. ignitus queens, the most favorable diapause treatment was CT-1M-CO2. A combination method of CO2– narcosis and cold temperature treatment yielded better results than that of single CO2–narcosis or cold temperature treatment on the colony development of diapause-broken B. ignitus queens. In the case of B. terrestris queens, we concluded that a combination method of CO2 and cold temperature treatment yielded better results than that of a single cold-temperature(up to2 months) treatment. In conclusion, the findings of the present study indicated that the combined application of CO2 and cold temperature was a favorable method for the colony development of diapause-broken B. ignitus and B. terrestris queens compared with only CO2–narcosisorcoldtemperaturetreatments. A combination method of CO2 and cold treatment reduced the side effect of CO2–narcosis and shortened the duration of cold treatment by at least 1month.

비벡터링(Bee-vectoring)은 벌을 이용하여 작물을 보호하는 기술이다. 꿀벌이나 뒤영벌(Bombus terrestris)이 모여 있는 벌통의 내부 또는 외부에 미생물제제를 담 은 분배장치를 설치하고 외부로 빠져나갈 때 미생물제제가 들어있는 분배장치를 통과하여 나가게 함으로써 수분 활동 시에 작물의 꽃이나 잎 부위에 미생물 제제를 퍼트려 병해충 방제에 이용할 수 있는 기술이다. 이 기술의 장점은 화분매개곤충들 의 방화활동을 이용하여 병해충 방제가 필요한 꽃이나 잎에 정확히 미생물 제제를 운반함으로써, 수분활동에 의한 농작물 수량증대 및 품질향상의 이점과 더불어 병 해충의 효과적인 방제를 할 수 있다는 점이다. 본 연구에서는 미생물제제를 효과적 으로 뒤영벌의 몸에 묻힐 수 있는 분배장치를 자체 제작하고 분배장치 유무에 따른 뒤엉벌의 시간 당 출입 비율을 비교하였다. 출입 비율은 뒤영벌이 가장 많이 활동하 는 오전 6시부터 오후 10시까지의 행동을 조사하였다. 분배장치와 미생물제제인 토박이(비티아이자와이엔타 423 수화제, 동부하이텍)를 설치하였을 경우 뒤영벌 은 시간당 평균 18.8회 나오고 평균 18회 들어가는 행동을 보였다. 분배장치만 설치 하였을 경우, 평균 21.6회와 24.2회의 출입 행동을 보였으나 분배장치를 설치하지 않은 경우에는 시간 당 평균 35.2회와 43.4회 출입 행동을 하였다.

Recently, the flies are used to pollinate onion for seed production in Korea. However, the use of flies for pollination has caused complaints by local residents in onion seed production owing to sanitation issues and smell. Therefore, we studied the effects of pollination with bumblebee (Bombus terrestris) and flies(e.g., houseflies, flesh flies, blue bottle flies and blowflies) at small net(1.8m×2.3m×1.8m) in the greenhouse(66m2) for onion seed production. Although the number of flies was about 70 times than number of bumblebee workers, flies and bumblebees showed equivalent pollination efficiency. Therefore, pollination by B. terrestris can be used as an alternative approach in seed production of onion. An analysis of the most appropriate colony of B. terrestris for seed production of onion in small net showed that colony composed of fifty to eighty workers per small net was the most efficient level. When early maturing onion cultivars ‘Salad-5’ and ‘Yeonsinwhang’, and mid-late maturing onion cultivar ‘Colossus’ were pollinated by B. terrestris and files in small net, B. terrestris showed rather efficient pollination efficiency than flies or was equivalent to flies. In addition, difference among onion cultivars was affecting pollination effect by flies but was not affecting pollination effect by B.terrestris.

Bumblebee, Bombus terrestris is widely used to pollinate various crops, especially tomato, in greenhouses. Mason bee, Osmia cornifrons, which is using as apple pollinator. Here we investigated characteristics on pollinating activity of B. terrestris and O. conifrons at different weather conditions in apple orchard. The foraging activity of O. conifrons was more sensitive than that of B. terrestris at weather conditions including temperature, illumination intensity and wind velocity. The pollinating activity of O. conifrons was very low than that of B. terrestris at a low temperature, cloudy, rainy and windy weather. Therefore, Our results indicated that B. terrestris was suitable for apple pollinator than O. conifrons, especially in low temperature and bad weather.

Bumblebee, Bombus terrestris is widely used to pollinate various crops, especially tomato, in greenhouses. Mason bee, Osmia cornifrons, which is using as apple pollinator. Here we investigated characteristics on pollinating activity of B. terrestris and O. conifrons at different weather conditions in apple orchard. The foraging activity of O. conifrons was more sensitive than that of B. terrestris at weather conditions including temperature, illumination intensity and wind velocity. The pollinating activity of O. conifrons was very low than that of B. terrestris at a low temperature, cloudy, rainy and windy weather. Therefore, Our results indicated that B. terrestris was suitable for apple pollinator than O. conifrons, especially in low temperature and bad weather.

Bumblebee, Bombus terrestris is widely used to pollinate various crops, especially tomato, in greenhouses. Mason bee, Osmia cornifrons, which is using as apple pollinator. Here we investigated characteristics on pollinating activity of B. terrestris and O. conifrons at different weather conditions in apple orchard. The foraging activity of O. conifrons was more sensitive than that of B. terrestris at weather conditions including temperature, illumination intensity and wind velocity. The pollinating activity of O. conifrons was very low than that of B. terrestris at a low temperature, cloudy, rainy and windy weather. Therefore, Our results indicated that B. terrestris was suitable for apple pollinator than O. conifrons, especially in low temperature and bad weather.

Bumblebees are widely used to pollinate various greenhouse crops. Among the different bumblebee species, Bombus ignitus is indigenous to Korea, China, Japan and Russia. B. ignitus undergoes one generation per year, and artificial hibernation is essential for year-round rearing of the bumblebee. Keeping the queens under low-temperature conditions for several months is an effective method for terminating their diapause and promoting colony development. In the present study, we investigated how cold temperature affects the artificial hibernation of B. ignitus queens. Under chilling temperatures of -2.5°C, 0°C, 2.5°C and 5°C with constant humidity >80%, the queens stored at 2.5°C exhibited the highest survival rates, which were 74.0% at one month, 67.0% at two months, 60.0% at three months, 46.0% at 4 months, 33.0% at 5 months and 24.0% at 6 months. Lower survival rates were observed at 0°C, 5°C, 7.5°C and 12.5°C. At 2.5°C the colony developmental characteristics after diapause were 1.2- to 1.5-fold greater than those when queens were stored at 5°C. Thus, 2.5°C and 70% R.H. were the most favorable chilling temperature and humidity conditions for terminating the diapause of B. ignitus queens.

Bumblebee venom serine protease inhibitors have been shown to inhibit plasmin activity. In this study, a bumblebee (Bombus ignitus) venom serine protease inhibitor (BiVSPI) that acts as an antimicrobial factor was identified. BiVSPI is a 55-amino acid mature peptide with ten conserved cysteine residues and a P1 methionine residue. BiVSPI was expressed in the venom gland and was present as an 8-kDa peptide in venom. Recombinant BiVSPI expressed in baculovirusinfected insect cells exhibited inhibitory activity against chymotrypsin, but not trypsin. BiVSPI also exhibited inhibitory activity against microbial serine proteases, such as subtilisin A (Ki 6.57 nM) and proteinase K (Ki 7.11 nM), indicating that BiVSPI acts as a microbial serine protease inhibitor. In addition, BiVSPI was also shown to bind directly to Bacillus subtilis, B. thuringiensis, and Beauveria bassiana, but not to Escherichia coli. Consistent with these results, BiVSPI exhibited antimicrobial activity against Gram-positive bacteria and fungi. These findings provide novel evidence for the antimicrobial function of this bumblebee venom serine protease inhibitor.

The bumblebee, Bombus terrestris, has played an important role as one of the alternative pollinators since the outbreak of honeybee collapse disorder. Recently, pathogens and parasites such as viruses, bacteria and mites, which affect the life span and fecundity of their host, have been discovered in B. terristris. In order to detect the microsporidian pathogen, Nosema Spp. in the field populations of B. terristris, we collected adults and isolated their genomic DNA for diagnostic PCR. The PCR primers specific for Nosema Spp. were newly designed and applied to gene amplification for cloning. Only small subunit ribosomal RNA(SSU rRNA) gene of N. ceranae was successfully amplified and sequenced among examined genes, which indicates that N. ceranae mainly infects the examined field population of B. terristris. To detect of SSU rRNA gene, two regions of SSU rRNA gene were selected by primary PCR analysis and further analyzed in quantitative real-time PCR(qRT-PCR). The qRT-PCR analysis demonstrated that SSU rRNA of N. ceranae was detected at concentrations as low as 0.85 ng/μl genomic DNA. This result suggests that the detection via qRT-PCR can be applied for the rapid and sensitive diagnosis of N. ceranae infection in the field population as well as risk assessment of B. terristris.

The bumblebee, Bombus ignitus (Hymenoptera: Apidae), is a valuable natural resource that is widely utilized for greenhouse pollination in South Korea. Understanding the magnitude of genetic diversity and geographic relationships is of fundamental importance for long term preservation and utilization. As a first step, we sequenced a partial COI gene of mitochondrial DNA (mtDNA) corresponding to the “DNA barcode” region and the complete internal transcribed spacer 2 (ITS2) of nuclear ribosomal DNA from 88 individuals collected in nine South Korean localities. The complete ITS2 sequences were longest among known insects, ranging in size from 2,034 bp ~ 2,052 bp, harboring two duplicated 112-bp long repeats. The 658-bp long mtDNA sequences provided only six haplotypes with a maximum sequence divergence of 0.61% (4 bp), whereas the ITS sequences provided 84 sequence types with a maximum sequence divergence of 1.02% (21 sites). The combination of the current COI data with those of published data suggest that the B. ignitus in South Korea and China are genetically a large group, but those in Japan can be roughly separated into another group. Overall, a very high per generation migration ratio, a very low level of genetic fixation, and no discernable hierarchical population were found to exist among the South Korean populations of B. ignitus, which suggests panmixia. This finding is consistent with our understanding of the dispersal capability of the species.

To substitute for bluebottle fly, Chrysomyia megacephala which is being used for pollinator in mango fruit, and improve the pollinating effect of mango fruit which is also being increased as high value added crop recently in Jeju island of korea, 2 kinds of pollinator were used in analyzing and surveying of foraging activities on mango fruit in Seogwipo province. This study was conducted using 3 species of pollinator, Apis mellifera, Bombus terrestris and Chrysomyia megacephala with 3 treatment in vinyl-house condition respectively. Species of mango fruit, Irwin, was used in this experiment. A number of foraging activity of Apis mellifera and Bombus terrestris in hive showed highest 11 AM, and showed normal foraging activity in high temperature condition(28℃). Pollinating ratio of Bombus terrestris was shown 100% and over 95% in case of Apis mellifera. This ratio suggest that the 2 species of insects is effective as pollinator on mango fruit compared with bluebottle fly. Daily pollinating activity of Apis mellifera and Bombus terrestris was shown peak in 11 AM, but showed even activity from 9 AM to 3 PM in case of Chrysomyia megacephala. The pollinating characteristics of 3 species depends on illuminance but temperature, especially in case of Bombus terrestris was more affected by change of illuminance. Visiting time of Bombus terrestris and Apis mellifera on this flower was shown 2.8 and 3.4 seconds respectively. But Chrysomyia megacephala showed longer 10 times with 32.5 seconds than other insects. This results suppose that Chrysomyia megacephala showed as resting behavior for almost time on the flower not foraging activity to pollinate.

Many bumblebee species have declined in number in recent decades, particularly in developing regions. Widespread declines of bumblebee species threaten the pollination levels of both wildflowers and crops. Here, we investigated the body weight and colony-developmental characteristics of Korean native bumblebee (B. ignitus) queens collected from 2000 to 2010 for conservation of native bumblebees for breeding. The average weight of 6,852 queens was 0.77 ± 0.44 g. The weight of B. ignitus queens collected in 2005 was the greatest, 0.87 ± 0.12 g, which was 1.0–1.3-fold heavier than any other year. The average oviposition rate was 81.6 ± 10.7%, and 2004 showed the highest rate, 95.0%. This value corresponded to 1.1-1.6-fold increases over the queens collected in the other years. The average rate of colony foundation was 60.9 ± 11.0%. Queens in 2008 exhibited the best performance, 75.4%, which was 1.0-1.9-fold higher than the other years. The rate of progeny-queen production averaged 27.0 ± 9.4% (Fig. 5) and peaked in 2001 at 43.2%; this value was 1.1–4.7-fold higher than other years. The average number of queens produced and number of generations begotten by queens was 27.6 ± 10.1% and 4.8 ± 2.0, respectively. Queens in 2000 averaged 9 generations of offspring, which was 1.1-3.1-fold greater than other years. These results indicate that the colony-developmental characteristics of the collected queens changed significantly between 2000 and 2010. In addition, there was no correlation between body weight and number of queens collected, although body weight was affected by collection year. Since 2008, the colony- developmental characteristics of queens have worsened.

Currently, honeybee colonies are not stable and suffer from the infection of pathogens, affecting the pollination. For the alternatives to this difficulty, Bombus terrestris has been imported and used for pollination in agricultural fields. Although imported insects for pollination are very useful, the potential risk exposing to novel pathogens has been raised. To assess the risk primarily, we designed and synthesized PCR primers for detection of pathogens and parasites in B. terrestris. The samples were obtained from companies importing B. terrestris or field collections and genomic DNAs not showing physical shearing were purified. PCR for detection of pathogen- or parasite-specific gene revealed several DNA fragments were amplified in expected molecular size including Kashmir Bee Virus, Varroa jacobsoni, V. rindereri, Acarapis woodi and Aspergillus flavus. These amplified DNA fragments are in the process of cloning for DNA sequencing to confirm the target gene amplification. We also have plans to optimize the PCR conditions for each amplified target gene and try to develop biomarkers for diagnosis.