【Aim】 Pyridoxal kinase (PLK) is a key enzyme related to VB₆ metabolism. In the previous study, the cDNA of PLK of the silkworm, Bombyx mori, was cloned, and several important and conservative amino acid residues were replaced in this protein by sequence alignment. This study aims to identify the effect of amino acid residues at specific position of PLK of B. mori on enzyme function. 【Methods】 The conserved sites Thr⁴⁷, Asn¹²¹, Ile⁵⁴, Arg⁸⁸ and Trp²³⁰ were site-mutated respectively by using over-lap extension. The expression plasmid pET-22b(+)-PLK was constructed and transformed to Escherichia coli Rosetta for induction and expression, and then the function of the recombinant protein was analyzed after expression product was purified using affinity chromatography. 【Results】 Compared with the wild type PLK, the PLK activities of the mutantions Thr⁴⁷, Ile⁵⁴ and Arg⁸⁸ were reduced by 82%, 58% and 85%, respectively, while the PLK activity of the mutantion Asn¹²¹ was hardly affected and the PLK activity of the mutantion Trp²³⁰ vanished. 【Conclusion】 In this study, the significance of selected amino acid residues of side chains on the catalytic function of PLK of B. mori was clarified.

ApisOBP3 from Acyrthosiphum pisum was known as its affinity to E-β-farnesene (EBF), and the volatile substances released by Drepanosiphum platanoidis contain no EBF which is a conventional component secreted by most of aphids. In this study, two odorant binding proteins named as DplaOBP3-1 and DplaOBP3-2 sharing high identities (95.8％ and 98.32%, respectively) with ApisOBP3 were identified in D. platanoidis. The lengths of coding sequences of both DplaOBP3-1 and DplaOBP3-2 are 426 bp with 69 bp coding for signal peptide at the 5′ end. DplaOBP3-1, DplaOBP3-2 and ApisOBP3 share 6 highly conserved cysteine residues at the same position among sequences. Based on the 3D structure of LmadPBP, a pheromone binding protein from cockroach Leucophaea maderae, we modeled the structures of DplaOBP3-1 and DplaOBP3-2, and all the five hydrophobic amino acids and the aromatic amino acid residue, Tyr84, located in the binding cavity probably participating in interacting with ligands have their counterparts in the structure of ApisOBP3. Further, four-arm olfactometer assay showed that D. platanoidis were significantly repelled by both EBF and crushed aphids （P=0.01）. The results suggest that volatiles from fresh crushed bodies could be the intraspecies alarm pheromone in D. platanoidis; D. platanoidis are repelled by allogenetic EBF whereas they never release EBF.

 【Objective】 This study aims to determine the activities of polyphenoloxidase (PPO) in the fruit fly Bactrocera tau (Walker) during various developmental stages. 【Methods】 The PPO activity and kinetic properties in the 1st, 2nd and 3rd instar larva, pupa and adult were determined with spectrophotomethrical method using catechol as the substrate. 【Results】 The PPO activities varied significantly during different developmental stages of B. tau. The enzyme activity in  the 3rd instar larva was the highest (434.42 U/mg) and that in pupa was the lowest (231.05 U/mg). At pH 6.5, the enzyme activities in the 1st, 2nd and 3rd  instar larva, pupa and adult were 265.42，  358.34, 444.42，  210.02 and 373.99 U/mg, respectively. However, PPO activities decreased dramatically at pH levels above 7.0 or below 5.0. At 34℃ and 37℃, PPO activities stayed at a high level, while the enzyme activities significantly decreased at above 40℃ or below 27℃. When catechol was used as the substrate, the measured Michaelis-Menten constant (K_m) and maximum velocity (V_max) of PPO in 2nd instar larva were 3.10 mmol/min and 476.19 mmol/L, respectively; but those in pupa were 0.63 mmol/min and 50.25mmol/L, respectively, indicating that the catalytic activity of PPO in 2nd instar larva to the substrate catechol was higher than that in pupa. When L-DOPA was used as the substrate, the measured K_m and V_max of PPO in the 3rd instar larva were 0.49 mmol/min and 188.68 mmol/L, respectively; in contrast, the K_m (0.25 mmol/min) and V_max (21.79 mmol/L) of PPO in pupa were relatively lower. 【Conclusion】 Our results indicate that the properties of PPO in B. tau at different temperature and pH values are closely associated with its developmental stage.

Diadegma semiclausum Hellen is a larval endoparasitoid of the diamondback moth, Plutella xylostella L. The parasitoid regulates its host physiology and development mediated by its maternal secretions, such as venom and polydnavirus (PDV), but the possible functions of venom of this PDV-producing endoparasitoid are unknown. The SDS-PAGE analysis of venom showed that the most protein bands ranged from 35 to 220 kDa with a few bands less than 15 kDa, but the most abundant bands were within 35-70 kDa, which were similar to venom proteins in other parasitoid-host systems. By in vitro primary-culture of host hemocytes with parasitoid venom fluid or Escherichia coli, the changes of spreading and phagocytic abilities of hemocytes from the parasitized or the non-parasitized host larvae indicated that the venom of D. semiclausum alone was harmless to the spreading and phagocytic abilities of plasmatocytes and granulocytes in larvae of the diamondback moth. However, the spreading ability of hemocytes from host larvae was obviously inhibited in the early stage of parasitism by D. semiclausum, but these hemocytes could still recognize the pathogens with a lapse of endocytosis; at 24 h post-parasitization, the hemocytes recovered their spreading ability and then granulocytes acted as phagocytes to engulf bacteria successfully. This study demonstrates that the parasitism of D. semiclausum can affect the phagocytosis of hemocytes from its host larvae through inhibiting the spreading ability of granulocytes.

In order to explore the effects of heavy metals on transfer, biomagnification and developmental duration in spiders, the changes of cadmium content in various instars of Pirata subpiraticus fed with Drosophila melanogaster flies reared on artificial diets were detected by atomic absorptions spectrometry and indirect effects of cadmium on its developmental duration were measured. The results showed that excessive cadmium in food could be transferred by food chain and accumulated in P. subpiraticus and the accumulation level increased with instar duration extended, which was significantly positively related with the number of fruit flies ingested and the cadmium content in food, respectively. It was estimated that 65.4% of the Cd²⁺ was assimilated by the spiders and the biomagnification factor was approximately 1.8. Cd²⁺ content in P. subpiraticus had no significant change in the observed 4 weeks of feeding with uncontaminated flies. Cadmium in food significantly led to lengthening of developmental duration and reducing of starvation tolerance in P. subpiraticus. The results might provide adequate basic theory on Cd²⁺ transmission, biomagnification and physiological tolerance along the soil-insects-predators.

Both destruxin A and destruxin B are bioactive molecules. In the present study, the effects of the two destruxins (destruxin A and destruxin B ) on Spodoptera litura SL-1 cells were compared with MTT assay, inverted phase contrast microscope (IPCM), fluorescence microscopy (FM) and flow cytometry (FCM) technologies. The results showed that both destruxins had an apparent effect on inhibition of cellular proliferation, and they had a positive relationship of time-concentration-efficacy. The IC₅₀ values were 7.80 and 20.73 μg/mL at 48 h after treatment with destruxin A and destruxin B, respectively. With inverted phase contrast microscopy observation, we found that both destruxins could cause cell rounded, celluar membrane shrunk, and apoptotic bodies formed. Most of cells were suspended and dead as vacuoles appeared and cytoplasm leaked out with extension of the treatment time. However, under the same concentrations tested, destruxin A showed stronger effect than destruxin B. With fluorescence microscopy observation after AO/EB staining, a relative higher fluorescent intensity of the cells induced by destruxin A was found as compared with destruxin B. Observations by flow cytometer showed that destruxin A and destruxin B could induce apoptosis in SL-1 cells, and the total apoptotic rates reached 78.88%±0.97% and 72.23%±2.29% at 48 h after treatment with 10 μg/mL destruxin A and destruxin B, respectively. This research demonstrated that destruxins possess a preferable activity for cell proliferation and apoptosis at the cellular level, and provided some theoretical support for their potential application in pest control.

SpltMNPV X₁₇ was isolated from the stock collected in Ogasawara Island, Japan, by in vivo cloning techniques. In order to reveal its genotype, the primer was designed according to the complete genome sequence deposited in GenBank under accession no. NC_011616. The polyhedrin gene of X₁₇ was amplified by PCR, and then the neighbor-joining tree based on nucleotide sequences of the polyhedron gene between SpltMNPV different genotypes including X₁₇ and 37 NPVs was constructed. The phylogenetic tree indicated that SpltMNPVs were broadly divided into three groups, i.e., SpliNPV (A) type, SpltMNPV (B) type and SeMNPV (C) type, which is consistent with other reported research results by DNA restriction endonuclease analysis. X₁₇, SpltMNPV-1 and SpltMNPVⅡ clustered into one clade, which was genetically away from the other SpltMNPV genotypes and belonged to SpltMNPV C genotype. Moreover, the genes 38.7kD, Lef-1, Lef-9, fp, p10 and p74 of X₁₇ were also cloned by PCR amplification, and homologous analyses of the genes between X₁₇ and SpltMNPV, SpltMNPVⅡ, SeMNPV and SfNPV were performed. The results showed that based on the six cloned ORFs the homology between X₁₇ and SpltMNPV is the lowest. The identity for amino acid sequence of the late expression factor 9 (lef-9) is the highest (69%), but that of the 38.7 kD amino acid sequence is only 26%. The homology of the most genes amplified between X₁₇ and SpltMNPVⅡor SeMNPV is higher than that between X₁₇ and another NPV analyzed here. The identities of the fp25K amino acid sequence between X₁₇ and SpltMNPVⅡor SeMNPV are both high (up to 95% and 96%, respectively), but that of the 38.7kD amino acid sequence is the lowest of the six ORFs (64%). It is reasonably concluded that X₁₇ is a new strain of SpltMNPV C gentype, named as SpltMNPVⅡ-1. This study provides some theoretical basis for further study and utilization of X₁₇ strain.  

The cytoplasmic incompatibility (CI) is the most common effect of Wolbachia on the reproduction of its arthropod hosts, and the expression of CI differs greatly among different populations. Using the Jiangsu (JS) and Liaoning (LN) populations of the two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), as experimental materials, 100% infected and uninfected Wolbachia lines were obtained by screening. The present study tried to evaluate some factors influencing the expression of CI in the spider mite by crossing experiment and Real-time quantitative PCR. These factors include age of host, temperature, host genes and Wolbachia density. The 1, 3, 5, 7-day-old virgin males were used to investigate the influence of host age on Wolbachia-induced CI. The results showed no effect of age on CI, suggesting that host age does not reduce the sperm modification induced by Wolbachia. The effect of temperatures (20℃, 25℃ and 30℃) on the CI induced by Wolbachia was also checked. Neither high nor low temperatures influenced the expression of CI. Wolbachia density in males of the JS population, as measured by quantitative PCR using the wsp (surface protein of Wolbachia) gene, was significantly higher than that in the LN population. In addition, in both the JS and LN populations, Wolbachia density increased with the age of male hosts. Wolbachia density also showed no effect on CI. We estimated the variability of CI expression between the JS and LN population of T. urticae was due to the interaction between Wolbachia and host genotypes. The results might provide foundation for understanding the mechanisms of reproductive manipulation induced by Wolbachia.

In order to clarify the cross-resistance and the change in related enzymes of the imidacloprid-resistant population of Aphis gossypii to other neonicotinoid insecticides, the bioassay method was used to determine the cross-resistance of different resistant populations of imidacloprid-resistant A. gossypii to dinotefuran and nitenpyram, and the synergism of three detoxification enzyme inhibitors to imidacloprid and other two neonicotinoid insecticides using cotton aphids of the imidacloprid-resistant population selected in the laboratory, the Xiajin resistant population in the field in Shandong and the susceptible population. The activities of detoxification enzymes and AChE of three populations of cotton aphid and the inhibition effect of insecticides were determined through biochemical analysis. The results showed that the imidacloprid-resistant population and the Xiajin resistant population exhibited no cross-resistance to dinotefuran, but showed 5.28-fold and 4.89-fold cross-resistance to nitenpyram, respectively. Dinotefuran could significantly inhibit the activities of CarE, GST and AChE of the imidacloprid-resistant cotton aphid. Nitenpyram showed little effect on the activities of CarE, GST and AChE of the imidacloprid-resistant cotton aphid. Carboxylesterase inhibitor TPP and mixed-functional oxidase inhibitor PBO had obvious synergism to imidacloprid and nitenpyram, while had little synergism to dinotefuran. Glutathione-S-transferase inhibitor DEM showed no obvious synergism to the three insecticides. Dinotefuran and nitenpyram could inhibit the activities of detoxification enzymes and AChE of the imidacloprid-resistant conton aphid, with dinotefuran showing significant effect. The results demonstrate the great application value of dinotefuran in control of the imidacloprid-resistant cotton aphid, and its structure can provide a reference to the development of neonicotinoid insecticides in the future.

Sterols are important secondary metabolites in plants with a variety of biological activities. In order to reveal the mechanism of sterols from plants against insect pests, the effects of sterol constituents (sterol A and sterol B) extracted from Xanthium sibiricum (Compositae) on feeding, enzyme activities in the hemolymph and midgut, and midgut tissues of 4th instar larvae of Pieris rapae treated by feeding on leaf-discs of crucifer Brassica oleracea were investigated. The results showed that the feeding of P. rapae larvae was obviously deterred by sterol A and sterol B with the AFC₅₀ of 0.0229 mg/mL and 0.0147 mg/mL, respectively, at 24 h after treatment. The activities of midgut amylase, protease and carboxylesterase (CarE) were significantly lower than those of the control within 36 h treatment, and sterol B showed stronger effects, which inhibited activities of midgut protease by 23.74% and 58.59%, and activities of midgut CarE by 49.01% and 83.03% at 24 and 36 h after treatment, respectively. Meanwhile, the content of hemolymph protein was reduced and the CarE activity in hemolymph was induced within the exposure time. Moreover, the microstructures of the tested larval midgut tissues exhibited such distinct pathological changes as the micro-rods of cylindrical epithelium dissolved and the goblet cells in the midgut epithelium deformed at 24 h after treatment. The results suggest that the inhibition of digestive enzyme activity and the damage of midgut tissues might be involved in feeding deterrence of the sterols from X. sibiricum against P. rapae larvae, and the proportion difference of plant sterols may affect their effects on insects.

In order to provide scientific suggestions for selecting new poplar strains resistant to insects, the resistance of 18 clones of Populus deltoids to 3rd instar larva of Clostera anastomosis was evaluated and analyzed in laboratory conditions （28±1℃, RH 70%, 16L∶8D）. The choice and no-choice experiments indicated that eleven of 18 clones had higher resistance to C. anastomosis larvae than I-72 (control). Main nutrients and secondary metabolites were measured, and activities of digestive and protective enzymes and their effects on food utilization and insect development were also studied. The results showed resistance of the clones tested in P. deltoides to C. anastomosis larvae was statistically and positively related with the content of phenolic compounds of their leaves. Differential effects of clones of P. deltoides on the growth and food utilization existed in larvae of C. anastomosis, as showed in the changes of the main indicators of growth and food utilization, such as larval weight, daily body weight gain, relative growth rate, approximate digestibility, efficiency of conversion of ingested food, and efficiency of conversion of digested food. Differential effects of clones of P. deltoides on food intake and feces excretion in larvae of C. anastomosis also existed. The results suggest that C. anastomosis are more sensitive to secondary metabolites and the clones of P.deltoides have apparently active effects on activities of SOD and CAT in C. anastomosis larvae.

【Objective】 There are many plum-peach intercropping or neighboring orchards in many regions in China due to their similar ecological environments. The experiment was carried out to clarify the effect of plum-peach intercropping on trophic structure of ground beetles, and to assess whether plum-peach intercropping is beneficial to pest control. 【Methods】 Ground beetles were investigated with pitfall trapping method in three mono-peach orchards and three plum-peach intercropping orchards in Linfen, Shanxi from April to October in 2006 and 2007, of which each experimental orchard is about 0.3 hm². The Carabidae species richness was compared between the two kinds of orchards. For the six dominant Carabidae species determined, two different species in combination were placed in one glass bottle with 10 beet webworms as food, and the bottles were placed under peach trees. The results of beetle attacking recorded from these bottles were used to determine the trophic level of Carabidae species. The relative abundances of carabids from each trophic level were compared between the two kinds of orchards. 【Results】 The 24 species of ground beetles were found, and the number of species showed no significant difference between in mono-peach orchard and in plum-peach intercropping orchard (P≥0.38). In addition, the six dominant species could be classified into four trophic levels: Lesticus magnus (Motschulsky) belongs to the senior predator, Scarites acutides Chaudoir the intermediate predator, Chlaenius bioculatus Chaudoir and Calathus halensis (Schaller) junior predators, and Harpalus tschiliensis Schauberger and Harpalus griseus (Panzer) herbivores. The relative abundance of carabids at different trophic levels was higher in plumpeach intercropping orchard than in mono-peach orchard. The results of T test showed that extremely significant difference (P≤0.002) and significant difference (P=0.013) in relative abundances of herbivores and junior predators existed between the two kinds of peach orchards in 2006, respectively, but no significant difference (P≥0.085) in relative abundance existed in carabids of other trophic levels in the surveyed two years. 【Conclusion】 It is concluded that plum-peach intercropping has no effect on the species component and richness of the ground beetles in the plum-peach intercropping orchard, but has some effects on population relative abundance of different trophic levels in the ground beetles. In general, plum-peach intercropping has significant effects on carabid abundance at different trophic levels of herbivores, but has no significant effect on carabid abundance at most trophic levels of predators.

Predation of the brown planthopper (BPH), Nilaparvata lugens (Stål) by its predators is particularly important in rice field ecosystem, but its studying is rather difficult. In this study, PCR cross-amplification was initially conducted on DNA extracted from 66 species of arthropods commonly found in rice fields. Only Lug-F1a/Lug-R1 was specific and efficient enough to detect species-specific DNA of N. lugens in the gut of wolf spider Pardosa pseudoannulata. Then, we assessed how digestion time, temperature, feeding amount of predators and the sex of predators can influence the effectiveness of detecting BPHs in spider guts using Lug-F1a/Lug-R1. The results indicated that digestion time, temperature and feeding amount significantly affected the detection of the primers, while the sex of predators had no significant influence. The detection was significantly and negatively correlated with digestion time and temperature, but positively with the feeding amount. Thus the species-specific primers of N. lugens had been successfully identified, which could be effectively used in rice fields. These primers under the conditions tested here provide a new detection tool efficient for studying the relationship between N. lugens and its predators.

Diapause is one of the primary developmental pathway for insects to escape from unfavorable environment. Knowledge of diapause regulation is essential for rational use of beneficial insects and development of effective pest management strategies. Diapause evokes a unique pattern of gene expression, such as heat shock proteins (Hsps) and acquisition of increased tolerance to environmental stresses. The purpose of this review is to consider the advance on heat shock proteins related to insect diapause, including the relationship between diapause and heat shock proteins, differential expression of heat shock protein gene at different life stages during diapuase and the comparative proteomic analysis between diapause and non-diapause individuals. Unlike other common stress responses that elicit synchronous up-regulation of all Hsps, however, expression of Hsps is different during diapause in different species. The up-regulation of Hsps during diapause is an important factor contributing to overwintering defense strategy and survival. This review may provide some reference to understanding the mechanisms of diapausing insect response to environmental stimuli.

In order to explore the mechanism of honeybee’s Zaohua disease caused by date nectar and pollen, worker honeybees of Apis mellifera ligustica suffered the disease were collected from Linyi, Taigu and Linxian, and the activities of amylase, protease, invertase, pectinase, trehalase, esterase (EST), glutathione-S-transferase (GST), cytochrome P450 (P450), acetylcholinesterase (AchE) were determined by spectrophotometry. The results showed that compared with the control (enzyme activity of the normal worker honeybee), the amylase activities of worker honeybees from Linyi, Taigu and Linxian were decreased by 73.17%, 60.74% and 70.50%, respectively; protease activities were decreased by 74.45%, 50.68% and 90.44%, respectively; and pectinase activities were decreased by 39.50%, 27.99% and 73.13%, respectively. However, sucrase and trehalase activities had no significant difference from those of the control （P＞0.05）. GST activities of worker honeybees from Linyi, Taigu and Linxian were enhanced by 1.29-, 1.47- and 1.30-fold, and P450 activities were enhanced by 3.31-, 1.66- and 2.12-fold, respectively. But EST and AchE activities had no significant difference from those of the control (P＞0.05). The activities of digestive and detoxification enzymes of worker honeybees, including amylase, protease and pectinase, are changed after their feeding on date nectar and pollen during date florescence, which may be related to the occurrence of Zaohua disease.

To understand the effects of temperature and photoperiod on egg diapause induction of Apolygus lucorum Meyer-Dür, the hatch rate of A. lucorum eggs laid by those rearing at 3 temperatures and 6 photoperiods were systematically investigated. The results indicated that the 1st instar larvae of A. lucorum was the sensitive stage. The daylength less than 14 h was beneficial to diapause, while the daylength more than 14 h was beneficial to growth and diapause inhabitation at three different temperatures (17℃， 20℃，and 23℃). Short daylength had a cumulative effect on diapause induction. The diapause rate decreased with the increase of temperature in the daylength from 10 h to 14 h, and temperature had no effect on diapause induction in the daylength more than 14 h. The critical photoperiods of A. lucorum were 13 h 10 min∶10 h 50 min （L∶D）， 12 h 58 min∶11 h 2 min （L∶D）and 12 h 51 min∶11 h 9 min （L∶D） at 17℃， 20℃ and 23℃， respectively. The critical daylength shortened with the increase of temperature. The results suggest that A. lucorum is a typical long-day insect, photoperiod is the main factor of diapause induction, temperature is the cofactor of diapause induction, and low temperature in autumn is beneficial to the diapause induction.