We analyzed a portion of mitochondrial COI gene sequences (658 bp) to investigate the genetic diversity and geographic variation of the swallowtail butterfly, Papilioxuthus L., and the cabbage butterfly, Pieris rapae (Lepidoptera: Papilionidae). P. xuthus showed a moderate level of sequence divergence (0.91% at maximum) in 15 haplotypes, whereas P. rapae showed a moderate to high level of sequence divergence (1.67% at maximum) in 30 haplotypes, compared with other relevant studies. Analyses of population genetic structure showed that most populations are not genetically differentiated in both species. The distribution pattern of both species appears to be consistent with category IV of the phylogeographic pattern sensu Avise (Avise et al. 1987): a phylogenetic continuity, an absence of regional isolation of mtDNA clones, and extensive distribution of close clones. The observed pattern of genetic diversity and geographic variation of the two butterfly species seems to reflect the abundant habitats, abundant host plants, and flying abilities in connection with the lack of historical biogeographic barriers.
In order to establish a year round indoor-rearing system for Copper butterflies; large copper butterfly, Lycaena dispar, the effect of temperature on larval development and diapause was investigated. Temperature has been suggested as an important factor regulating the developmental rate, length of life, and survival rate from insect. As temperature increased, the developmental period was gradually shorten. The developmental periods of large copper larvae had a range of 11.0 days to 28.5 days at 30℃ and 17.5℃ respectively. The highest emergence rate was 94.2% recorded at 20℃. And the low emergence rate was 72.7% under 17.5℃. We investgated the sensitive stages to diapause induction in the larger copper effect of temperature and photoperiod. The experiment involves transfer of individuals from diapause averting (LD 16:8h, 25℃) to diapause inducing condition (LD 8:16h, 20℃) at various stages. Diapause was induced in 95.2% insect transferred at hatching larvae, in 15.6% of insects transferred at 2nd stadium molt, in 0% of insects transferred at after 3rd stadium molt. Percentage diapause induction increase with the length of short days and low temperature experienced. The main stage sensitive to photoperiod and temperature induction of diapause determination is the early first larval instar. The diapause induction began 14 days and ended 20 days after hatching larvae (LD 8:16h, 20℃).
Cytochrome P450s (P450s) are known to oxidize a variety of insecticides including pyrethroids, thereby conferring metabolic resistance in diamondback moth (DBM), Plutella xylostella. Synergism assay with piperonyl butoxide indicated that the enhanced activity of P450 is associated with pyrethroid resistance in a cypermethrin-resistant (CR) strain. However, there were little differences in the basal transcription levels of all the P450s examined between susceptible (Sus) and CR strains, suggesting that constitutive overexpression of P450 is not likely involved in the cypermethrin resistance but induction of P450 by cypermethrin is rather associated with metabolic resistance. To determine the conditions resulting in maximum levels of P450 induction, several factors including the way of adminstration (topical application vs. leaf dipping), exposure dose and exposure duration were examined. In general, leaf dipping method resulted in greater levels of induction in a wider array of P450s. The conditions of ‘low dose (sublethal dose or concentration) and short exposure (less than 3 hr)' to cypermethrin were more efficient in P450 induction than those of ‘high dose (around LD50 or LC50) and long exposure (more than overnight)’, which have been employed in many other studies to date. Cross-strain comparison revealed that 9 of 11 P450s were induced 1.4-2.2 fold in CR whereas only 3 P450s in Sus under the optimal induction conditions, demonstrating that metabolic resistance in CR strain is actually conferred by the mechanism of selective P450 induction when exposed to cypermethrin.
For the physiological study on environmental impact of genetically modified (GM) pepper plant on non-target but three-trophically related insect species, we investigated behavioral responses of Aphis gossypii and Aphidius colemani in Y-tube olfactometer to the cucumber mosaic virus (CMV)-resistant transgenic pepper plant (H15 GM line) expressing coat protein gene of CMV and its wild type pepper plant (untransformed, susceptible to CMV pathotype II, P2377 inbred line) in relation to CMV infection. CMV-infected plants were prepared with the 30 min of inoculation by the winged A. gossypii viruliferous or mechanical inoculation using CMV-Fny, and with molecular diagnosis using reverse transcriptase-polymerase chain reaction (RT-PCR) over 2 weeks after inoculation. In this study, time for attraction responses (attraction time) of A. gossypii were not significantly different in the pepper strain, and the virus infection of plant. However, the attraction time of A. colemani was significantly different between the GM plant and the non-GM plant. In addition, the attraction time of A. colemani to the GM plant was significantly decreased according to the CMV infection. For further study, the volatile organic compounds (VOCs) emitted by these plants were collected with an entrainment kit and analyzed by Gas Chromatography (GC) on HP-1 column. The specific VOCs related to CMV infection were detected in the GM plant over 4 weeks after inoculation of CMV in this study. Thus, it is suggested that VOCs of the GM plant in this study may be produced more as a signal attracting A. colemani in relation to CMV infection.
This study was carried out to investigate the effect of each single component of the synthetic sex pheromone in the rice green caterpillar, Naranga aenescens and rice leaf roller, Cnaphalocrocis medinalis. Trap catches with each single component or binary mixtures of the sex pheromone in N. aenescens were significantly lower than that with an optimum mixture (2: 1: 4) of Z9-14:Ac, Z9-16:Ac and Z11-16:Ac. But only few number of C. medinalis male was caught in trap baited with each component of the sex pheromone excepting Z13-18:Ald. In an optimum composition of the sex pheromone in C. medinalis, trap catches baited with an 11: 100: 11 mixture of Z11-18:Ald, Z13-18:Ald, and Z13-18:Ac without two alcohol components (Z11-18:OH and Z13-18:OH) was higher than that baited with an optimum mixture of Z11-18:Ald, Z13-18:Ald, Z11-18:OH, Z13-18:OH, and Z13-18:Ac. The efficacy of the sex pheromone lure of N. aenescens kept over 4 weeks. The occurrence of N. aenescens caught in trap baited with sex pheromone gave three peaks and C. medinalis showed two distinct peaks throughout the flight season. Therefore, it can be expected that pheromone-baited trap provides a simple tool for monitoring N. aenescens and C. medinalis in Korea.
We tested differences in female pheromone production and male response in three species of the genus Adoxophyes in Korea. Females of all three species produced mixtures of Z9-14:OAc and Z11-14:OAc as major components but in quite different ratios. The ratio of Z9-14:OAc and Z11-14:OAc in pheromone gland extracts was estimated to be 100:198 for A. honmai, 100:23 for A. orana, and 100:3880 for A. sp. Field tests showed that males of each species were preferentially attracted to the two-component blends of Z9-14:OAc and Z11-14:OAc mimicking the blends found in pheromone gland extracts of conspecific females. The effects of minor components identified in gland extracts on trap catches varied with species. Addition of 10me-12:OAc or E11-14:OAc to the binary blend of Z9-14:OAc and Z11-14:OAc significantly increased captures of A. honmai males, whereas E11-14:OAc exhibited a strongly antagonistic effect on catches of A. sp. males. Moreover, Z9-14:OH or Z11-14:OH added to the binary blends increased attraction of male A. orana but not A. honmai and A. sp. males, suggesting that these minor components, in addition to the relative ratios of the two major components, play a noteworthy role in reproductive isolation between Adoxophyes species in the southern and midwestern Korea where these species occur sympatrically.
As an effective generalist predator of aphids and other hemipteran pests, Harmonia axyridis has been a successful biological control agent. Interestingly, it was known that there were varied in color patterns on H. axyridis elytra. In fact, Seo & Youn (2007) reported that H. axyridis had five color patterns, for example, succinea 1, 2, conspicua, spectabilis, and axyridis. But there are uncertain that H. axyridis elytra colour patterns are regulated by genetic polymorphism. So we tried to what is the reason that color patterns are greatly variable. To identify DNA markers linked to a elytra polymorphism, amplified fragment length polymorphism (AFLP) analysis was performed on DNA samples from four female succinea, conspicua, spectabilis and Coccinella septempunctata which is another species in Coccinellidae. AFLP analysis with the restriction endonuclease combination EcoRⅠ and MseⅠwas performed. Using 12 AFLP primer pairs, nine AFLP fragments which is specific between succinea, conspicua, spectabilis was identified. These nine AFLP fragments were isolated, cloned and sequenced. Subsequent UPGMA cluster analysis revealed three major group of H. axyridis populations. These genetic tree showed that H. axyridis elytra colour diversity was affected by genetic polymorphism. For more genetically understanding elytra colour genes, different primer combinations may be need to be generate enough polymorphic markers. These genetic analyses may be facilitate the understanding of molecular mechanism behind wing colour pattern formation.
To isolate plant insecticidal factors, a series of survivorship tests were conducted using artificial diets mixed with seed extract of a mungbean variety (Vigna radiata), which gave a high mortality to the bean bug, Riptortus clavatus. Seed flour was extracted with methanol and ethanol solutions, and then the crude extract was first fractionated with layers of ether, n-butanol and water in solvent/solvent partition. Layers of ether and n-butanol fractions showed insecticidal effects to the bean bug, respectively. The n-butanol fraction was further isolated with several fractions on a reversed-phase column chromatography, and its 100% methanol fraction showed a high insecticidal effect. The fraction was further isolated on a normal-phase column chromatography, and the 70% methanol fraction showed the highest insecticidal effect. The final fraction was analyzed and fractionated on a reversed-phase HPLC. The active isolates were almost insoluble in any organic solvents and did not show any absorption in ultra-violet and visible light wavelength range. On the other hand, the ether fraction of crude extract was isolated on a normal phase column chromatography, and its hexane fraction showed the activity. The hexane fraction was further isolated on a reversed-phase HPLC and tested for the survivorship of the bean bug.
Innate immunity responses are triggered by the immune challenge and therefore involve signaling processes. The cellular response is initiated by hemocytes and mainly involves phagocytosis and encapsulation of intruders by these cells. To address whether Hc-STAT is activated upon bacterial challenge, we examined the subcellular location of STAT protein in hemocyte by immunostaining. A new insect member of the STAT family of transcription factors (Hc-STAT) has been cloned from the lepidopteran, Hyphantria cunea. The domain involved in DNA interaction and the SH2 domain are well conserved. The gene is transcribed at a low level during all stages of development, and the protein is present in hemocytes, fat body, midgut, epidermis, and Malphigian tuble (Mt). Especially, hemocytes and Mt showed transcriptional activation of Hc-STAT upon Gram (-) bacteria and fungal challenge. Gram (-) bacteria and fungal challenge specifically results in nuclear translocation of Hc-STAT protein and induction of DNA-binding activity that recognizes a STAT target site in H. cunea hemocyte. In vitro treatment with pervanadate translocates Hc-STAT to the nucleus in hemocyte cells. Here we report the first evidence for the involvement hemocyte JAK/STAT pathway upon microbial infection in lepidopteran insect.
Reactive oxygen species (ROS) is toxic to living organisms, because its high reactivity causes oxidative damage to proteins, nucleic acids, and lipids. Superoxide dismutase (SOD) is an enzyme facilitating the removal of superoxide anions from living organisms. This study focused on the cloning of MnSOD cDNA from Hyphantria cuneaand its induction upon bacterial infection and various stresses. The open reading frame of MnSOD is composed of 645 bp, encoding 215 amino acid residues. The theoretical molecular mass and pI of putative MnSOD was evaluated to be 24276 Da and 9.14, respectively. The MnSOD from H. cunea is highly similar to human MnSOD (59.5%) as well as Bombyx mori MnSOD (76.2%). MnSOD showed no big induction upon bacterial infection and stresses, compared to that of Cu/ZnSOD.
The larvae of Asian rhinoceros beetle, Trypoxylus [Allomyrina] dichotoma (Coleoptera: Scarabaeidae) feed on decaying organic matter and can be found in compost heaps or old dead logs. Mass rearing of this insect with a sawdust-based diet has made it popular commercial pet in many countries of Asia. Recently, lignocellulose, or biomass has been focused as renewable and sustainable resources for energy and many chemical products. It is well-known that gut symbiotic microbes in consortia are inevitable for the lignocellulose digestion in many insects. The cellulase and xylanase activity of the crude extracts from each gut contents and tissues, feces, and blood as a control from Asian rhinoceros beetle were analyzed with the overlaid agar-plate assay and spectrophotometric reduced sugar determination. In the plate assay, the highest activity of cellulase and xylanase was observed from midgut tissue extracts, and next from midgut contents, and minor activity from feces. Whereas no enzyme activity was detected from the samples from blood, crop, and hind gut. Similar results were confirmed from the reducing sugar assay with substrates of Avicell and xylan from birchwood, respectively. Further research on the biochemical and genetic property of symbiotic microbes and their enzymes relating on the lignocellulose degradation is planned.
In order to establish artificial indoor rearing techniques for the black soldier fly (BSF), we developed indoor rearing instructions and collection manual for each developmental stage of the fly. The fly collection was conducted between June and October at 1 month interval. Calf feed and food waste were most effective to attract adults. Collection efficiency is higher from the trap installed in a shady spot than that in an open spot. It was highest to collect flies in August and September. As the egg-laying medium for the artificial egg collection, calf feed and food waste were most effective. The optimal number of the medium (W*D*H=60*40*15cm) was 8 for 2000 adults (male:Female=1:1) in the egg-laying net (W*D*H=4*2*2m). Flower foams and wooden blocks with holes were used as egg-laying sites. Adult females preferred the holes on average 3~5mm in diameter and 7~10mm in depth for oviposition. Larvae reared in low density (2~4 individuals/㎠) showed superior practical traits than those reared in an overcrowded environment (6~10 individuals/㎠). It is important to place the fly pupae in moist sawdusts (humidity: 20~40%), since a pupa tends to hide in a refuge. Adult insects should be employed for laying eggs for food waste processing immediately after emergence. When treated at 10℃ for 10 days after pupation, emergence rate of the insect was still 93.3%. By keeping pupae at the low temperature, emergence timing could be manipulated with about 10 day flexibility.
Chrysanthemum flower model trap (CFMT) developed to attract the flower thrips was evaluated for its optimal trap density that is required to manage the population of flower thrips in a strawberry greenhouse. CFMTs were installed in different rates per unit area (10m×5m), i.e., control (no trap), low (5 traps), medium (10 traps), and high (20 traps), in three different strawberry greenhouses at Namhoo, Andong. From each plot, 10 flowers were sampled every week from 16 May 2008 to 6 June 2008. Five traps were also randomly collected from each plot every week to record the number of flower thrips trapped. The seasonal number of both female and male thrips collected in the flowers was significantly lower in the plots with high density of traps compared to the control. In the plot with medium level of traps, less seasonal number of male F. intonsa were found in the strawberry flowers compared to the plot with low level of traps or control. When the numbers of thrips in the flowers were plotted against the numbers of thrips trapped in the CFMTs, significant correlation was found in female thrips only. However, there was no significant difference in seasonal number of male and female thrips trapped in the CFMTs randomly collected from the plots among the treatments. In conclusion CFMT used in medium or high density in the strawberry greenhouse reduced infestation of F. intonsa in the flowers and can be a good addition to integrated pest management tactics against anthophilous thrips.
Propylea japonica is small ladybug(approximately 4-5mm) and is met with everywhere in Korea. When there was no aphid, the survival rate of P. japonica was 0% among another instar bugs and 26.7~32.8% among same instar bugs after 48 hours. When there were sufficient aphids, the cannibalism of P. japonica was low, so the survival rate of P. japonica was 78.6~95.8% among another instar bugs and 80.6~100% among same instar bugs after 48 hours. In the comparison of the number of eggs at different oviposition site, P. japonica preferred the corrugated cardboard. In experiment using T-tube, the rate of decoy was 81% at cotton aphid compared with pepper leaf and 62% at cotton aphid compared with artificial diet. In the artificial diet using shrimp and chicken liver, the developmental period from egg to adult was 17days and the survival rate from egg to adult was 66.7% but the vitality was not good. These data suggest that P. japonica can be used effective natural enemy for control of aphids and is needed to research about artificial diet, mass rearing system and control effect in field.
The injury of red bean (Vigna angularis) by the legume pod borer, Maruca vitrata (Lepidoptera: Pyralidae), was evaluated through an artificial inoculation experiment in cages randomly arranged in a field. Virgin adult pairs (0, 1, 2, 4 and 8) were released into screen-nets at the flowering stage of red bean, and the damage was investigated at harvest. Any insecticides were not sprayed through all plant-growing season. The absolute number of uninjured pod in a net showed a decreasing tendency according to the increase of the number of released adult pairs, but did not show a statically significant difference among treatments. The total production (number of pod and seed) of red bean was highly variable even in a treatment. The variation seemed to be caused by pre-infestation by M. vitrata and other insect species before netting. Instead, therefore, relative damage was compared among treatments. The total ratio of injured pod in a net increased according to the released adult number. The result clearly indicated that the mating ratio and fecundity within the screen-net also increased according to the number of released pairs and that the hatched larvae affected the damage. The corrected rate for pod injury at 8 pair release was about 46%. On the other hand, the ratio of injured seed per injured pod was about 80% regardless of treatments. The result suggested that one pod continuously be attacked till almost disappearance of itself.
Determining age distributions of insect is required for many basic ecological studies. Most of insect developmental studies, the head capsule width and other developmental traits have been widely used to distinguish the instars. Unlike to other insect species, collembolan continues to molt after they reach sexual maturity. So these traits sometimes make it difficult to conduct developmental studies. Model animal used in this study was Paranura rosea a native collembolan speciese. To determine the age groups of P. rosea, we periodically (1week) made digital images of P. rosea (over 100 different images) under the magnifier (DIMIS-M) for consecutive 20 week. And then the head capsule width and the surface area of P. rosea were analyzed using a image software (ImageJ). In this test the head capsule widths and surface area were set as developmental parameters. Both two developmental parameters showed time-dependent increase patterns. And they gave good agreements between predicted developmental parameters and observed parameters (r2=0.83 and 0.71). Based on the reproducibility of juveniles and statistical mean separation tests (tukey's test), both two developmental parameters could be divided into 3 age groups. The information given this study will be used as a basis for understanding the population dynamics and other ecological research.
Consistent individual difference in a behavior is recognized as an important evolutionary force in niche variation and may be a personality trait, an inherent property of the individual. To understand whether aggressiveness is a personality trait, independent of resource holding potential (RHP) and resource value in animal aggression assumed by game theory models, I determined individual variation in aggressiveness by measuringboth positive phonotactic and acoustic responses to the broadcast stimulus repeatedly over the three-week period in males of Gryllus rubens. Factors tested included amplitude levels of the broadcast stimulus, body size, wing dimorphism, and age. The analyses of the generalized linear mixed models revealed that RHP factors such as age, wing dimorphism, and body size, were not significant for both positive phonotactic and acoustic responses. However, there seemed to be more aggressive responses to higher levels of amplitudes. The significant variance-covariance structures of the models indicated that some individuals consistently aggressive or nonagressive during the experimental periods. Because potential factors for resource values were controlled, and because potential RHP factors would not explain variation in aggressive responses, I concluded that aggressiveness is a personality trait that is not characterized by factors that I tested.
Cryptic mealybug (CMB), Pseudococcus cryptus, is a serious scale pest on citrus, especially grown in plastic greenhouse on Jeju island, Korea. Binomial sampling plan was developed using the relationship between infested proportion of twig with less than T scales (pT) and mean density per twig (m) according to the empirical model ln(m)=α+βln(-ln(1-pT)). T was defined as tally threshold and set to 1, 2, 3, 4 and 5 scales per twig in this study. T=3 was the best tally threshold for estimating densities of CMB based on the precision of the model. Binomial model with T=3 provided reliable predictions of mean densities of CMB observed on commercial citrus orchards. The obtaining model was evaluated by resampling validation of Wald's sequential binomial plan and by fit at 95% confidence intervals for independent collected data sets. Binomial sampling program for decision-making of CMB population level was obtained at the action threshold 0.10, proportion of twig infested with more than 3 scales.
Pine wilt disease (PWD) is one of the most serious forest tree diseases and now the damaged area covers 7,877 ha and more than 60 cities in Korea. The causal agent of PWD, Bursaphelenchus xylophilus is transmitted by the insect vectors, pine sawyers (Monochamus spp.). Spread of PWD damage and control effect for PWD are directly related to dispersal potential of Monochamus spp. vectors. Recently, harmonic radar technology has been applied as a means for tracking individual dispersal of tagged insects. Tracking insects with harmonic radar system is similar to a mark-and-recapture technique, in which an organism is tagged, released, and recaptured. Our objective is to develop a harmonic radar tracking system by using lightweight tags to investigate dispersal of Monochamus spp., insect vectors. The harmonic radar system consists of two parts: a commercially available, light-weight (1.6 kg), handheld transceiver (RECCO Rescue System) and a small tag that reflects the transceiver signal (917 ㎒) at harmonic frequencies (1,834 ㎒). A tag consists of a miniature diode (Toshiba ISS350) with wires (36-gauge tinned copper) soldered using conductive epoxy on each side to produce a dipole antenna. These tags were evaluated in an open field to assess their maximum detection range, and at tag length of 13 ㎝, the tag was detected at >100 m.
The first occurrence of oak wilt disease(OWD) in Korea was reported in 2004 and a Platypodid beetle, Platypus koryoensis, was known to contribute significantly the occurrence of the disease. To reduce the oak wilt disease, it is necessary to control density of P. koryoensis under injury level. This experiment conducted to clarify the flying period of adult P. koryoensis. Four forests containing dead trees by OWD were selected and the location of the experimental sites were Mt. Surak in Seoul, Goyang-si, Paju-si in Gyeonggi-do, Hongcheon-gun in Gangwon-do. Sticky trap or vertical multi-funnel trap was settled on Quercus mongolica trees located in infested forest. Density of P. koryoensis was survey with one or two week interval from May to October in 2007 and 2008. The number of the beetle collected by the vertical trap and sticky trap was positively correlated (r2=0.69). The optimum flying period of the beetle was ranged from late June to late July with geographical variations.