Acta Entomologica Sinica

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Monthly, Founded in 1950
Supervisor:Chinese Academy of Sciences
Sponsor:Institute of Zoology,Chinese Academy of Sciences
The Entomological Society of China
Domestic postal code: 2-153
Foreign issuance code: Q61
ISSN 0454-6296
CN 11-1832/Q

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20 May 2008, Volume 51 Issue 5

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RESEARCH PAPERS

Select Cloning, expression and sequence analysis of Bmimd , an innate immunity gene in the silkworm, Bombyx mori

ZHANG Yu-Li

2008, 51(5): 459-465.

Abstract ( 3449 )

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The cascade of innate immunity in organisms was regulate d by a set of genes. The gene immune deficiency of Drosophila plays an important role in the pathway of humoral immunity. We got a homolog of Drosophila imd from Bombyx mori successfully through silico cloning, and named it Bmimd. The gene is 1 092 bp in length, consisting of 4 exons and 3 introns, containing an ORF of 750 bp, encoding a protein with 250 amino acids, and the predicted molecular weight is 28.6 kD. Bmimd contains a death domain, which is similar to mammalian RIP by sequence analysis. Prokaryotic expression of the gen e recombinant with six-His tag and a Nus·Tag was successfully carried out through sub-cloning into PET-50b vector. The result of Western blotting showed that the gene was expressed in head, fat body, germen, cuticle and midgut, but its ex pression was not detected in silk glands of the silkworm.

Select Eukaryotic expression of the cocoonase gene from Bomb yx mori and biological activities of the expressed product

WU Yu-Dan

2008, 51(5): 466-472.

Abstract ( 4109 )

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In order to research the biological activity of cocoonase expressed through eukaryotic expression system, cocoonase gene (GenBank accession number: EF428980) was cloned into baculovirus transfer vector pFastBac^TM1 to obtain pFast-cocoonase, and this recombinant vector was then transformed into DH10Bac competent cells. Cocoonase gene was verified to have been successfully transferred to recombinant bacmid by PCR. BmN cells were transfect ed with the recombinant bacmid through lipofectin mediated method to obatin rec ombinant baculovirus. SDS-PAGE analysis showed that a specific band of about 27.6 kD existed on the lane of expression products of recombinant baculovirus, which was consistent with predicted size of the target protein. SEM micrographs were taken after treating silkworm silk with the expressed protein, and the results s howed that the expressed protein could hydrolyze sericine to some extent.

Select Cloning and prokaryotic expression of the cDNA sequence encoding ubiquiti n from Musca domestica

JIN Feng-Liang

2008, 51(5): 473-479.

Abstract ( 3407 )

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Ubiquitin proteasome pathway is the most important and hi ghly selective proteolytic pathway which plays an important role in degrading the intracellular proteins selectively. The cDNA sequence encoding ubiquitin from  Musca domestica was cloned by RT-PCR and sequenced. Sequence a nalysis showed that the length of this ORF is 228 bp, encoding 76 amino acids, which was named Mdubi and registered in GenBank with accession no. DQ115796.Multiple sequence alignment indicated that Mdubi was very similar to the homologous proteins of other eukaryotic species and it shared more than 94 % amino acid sequence identity with ubiquitins from other eukaryotic species. The expression of Mdubi transcript was quantified by the semi-quantitative RT-PCR, and the results demonstrated that the expression of Mdubi was ubiquitous and not influenced by stimulation of Escherichia coli.The Mdubi was inserted into expression vector pQE30 in vitro and transfo rmed into E. coli M15.The M15 strain, containing Mdubi recombinant plasmid, expressed a 9.6 kD protein with 6His tag at N-terminus in agreement with the expected molecular weight after the induction with IPTG. The fusion protein was purified by Ni²⁺-NTA column and used to raise antiserum. The successful cloning and expression of the coding sequence of M.domestica ubiquitin provided a basis for further study on its function.

Select Cloning and activity analysis of insect antifreeze protein gene from Pterocom a loczyi(Coleoptera: Tenebrionidae)

MA Ji

2008, 51(5): 480-485.

Abstract ( 3598 )

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To investigate antifreeze protein gene in the beetles liv ing in Xinjiang Junggar desert, Pterocoma loczyi, a

large size desert beetle, was chosen for antifreeze gene cloning. RT-PCR technique was employed to obtain Pterocoma loczyi antifreeze protein gene. One of the obtained cDNA fragment was named Plafp 743.Sequence analysis showed that Plafp 743 encodes 112 amino acids with the repeated constructive sequence CTX₁X₂X₃X₄CX₅X₆X₇X₈X₉, which is the special motif for some of the known insects antifreeze proteins. Recomb inant plasmid pGEX-4T-1-Plafp743 was transferred into Escherichia coli BL21 and induced by IPTG. The expressive product was analyzed by SDS-PAGE, and the result showed that pGEX-4T-1-Plafp743 was expressed in fusion protein with relative molecul ar mass of 36 kD. The fusion protein, named GST-PLAFP, was purified by Glutathione Sepharose 4B, and then identified by Western blotting with mouse antiserum against antifreeze protein from Dendroides canadensis. Then, GST-PLAFP with different concentrations was added to E. coli culture for various days at -6℃ to check its cryoprotective effect on bacteria survival at subzero temperatures. This cryoprotective experiment showed that the fusion protein GST-PLAFP could protect bacteria from freezing to death at conce ntrations as low as 30 μg/mL. The results suggest that AFP might be a widely adopted strategy for overwintering of desert beetles.

Select Characterization of the nonstructural gene promoter of Periplaneta fuliginosa densovirus

YANG Bo

2008, 51(5): 486-491.

Abstract ( 3783 )

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To understand the mechanism of transcriptional regulation of Periplaneta fuliginosa densovirus (PfDNV) nonstructural genes，5′-flanking sequence of ns3 gene (325 bp), which encompasses the region from the

5′-terminus of the viral genome to the translation init iation codon of the ns3 gene, was cloned. This construction was subsequently used as a template for generation of six 5′-or 3′-end unidirectional truncation mutants and two site directed mutants in the PCR amplification. These amplified products were subcloned into the report plasmid pGL3-basic and the resultant plasmids were transfected into several insect cell lines. The results indicated that the 325 bp DNA sequence possessed promoter activity in insect cell lines including Sf9, Ld652, Tn368, and S2 cells. Subsequent promoter deletion analysis showed that the promoter had a TATA dependent and TATAindependent transcriptional activity. In addition, we found that the tr a nscriptional activity of this promoter could be transactivated by the viral nons tructural protein NS1 in a concentration dependent fashion in S2 cells.

Select Bioactivities of two tebufenozide derivatives against five lepidopteran pests

CUI Quan-Min

2008, 51(5): 492-497.

Abstract ( 3685 )

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In order to compare the toxicities of tebufe nozide and its derivatives, the toxicities of tebufenozide derivatives 0593 (Ⅲf) and 0673 against five lepidopteran pests,i.e., Spodoptera exigua (Hübner), Prodenia litura (Fabricius), Plutella xylostella (Linnaeus), Helicoverpa armigera (Hübner) and Ostrinia furnacalis (Guenée), were investigated with leaf-dip, dipping and topical application compared with the tebufenozide. The effects of teb ufenozide and 0593 on growth and development of S. exigua were studied with leaf-dip. The results showed that the contact toxicities of 0593 and 0673 to 4th-instar larvae of S. exigua were 11.3 and 7.4 times that of tebufenozide, respectively. The contact toxicit ies of 0593 and 0673 to 4th-instar larvae of P. litura were 30.4 and 24.7 times that of tebufenozide, respectively. The toxi cities of 0593 and 0673 to 3rd-instar larvae of P. xylostella with dipping method were 4.7 and 4.5 times that of tebufenoz ide, respectively, and the contact toxicities of 0593 and 0673 to 3rd-instar larvae of H. armigera were 15.5 and 15.2 time s that of tebufenozide, respectively. The contact toxicities of 0593 and 0673 to 4th-instar larvae of O. furnacalis were 1.3 and 2.0 times that of tebufenozide, respectively. The toxicities of 0593 and 0673 to the five pests were higher than that of tebufenozide, but the differences of toxicities between 0593 and 0673 were not significant. Treated with tebufenozide and 0593, the 2nd-instar larvae of S. exigua died unceasingly after 72 h; the pupation rate and the number of normal pupae were decreased; in addition, adult emergence ratio, eggs laid per female and egg hatchability were also reduced significantly, and the fecundity was so affected significantly. Tebufenozide de rivative 0593 showed more disadvantageous influence than tebufenozide on the fec undity of S. exigua at the same concentration.The results suggest that tebufenozide derivative 0593 is a pesticide with higher bioactivity against lepidopteran larvae and stronger harmful effects on their development and fecundity than tebufenozide, so it is worthy to exploit

Select Insecticidal effects of Oedaleus asiaticus entomopoxvirus in combination with chemical insecticides and effects on the main activities of detoxification enzymes of its host

YANG Xin-Hua

2008, 51(5): 498-503.

Abstract ( 3794 )

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Third-instar nymphs of Oedaleus asiaticus were fed on artificial diet mixed with combinations of O. asiaticus entomopoxvirus (OaEPV) and the insecticides of malathion, chlorpyrifos, β-cypermethrin, cyfluthrin and deltamethrin, in which OaEPV was used as a synergist. Mortalities were observed, and LC₅₀ values and the synergic ratios were determined. The specific activities of two enzymes (carboxylesterase, CarE; glutathione S-transferases, GSTs) in relation to the pest resistance to insecticides were also assayed. The results showed that combinations of OaEPV and chlorp yrifos, β-cypermethrin, cyfluthrin, deltamethrin had no distinct synergism, whereas the combination of OaEPV and malathion was more effective than malathion alone in some degree. The synergic ratio of OaEPV to malathion was 1.42 times. Except del tamethrin, the specific activities of CarE in the midgut of O. asiaticus nymphs treated with mixtures of OaEPV and other insecticides mentioned above were inhibited obviously as compared to insecticide treatments alone. The highest inhibition ratio of CarE specific activity, 4.21 times, appeared when OaEPV was m ixed with malathion. The specific activities of GSTs in the midgut of O. asiaticus treated with the mixtures of OaEPV and cyfluthrin or OaEPV and delta methrin were evidently inhibited as compared to insecticide treatments alone. The specific activities of CarE and GSTs in the fat body of O. asiaticus treated with the mixtures, however, were not significantly changed as compared to insecticide treatments alone. The results suggest that OaEPV enhances the insecticidal effects of chemical insecticides against O. asiaticus by inhibiting the specific activity of CarE in the midgut.

Select Histological effects of fenoxycarb on the Oriental rat flea, Xe nopsylla cheopis (Siphonaptera: Pulicidae)

ZHANG Ying-Chun

2008, 51(5): 504-508.

Abstract ( 3475 )

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【Objective】 To provide the fundamental dat a of controlling the flea species of media of plague and to probe insecticidal m echanisms of fenoxycarb to the early third instar larvae and the newly emerged a dults of Xenopsylla cheopis (Rothschild, 1903) through histological observations. 【Methods】 The early third instar larvae and the newly emerged adults of X. cheopis were treated with fenoxycarb by microliter syringes. The changes of tissues were determined by the methods of histology, microphotographics and statistics. 【Results】 The cuticula of the third instar larvae of X. cheopis treated with fenoxycarb were thicker, the reproductive cells of ovarial rudiments became atrophied, and interstitial tissues of sperma gonia became less. The testicular plug of the newly emerged adults of X. cheopis treated with fenoxycarb disappeared faster than treated with a cetone, the salivary gland cells were damaged seriously, and the midgut epitheli a become atrophied. 【Conclusions】 (1) Fenoxycarb caused the death of X. cheopis larvae by disturbance of metamorphosis and changes in cuticula, ovarial rudiments and testicular rudiments. (2) Fenoxycarb escalated disappearance of testicular plug in the newly emerged male adults. (3) Fenoxycarb caused the death of the newly emerged adults of X. cheopis by damage of salivary gland cells and atrophy of midgut epithelia.10 Refs. In Chinese

Select Comparisons among soil collembola community characteristics in re lation to different vegetation restoration treatments in the moderate degraded g rasslands in the Songnen Plain of Northeast China

WU Dong-Hui

2008, 51(5): 509-515.

Abstract ( 3671 )

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This study aimed to analyze the differences of soil collembola community characteristics under different vegetation reclamation treatments in the moderate degraded grasslands in the Songnen Plain of Northeast China.

Based on the data of soil collembola collected in the Songnen grasslands of different treatments from May to October in 2005, the community parameters such as group number, individual density, and diversity index were calculated. The results showed that a total of 1 156 soil collembola individuals were captured, which fell into 9 families and 19 genera. In contrast to over grazing, both fencing enclosure and planting alfalfa could improve soil properties and substantially increase group number, individual density, and diversity of soil collembola in the moderate degraded grasslands. This suggests that fencing enclosure and planting alfalfa were helpful to restore the soil environment and ameliorate collembol a community. However, the individual density of collembolans in planting alfalfa treatment was significantly higher than that of fence enclosure treatment. Further information came from the comparison of collembola taxa that Isotomidae, E ntomobryidae and Sminthuridae were significantly restored after reclamation. Comparing collembola taxa in fencing enclosure vs. plantin g alfalfa treatments, we found that fencing enclosure maybe more favorably restored Entomobryidae and Sminthuridae, and planting alfalfa perhaps more successfully reclaimed Isotomidae. At sites of the fencing enclosure treatment, the individual density of Entomobryidae reached a level of 5.14 times that of over grazing treatment, and the individual density of Sminthuridae reached 2.38 times that of over grazing treatment. At sites of the planting alfalfa treatment, the individual density of Isotomidae reached 3.33 times of over grazing treatment.

Select Effects of meal size and temperature on the detectable period of Sitobion avenae (Fabricius) (Hemiptera: Aphididae) in Adonia variegata (Goeze) (Coleoptera: Coccinellidae)

GAO Shu-Jing

2008, 51(5): 516-520.

Abstract ( 3451 )

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The effects of meal size and temperature on the detectable period of Sitobion avenae (Fabricius) in Adonia variegata (Goeze) were studied using a monoclonal antibody based indirect enzyme-linked-immunosorbent assay (ELISA), which can be used to evaluate the control effects of predatory natural enemies on S. avenae in wheat fields. The results showed that the curves of the middle decayed product of S. avenae in the gut of the predator, A. variegata, exhibited single peak modes at different temperatures and meal sizes. Temperature had a significant effect on the prey detectable period. With temperature increasing, the prey detectable period decreased. Especially when temperature was up to 30℃，the decay rate of prey increased sharply and the detectable period was very short, only 1.18 day. Meal size also affected the prey detectable period remarkably. While the number of preys consumed by the predators increased from one aphid to three aphids, the prey detectable period increased from 2.80 to 4.25 days at 25℃

Select An approximate variance estimator for index of population trend developed with delta-method and its application

TAO Fang-Ling

2008, 51(5): 521-525.

Abstract ( 3499 )

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On the basis of Watt's mathematical model of the index of population t rend (I), an approximate variance estimator of I was developed with delt a method in this paper. Taking the natural population life tables of rice leaf roller Cnaphalocrocis medinalis (Lepidoptera: Pyralidae) published by Wu et al. (1986) as an example, it was applied in evaluating the control effectiveness of biologica lagent (Trichogramma japonicum Ashmead) and chemical insecticide (powder, 1.5% mevinphos + 3% alpha-hexachloro cyclohexane). By Z-test criterion, biologically and statistically sound conclusions were drawn that at the generation level, the suppression effectiveness of trichogramma wasp against rice leaf roller was better than the blank control with P =0.0111 and smaller I value (0.0390 versus 0.1768), and it was also statistically better than the insecticides with P =0.0036 and smaller I value (0.0390 versus 0.3035). The insecticides was not statistic ally worse than the blank control with greater I value (0.3035 versus 0.1768), and P =0.2236. Although these conclusions are close to that drawn by Wu et al. (1986) only based on the index of population trend point estimates, they are supported by an accuracy measurement P-value.

Select A taxonomic study of the genus Eoscyllina Rehn (Orthoptera: Acridoidea: Ac rididae) from China with description of a new species

ZHENG Zhe-Min

2008, 51(5): 526-529.

Abstract ( 3067 )

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The genus Eoscyllina Rehn from China was reviewed, with five species recorded, including a new species: Eoscyllina heilongjiangensis Zheng et Xu, sp.nov. Key to species of Eoscyllina from China is provided. Type specimens are kept in the Institute of Zoology, Shaanxi Normal University.

Select Advances in insect hemolin structure and function

YAO Hui-Peng

2008, 51(5): 530-536.

Abstract ( 3802 )

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Hemolin is the only member of the immunoglobulin superfamily in invertebrates so far. It plays an important role in Lepidoptera immunity. Based on more than 40 literatures on hemolin, we found that hemolin appears only in Lepidoptera, and exists in the two modes, dissoluble and infusible, which show different biological functions. Dissoluble hemolin could resist invading pathogens , such as bacterium and virus by some enzymes and proteins, while the infusible hemolin may have catabatic effect on cell-cell adhesion and virus or bacterium invading cell.

Select Roles of salivary components in piercing-sucking insect-plant interactions

YAN Ying

2008, 51(5): 537-544.

Abstract ( 4746 )

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Recent researches of piercing-sucking insect saliva have revealed that salivary components play important roles in relationships between piercing-sucking insects and plants. For most piercing-sucking insects, they will secrete gell saliva and watery saliva when they feed on plants. The gell saliva will be secreted at the early stage of feeding to form salivary sheath, which is supposed to surround and protect the stylet. This saliva can help piercing-sucking insects to feed on plants directly and indirectly. Meanwhile, the watery saliva contain s many components like pectnase, cellulose, polyphenol oxidase, peroxidase, alkaline phosphatase, sucrase, etc .These components can help piercing-sucking insects in plant tissue penetration, food digestion, detoxification of plant secondary substances and break down of plant defence reaction. Paradoxically, salivary components can also trigger defence reaction of plants. They can either elicit pl ant wounding messengers to trigger the direct defence reaction or elicit the production of plant volatiles to attract carnivorous natural enemies. Many piercing-sucking insects can cause plant pathological reaction differentially and some researches assumed that piercing-sucking insect salivary enzymes, such as polygalacturonase, alkaline phosphatase, sucrase, polyphenol oxidase, etc ., might be responsible for that. However, there have been no direct evidences for these hypot heses. Moreover, the amino acids and proteinase in piercing-sucking insect saliva are responsible for gall-forming of plants. It has been demonstrated that piercing-sucking insects can change salivary components to adapt different host plant species and meet different physiological requirements. Researches in salivary components of piercing-sucking insects may elucidate mechanisms in outbreaks, damages and virus transmission of piercing-sucking insect pests and piercing-sucking insect-plant coevolution, and may also have significance in guiding insect pest management.