Vertical transmission of symbiont is known as the most effective way to deliver their offspring. However, it has beenreported that the bean bug, Riptortus pedestris, acquires its gut symbiont, Burkholderia sp., from its environment (e.g.soil) during the 2nd instar nymphal stage. Nevertheless, it is not unknown how they locate their symbiont. For this reason,dual-choice experiments were conducted in both solitary (a nymph) and group conditions (100 nymphs) to investigatewhether or not R. pedestris has preference on Burkholderia sp. treated or untreated solution in laboratory conditions. Inthese experiments, there was no significant difference in the preference between the two treatments. We are testing inthe Y-tube olfactometer system to evaluate the response of R. pedestris on possible chemical cues from Burkholderia sp..

The bean bug, Riptortus pedestris, is a pest of leguminous crops. Recent studies have revealed an interesting relationship between R. pedestris and a gut symbiont Burkholderia sp.. R. pedestris mainly acquires Burkholderia sp. during 2nd instar period, which leads to enhancement of growth and reproductive rate of the host. Nevertheless, how R. pedestris acquires its symbiont is not yet clearly described. Therefore, we investigated whether 2nd instar nymphs have preference on Burkholderia sp. compared to untreated control in laboratory settings. In the choice test, no significant difference was found in the preference by R. pedestris between the two treatments. To further investigate single individual’s behaviors (its first choice, approaching frequency to each treatment, and residence time on the treatment). We video-recorded single individuals in the choice setting for 12 hours. Again, there was no significant difference in any of the behavioral traits recorded. Further choice tests will be conducted in the Y-tube olfactometer systems and semi-field conditions.

The soybean bug, Riptortus pedestris, is a polyphagous species and an important pest of soybean. We investigated the development periods and survivorship of R. pedestris egg and nymph stage. The study was conducted at eleven constant temperatures of 12, 14, 16, 18, 20, 24, 28, 32, 34, 35, and 36℃, respectively. Egg could not hatch under 14℃. Nymph could not develop to adult at 16℃. The lower developmental threshold (LDT) and thermal constant (K) of R. pedestris nymph stage were estimated by linear regression. Higher temperature threshold (TH) and lower temperature threshold (TL) were calculated by two nonlinear functions (Performance and SSI model). The adult emergence frequency of R. pedestris over full range of constant temperatures was simulated using developmental rate functions and Weibull function.

Riptortus pedestris is a polyphagous species and an important pest of soybean. We investigated the phenology of R. pedestris at 14 locations - Jeonju, Naju, Goheung, Namhae, Milyang, Gunwi, Andong and 7 areas of Jeju. Commercial pheromone monitoring traps were used. The traps were baited with lures containing a 1:5:1 ratio of myristyl isobutyrate, (E)-2-hexenyl (E)-2 hexenoate and (E)-2-hexenyl (Z)-3-hexenoate. There was difference the first adult catch and nymph emergence among locations. To develop spring emergence distribution pattern, we used the trap data from each location. Spring emergence by sampling date was cumulated and analyzed relative to the Julian date using Weibull distribution function.

R. pedestris의 중장인 M4에 Burkholderia 균이 공생하고 있으며, 이 공생균은 숙주의 면역에 이로운 영향을 준다. 본 연구실에서 외부 병원체의 침입에 따른 면역 반응으로 항균활성을 가지는 3 종류의 항균 펩타이드를 정제하였다. 그 중에서 Rip-thanatin은 18개의 아미노산으로 이루어져 있으며, 2개의 시스테인에 이황화 결합이 있는 펩타이드로, R. pedestris에 많은 수의 유도체가 존재하고 있다. 이 Rip-thanatin은 병원체를 숙주에 주사하였을 때 공생균을 가지는 곤충의 M4 중장에서 공생균을 가지지 않는 곤충보다 유전자 발현양이 유의적으로 높아지는 사실을 관찰하였다. 따라서, Rip-thanatin이 중장의 면역반응에서 중요한 역할을 하며, 이는 공생균의 유무에 의해 영향을 받는 것으로 규명되었다. 본 연구진은, 숙주의 면역반응에 있어서 Rip-thanatin의 중요성을 규명하기 위해, 다양한 병원체에 대한 Rip-thanatin의 유전자 발현 정도를 관찰하고, 여러 가지 Rip-thanatin의 유도체를 합성하여 항균 활성과 Rip-thanatin의 구조적 상관관계를 규명하고자 한다.

In order to establish symbiotic host-bacterial relationships, symbionts in insects evolved a mechanism to overcome host immune responses. Here we provide the resistance of symbiotic bacteria on the insect immune system. As a result, through the transposon mutagenesis, we found a salivary gland (SG) susceptible mutant. The disrupted gene was identified as nlpB involved in lipoprotein synthesis. The nlpB, bla double deletion mutant was sensitive to SG like nlpB-Tn5 inserted mutant. This mutant increases outer membrane permeability. It provides an explanation for SG susceptibility, because the antimicrobial peptide in SG would be able to translocate across the outer membrane more easily than in the wild type. These results indicate that nlpB and bla are likely to be important factors in terms of determining resistance against SG of Riptortus that is connected with the successful colonization of the Riptortus midgut.

곤충병원균인 Pseudomonas entomophila를 Riptortus pedestris의 체내에 주사하였을 때, 곤충을 사멸시키는 효과를 나타내는 물질을 찾기 위하여, P. entomophila를 배양한 배양액을 이온교환 크로마토그래피 및 크기배제 크로마토그래 피를 통해 살충 효과를 나타내는 물질을 정제 하였다. 그 결과 R. pedestris에 대해 살충효과를 보이는 독성 물질은 metalloproteinase의 일종인 AprA 단백질인 것으로 나타났다. 정제 된 AprA를 R. pedestris에 주사하였을 때, 곤충이 매우 빠른 속도로 사멸하는 양상을 보였다. AprA 단백질이 어떠한 작용 기전으로 R. pedestris에 독성 효과를 보이는지 알기 위하여, 정제 된 AprA를 R. pedestris의 체내로 주사한 후 hemocyte (혈구)의 숫자를 조사한 결과, AprA를 주사 한 직후에는 혈구의 숫자가 증가하였으나, 1시간 이후부터는 혈구의 수가 급격히 감소하기 시작하였고, 결국은 모든 곤충이 사멸하였다. 본 연구진은 AprA가 R. pedestris의 면역반응을 어떻게 억제하는지 알기 위해, AprA가 R. pedestris의 혈구를 제거하는 기작을 보고하고자 한다.

An entomopathogenic fungus, Beauveria bassiana ARP14, was isolated from field collected Riptortus pedestris (F.). Lethal concentration of ARP14 was compared with commercialized GHA strain against R. pedestris and its egg parasitoids Ooencyrtus nezarae Ishii and Gryon japonicum (Ashmead). Mortality and infection rate were evaluated in five concentrations, i.e., 1×10 9 , 1×10 8 , 1×10 7 , 1×10 6 , and 1×10 5 conidia/mL using glass scintillation vial. The LC50 of 2nd, 4th instar, and adult stage of R. pedestris was not different between the two strains. The infection rate of ARP14 was higher than that of GHA in 4th instar and adult stage in 1×10 8 conidia/mL. The infection rate was low in ARP14 than GHA in both egg parasitoids. Therefore, our results suggest that the ARP14 strain can be used against R. pedestris with less negative effect on egg parasitoids than GHA strain.

Riptortus pedestris (Fabricius) is one of major soybean pests in Korea. Its feeding during pod formation period significantly reduces the yield of soybean. Management tactics commonly used include insecticide such as etofenprox and aggregation pheromone trap, but they have been often found ineffective. As an alternative methods, parasitoids such as Gryon japonicum (Ashmead) and Ooencyrtus nezarae Ishii could be conserved by directly releasing non-viable host eggs in the field, by incorporating the non-viable host eggs into aggregation pheromone trap, by planting a trap crop of natural enemies, or by combining selective insecticides with a trap crop of natural enemies. I present study results of these tactics with discussion here.

Many insects possess symbiotic microorganisms in their bodies, wherein host-symbiont intimate interactions occur. Despite recent advances in omics technology, the molecular bases of the symbiotic associations remain unclear. The bean bug Riptortus pedestris harbors an environmentally acquired Burkholderia symbiont in their midgut crypts. Unlike other insect symbionts, the Burkholderia symbiont is easily culturable and genetically manipulatable outside the host insect. In conjunction with the experimental advantages of the bean bug, the Riptortus–Burkholderia association is an ideal model system for approaching the molecular bases underpinning insect-microbe symbioses. In this presentation, the current knowledge of this model system is summarized.

The bean bug, Riptortus pedestris (Hemiptera: Alydidae), is one of the major pests on leguminous crops especially in Korea and Japan. Because of its economic importance as crop pest, A large proportion of research on this species have focused on applied questions related to pest management tactics. In addition to the applied research, recent studies have also revealed an interesting relationship between R. pedestris and a gut symbiont Burkholderia sp. as an ideal model system for insect-microbe symbiotic interaction. Under this circumstance, our research group initiated a series of research projects to better understand behavioral ecology of R. pedestris and address its implications for both basic and applied questions. This presentation will cover various behavioral studies with R. pedestris including sublethal effects of insecticides, harmonic radar tracking, overwintering ecology, and symbiont acquisition mechanisms

Recent studies suggested that gut symbionts modulate insect development and reproduction. However, how gut symbionts modulate host physiologies and what types of molecules are involved in these changes are still unclear. When we analyzed hemolymph proteins and transcriptional levels of host insects, hexamerin-α (Hex-α), hexamerin-β (Hex-β) and vitellogenin-1 (Vg-1) were highly expressed in symbiotic insects (Sym) compared to aposymbiotic insects (Apo). Depletion of Hex-β by RNA interference in 2nd Sym-nymphs delayed adult emergence, whereas Hex-α and Vg-1 RNA interference in 5th nymphs decreased reproduction of female insects and caused loss of color of laid eggs. Also, the levels of JHSBIII of Riptortus host were 3-fold higher in the Sym-female insects compared to the Apo-insects. These results demonstrate that the Burkholderia gut symbiont modulates host development and egg production through regulating the expression of three host storage proteins by controlling of brain hormone.

Five phaP family genes and one phaR gene have been identified in the genome of Burkholderia gut symbiont. PhaP proteins function as surface proteins of polyhydroxyalkanoate (PHA) granules, and PhaR protein acts as a negative regulator of PhaP biosynthesis. To address the biological roles of four phaP family genes (phaP1, phaP2, phaP3, and phaP4) and the phaR gene during insect-gut symbiont interaction, these Burkholderia mutants were fed to the second instar nymph. The ΔphaR mutant decreased the colonization ability in the host midgut compared to wild-type Burkholderia cells and negatively affected the host insect’s fitness compared with wild-type infected host. These results demonstrate that PhaR plays an important role in the biosynthesis of PHA granules and it is significantly related to the colonization of the Burkholderia gut symbiont in the host insects’ midgut

Morphology of antennal sensilla and their distribution were investigated in male and female adults of Ooencyrtus nezarae, an egg parasitoid of Riptortus pedestris, using scanning electron microscopy. Antennae of O. nezarae was composed of scape, pedicel and seven flagella in both sexes. Six types of sensilla (s. trichodea, s. basiconica, s. chaetica, s. campaniformia, s. sickle-shaped and unknown s.) were identified from both sexes. Among them, s. trichodea and s. sickle-shaped were multiporous, others are not. They distributed in varying numbers. Sexual dimorphism was clearly observed in the distribution of s. trichodea (only on male antennae) and unknown sensilla (only on female antennae). These findings would be helpful for further studies on detailed sex specific-receptive functions of each antennal sensilla.

단감 해충의 효과적인 방제를 위하여, 현재까지 주로 화학 살충제가 사용되어 왔으나, 최근 화학 살충제의 과다 사용은 해충 저항성과 농약 잔류 문제라는 심각한 문제를 야기하고 있다. 본 연구에서는 친환경 유기농자재 (Eco-Friendly Organic Pesticides; EFOP)의 주요 단감 해충 인 감꼭지나방과 톱다리개미허리노린재에 대한 살충 활성을 검토하였다. 현재 유통되고 있는 친환경 유기농자재 11 종을 실험실 조건에서 대상 해충에 분무 처리하였다. 대조 약제는 화학 살충제인 buprofezin+dinotefuran (20+15) 수화제를 사용하였다. 감꼭지나방에 대한 유기농자재의 살충 효과를 평가하는 과정에서, 감꼭지나방 유충의 개체수 부족 문제로 인하여 배추좀나방 유충을 1차 실험충으로 사용하였으며, 세 종류의 친 환경 유기농자재를 사전 선발하였다. 선발된 친환경 유기농자재는 목초액 (EFOP-1), 회화나무, 양명아주, 멀구슬나무의 혼합추출물 (EFOP-2), Bacillus thuringiensis subsp. aizawai NT0423 (EFOP-11)이었으며, 선발된 유기농자재의 감꼭지나방 유충에 대한 살충력 실험 결과, EFOP-2 의 처리구에서 생충율이 27.7% (5 일차), 13.3% (7 일차), 6.7% (10 일차)로 가장 높은 살충 효과를 보였다. 톱다리개미허리노린재에 대한 살충 효과 평가에서는 회화나무, 양명아주, 멀구슬나무의 혼합추출물 (EFOP-2, EFOP-9)과 고삼, 데리스의 혼합추출물 (EFOP-10)이 높은 살충 효 과를 보였다. 특히 EFOP-2의 처리구에서 생충율이 20.0% (5 일차), 16.7% (10 일차)로 가장 높은 살충 효과를 보였다. 결론적으로, 친환경 유기 농자재인 EFOP-

Chemical pesticides have been used to control persimmon pests, however the overuse of the pesticides caused insect resistance. Herein we investigate the potential of eco-friendly organic pesticides (EFOP) on the control persimmon pests, Stathmopoda masinissa and Riptortus pedestris. Eleven commercially available EFOP were sprayed on the target insects in laboratory conditions. The chemical pesticide, buprofezin+dinotefuran wettable powder served as a positive control. In the bioassay against persimmon fruit moth, the mixture of Chinese scholar tree, goosefoot and subtripinnata extracts (EFOP-2) treatment showed the highest control efficacy: 27.7% (5 days) of survival rates. In the bioassay against bean bugs, the EFOP-2 showed the highest control efficacy: 20.0% (5 days) of survival rates. These results suggest that the mixture of three plants extracts (EFOP-2) has high and multiple potential in the management of the persimmon pests.

A viral genome was assembled de novo from next-generation sequencing (NGS) data from bean bugs, Riptortus pedestris, infected with an entomopathogenic fungus, Beauveria bassiana (Bb), and was further confirmed via the RACE method. Based on the phylogenetic analysis of the RdRp sequences, RiPV-1 was clustered in the unassigned insect RNA viruses with two other viruses, APV and KFV. These three viruses were suggested to constitute a new group of insect RNA viruses. Interestingly, RiPV-1 replication was increased dramatically in bean bugs 2 to 6 days after fungal infection. In conclusion, a novel insect RNA virus was found by NGS data assembly. This virus can provide further insight into the interaction between virus, fungus and the host.

It is not uncommon in the nature for insects to have symbiotic relationships with microorganisms. The bean bug, Riptortus pedestris, which harbors symbiont Burkholderia sp., is a serious agricultural pest in South Korea and Japan. It is reported that the symbiont has positive effects on the fitness of R. pedestris. However, virtually no information is available for how this insect finds the symbiont in the nature and how the behavior of R. pedestris is affected by the symbiont. To investigate the symbiont acquisition mechanism, the frequencies of R. pedestris approaching to symbiont or untreated control treatment were compared in a plastic cage (465×290×260mm) (n=20). In the experiment, there was no detectable difference between the two treatments. Further study using video tracking systems will help us to better understand detailed behaviors for symbiont acquisition.

Climate change can affect major changes to the dynamics of individual insect. The increase of surface temperature would cause multivoltine species to increase the number of generations (i.e. voltinism) per year. Determining the voltinism was based on the climate data and biological parameters. The soybean bug, Riptortus pedestris (Fabricius) (Hemiptera: Alydidae) is one of major pests of soybean and has multi host plants. To estimate the voltinism of R. pedestris, the climate data were obtained from the web site of the Korea Meterological Administration(KMA) from 1966 or 1973 to 2015 at 77 locations, respectively. We used the lower developmental threshold (15.0°C) and thermal constant (312.5DD) for R. pedestris from egg to adult emergence. Degree-days were calculated by using simple cutoff method and cumulating degree-days (CDD) were obtained by summing calculated degree-days. The voltinism of R. pedestris was calculated by dividing CDD per specific period by the thermal constant requirement of R. pedestris development from egg to adult emergence. The number of generations varied depending on the season, with the lowest in February and the highest in August at each location. At least one generation per month would be possible from July to August. The average voltinism from 1966 to 2015 was highest at Seoguipo, and was followed by Daegu, Jejusi, Gwangju and Jeonju. The average voltinism from 1966 to 2015 was lowest at Ulreundo. The voltinism of R. pedestris observed the last 50 years was attributable to human activities at some locations.

The bean bug, Riptortus pedestris (Hemiptera: Alydidae), is a major pest of soybean and tree fruit in Korea. To protect crops, chemical insecticides have been widely used. However, the effects of insecticides is not only reduce the pest population but also change of pest behavior. In this study, we tested the effects of two insecticides, etofenprox (pyrethroid) and acetamiprid (neonicotinoid), on the behavior of R. pedestris. Two insecticides were diluted with water based on the conventional spray dose. After exposed to insecticide residue, adult R. pedestris was put individually into a clean petri dish with soybean. The behavior of R. pedestris was video-recorded for 16 hours. The frequencies of approaching to food and feeding were compared to the untreated control group. R. pedestris exposed to etofenprox made more frequent approaching to the food, but those exposed to acetamiprid made less frequent approaching compared to the control group. When compared to the control group, the feeding frequency of acetamiprid-exposed group was significantly reduced but etofenprox-exposed group showed more frequent feeding activity. In both treatments, the feeding probability, calculated by feeding frequency divided by approaching frequency, was reduced compared to the control. We conclude that acetamiprid may protect crops by reducing the frequency of R. pedestris to approach and feed on crops.