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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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Research advances in symbiotic microorganisms in insects and their functions
WANG Wei-Xia, ZHU Ting-Heng, LAI Feng-Xiang
Acta Entomologica Sinica    2021, 64 (1): 121-140.   DOI: 10.16380/j.kcxb.2021.01.013
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Symbiotic microorganisms can account for 1%-10% of insect biomass, including acteria, fungi, archaea and viruses. Insects and symbiotic microorganisms co-evolve to form holobionts. Symbiotic microorganisms play an important role in the biological characteristics, diversity formation, ecological adaptability and stress resistance of insects. Crop pest insects seriously affect agricultural production. In this article, the research advances in the diversity, research methods, and functional mechanisms of insect symbiotic microorganisms, the interaction between symbiotic microorganisms and their application in pest control since 2000 were reviewed and prospected. With the continuous development and application of advanced research methods such as molecular microbial ecology and metagenomic sequencing, breakthroughs have been made in the research of symbiotic microorganisms of agricultural pest insects. It was found that symbiotic microorganisms mainly affect host insects in the following ways: (1) Synthesis of nutrients or production of digestive enzymes to promote host growth and development and to expand host ecological niche; (2) Production of protective metabolites to directly protect the host against stress or indirectly protect the host by regulating the defense response of host plants; and (3) Production of active substances to regulate host propagation, mating, aggregation and movement. The abundance and community composition of insect symbiotic microorganisms maintain dynamic changes in a certain spatial-temporal range and have an important impact on host phenotype, which is the result of the benefit trade-off among host, environment, and interactive microorganisms. We suggest that future research should focus on clarifying the molecular mechanisms underlying the formation and maintenance of symbionts, ascertaining the complex interactions among symbiotic microorganisms, host insects, plants, natural enemies and the environment in more spatial-temporal dimensions, and designing green and efficient pest control strategies through targeted regulation of pest insect symbionts.
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Response of miRNAs related to wing differentiation and their predicted target genes to ecdysone and the confirmation of target gene of miR-92a-1-p5 in the pea aphid, Acyrthosiphon pisum (Hemiptera: Aphidae)
MA Tian-Tian, YANG Zong-Lin, CHANG Mei-Ling, HUO Chun-Yue, KAN Yun-Chao, LI Dan-Dan
Acta Entomologica Sinica    2021, 64 (4): 419-427.   DOI: 10.16380/j.kcxb.2021.04.001
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 Abstract: 【Aim】 Ecdysone plays important roles in wing dimorphism of the parthenogenetic aphid. In the previous study we found that five microRNAs (miRNAs) also play pivotal roles in the wing dimorphism of the pea aphid, Acyrthosiphon pisum, but whether ecdysone and miRNAs have interaction in wing differentiation of aphids is unclear. This study aims to explore the effect of ecdysone on the expression of miRNAs and their predicted target genes, and to reveal the interaction of ecdysone and miRNAs in wing differentiation of aphids. 【Methods】 The five miRNAs (Let-7, miR-92a, miR-92b, miR-92a-1-p5 and miR-277) related to the wing dimorphism of A. pisum were selected. The 2nd instar nymphs of parthenogenetic A. pisum were exposed to 0.1 mol/L ecdysone analog 20-hydroxyecdysone (20E) for 10 min and 30 min, respectively, and sampled at 48 h after treatment. The expression levels of the five miRNAs and their predicted target genes after 20E treatment were detected by qPCR. The predicted target gene of miR-92a-1-p5, flightin, was verified by dual luciferase activity assay. Finally, the expression of miR-92a-1-p5 in the 4th instar nymphs of parthenogenetic A. pisum was knocked down with nanocarrier/detergent to verify the interaction between miR-92a-1-p5 and its predicted target gene flightin. 【Results】 The expression of the five miRNAs in the 2nd instar nymphs of parthenogenetic A. pisum could be extremely significantly induced by the treatment of 0.1 mol/L 20E for 30 min. But when the 2nd instar nymphs were exposed to 0.1 mol/L 20E for 10 min, the expression levels of miR-92a-1-p5 and miR-92b extremely significantly decreased, while those of Let-7 and miR-277 increased extremely significantly compared with the control. The expression trends of Let-7 and its predicted target gene abrupt were opposite in the 2nd instar nymphs of parthenogenetic A. pisum after 20E treatment. The expression levels of wingless and Uba1, which are the predicted target genes of miR-92a and miR-277, respectively, decreased extremely significantly in the 2nd instar nymphs of parthenogenetic A. pisum exposed to 0.1 mol/L 20E for 10 min, showing the opposite trend with those of the corresponding two miRNAs. The expression level of flightin, the predicted target gene of miR92a-1-p5, decreased extremely significantly in the 2nd instar nymphs of parthenogenetic A. pisum exposed to 0.1 mol/L 20E for 30 min, exhibiting an opposite expression trend to that of miR-92a-1-p5. Dual luciferase activity assay results showed that after co-transfection of the mimics of miR-92a-1-p5 and the flightin CDS overexpression vector pmirGlO [flightin] the luciferase activity was extremely significantly decreased by 40% compared to the control transfected with NC mimics. Knocking down the expression of miR-92a-1-p5 in the 4th instar nymphs of parthenogenetic A. pisum extremely significantly decreased its expression by 83%, while extremely significantly enhanced the expression of its predicted target gene flightin by 48%, confirming that flightin is the target gene of miR-92a-1-p5. 【Conclusion】 Ecdysone can induce the expression of miRNAs in A. pisum. miR-92a1-p5 may be involved in wing differentiation of parthenogenetic aphids by regulating flightin gene. Nanocarrier/detergent can achieve effective miRNA interference in aphids. This study lays a foundation for further exploring the interaction of ecdysone and miRNAs in wing differentiation of parthenogenetic aphids.
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Cloning and functional analysis of salivary protein gene Sm13498 of the grain aphid, Sitobion miscanthi (Hemiptera: Aphididae)
FU Yu, WANG Qian, ZHANG Yong, CHEN Ju-Lian
Acta Entomologica Sinica    2021, 64 (9): 1009-1019.   DOI: 10.16380/j.kcxb.2021.09.001
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【Aim】 The grain aphid, Sitobion miscanthi, is a dominant cereal aphid species in the major growing areas of wheat (Triticum aestivum) of China. Sm13498 is a salivary protein specifically expressed in the salivary glands of S. miscanthi. This study aims to investigate the potential role of the functionally unknown salivary protein Sm13498 of S. miscanthi in modulating plant defense. 【Methods】 Based on the sequencing data of the salivary gland transcriptome of S. miscanthi, the full-length cDNA sequence of Sm13498 gene was cloned by PCR and analyzed by bioinformatics. The expression levels of Sm13498 in apterous adults of S. miscanthi feeding on T. aestivum leaves for different time were determined by RT-qPCR. The secretion function of the signal peptide of Sm13498 was verified by yeast secretory system. The function of Sm13498 and its subcellular localization in Nicotiana benthamiana was examined using Agrobacterium tumefaciens-mediated transient expression technique. 【Results】 The full-length cDNA sequence of Sm13498 of S. miscanthi was cloned (GenBank accession no.: MW346655). Its open reading frame (ORF) is 783 bp in length, encoding 260 amino acids with the predicted molecular weight of 28.01 kD and amino acids 1-22 predicted to be N-terminal signal peptide. Phylogenetic analysis showed that Sm13498 was most closely related to LOC100159087 precursor, an uncharacterized protein of Acyrthosiphon pisum, deposited in GenBank under the accession no. NP_0013135481, sharing 71.7% amino acid sequence identity. The RT-qPCR results revealed that the expression level of Sm13498 reached the peak in apterous adults of S. miscanthi feeding on wheat leaves for 12 h. Saccharomyces cerevisiae strain YTK12 containing the signal peptide fragment of Sm13498 grew normally on the YPRAA medium in the yeast secretion system, and catalyzed the conversion of colorless 2,3,5-triphenyltetrazolium chloride (TTC) to insoluble dark-red-colored triphenylformazan (TTF), confirming the secretion activity of the predicted signal peptide. The transiently expressed Sm13498 in N. benthamiana mediated by A. tumefaciens could suppress the programmed cell death induced by Bcl-2-associated X protein (BAX) and pathogen elicitor INF1. Subcellular localization results indicated that the fusion protein  Sm13498-GFP was localized in the cytomembrane of N. benthamiana leaves. 【Conclusion】 The results suggest that the salivary protein Sm13498 of S. miscanthi may be involved in the suppression of plant defense responses. This study lays a foundation for identifying the salivary effectors of S. miscanthi and understanding the high adaptability of wheat aphids to wheat varieties.
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Research progress of olfactory binding proteins in insects 
WU Fan, ZHANG Li, QIU Yi-Lei, LI Hong-Liang
Acta Entomologica Sinica    2021, 64 (4): 523-535.   DOI: 10.16380/j.kcxb.2021.04.011
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 As the first participant in the olfactory system, olfactory binding proteins are mainly expressed in the hemolymph of peripheral olfactory system and play roles in recognizing, binding and transporting odors or pheromones to odor receptors. In recent years, with the application of novel modern biological technology, more olfactory binding proteins have been identified, and their diverse functions have been revealed. In this article, we summarized the research progress in the molecular characteristics, structure, functions and application of olfactory binding proteins in recent years. Generally, olfactory binding proteins are classified into three families, odorantbinding proteins (OBPs), chemosensory proteins (CSPs) and Niemann-Pick type C2 proteins (NPC2). Based on the α-helix and β-sheet, olfactory binding proteins form stable globular structure, which is beneficial for them to adapt to a variety of environments and tasks. And diverse functions of olfactory binding proteins are particularly important for insect physiology and behavior. Nowadays, olfactory binding proteins are used in biological control, variety selection and breeding, biosensor making, and so on. This review provides references and some new ideas for further research on olfactory binding proteins in insects.
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Molecular mechanisms of expression regulation of insect cytochrome P450 genes involved in insecticide resistance
ZHU Jiang, QIU Xing-Hui
Acta Entomologica Sinica    2021, 64 (1): 109-120.   DOI: 10.16380/j.kcxb.2021.01.012
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The frequent and extensive use of insecticides leads to the evolution of insecticide resistance. It is well recognized that over-expression of cytochrome P450s involved in insecticide detoxification is one common and major mechanism of resistance to
various classes of insecticides in insects. However, the molecular mechanisms for overexpression of insecticide resistance-related P450 genes in insects have not been well identified until now. In the recent decade, with the development of biological science and technology, many significant advances have been achieved in this direction. In this article, an overview of the current knowledge with respect to expression regulation of insect cytochrome P450 genes was provided. In addition to gene duplication and amplification, up-regulation of P450s at the transcriptional level can be ascribed to the interaction between the cis-regulatory elements and the trans-acting factors. Transcription factors such as CncC, CREB and AhR directly regulate the expression of P450s. The G-protein-coupled receptors and their downstream effectors play indirect roles in the transcriptional regulation of P450 genes. The universality of the role of CncC:Maf/Keap1 in the expression regulation of P450s is supported by studies on a variety of insect species. Increasing reports confirm the involvement of microRNAs in the regulation of P450 expression. Current findings reveal the diversity of regulatory factors and signal transduction pathways, and the complexity of underlying mechanisms in the regulation of P450 expression.
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Morphology, histology and ultrastructure of the compound eyes of the last instar nymphs and adults of Meimuna mongolica(Hemiptera: Cicadidae)
LAN Ying, WEI Cong
Acta Entomologica Sinica    2020, 63 (12): 1441-1451.   DOI: 10.16380/j.kcxb.2020.12.002
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【Aim】 This study aims to clarify the differences in the compound eyes of the last instar (5th instar) nymphs and adults of the cicada Meimuna mongolica, so as to ascertain the changes in the morphological structure and functions of compound eyes of nymphs and adults of Cicadidae in the process of ecological niche shift. 【Methods】 The compound eyes of the last instar nymphs and adults of M. mongolicawere observed and compared at the morphological, histological and ultrastructural levels using light microscopy, scanning electron microscopy and transmission electron microscopy. 【Results】 The compound eyes of M. mongolica are of apposition type. The color of the compound eyes of the last instar nymphs change from white to red and dark brown gradually before adult emergence, whereas the compound eyes of the adults are light brown. Sensilla basiconica, sensilla trichoid and sensilla chaetica were observed on the surface of the compound eyes of the last instar nymphs, but no sensillum was found on the surface of the compound eyes of adults. The surface of the white compound eyes of the last instar nymphs has a complete cornea, and no ommatidial facet was observed. The red and dark brown compound eyes of the last instar nymphs are composed of many irregular, hexagonal or pentagonal ommatidia, whereas the compound eyes of adults are composed of equilateral hexagonal ommatidia. The white compound eyes of the last instar nymphs are composed of a large amount of clustered cells, and no ommatidium was observed. In each ommatidium of the dark brown compound eyes of the last instar nymphs, the nuclei of secondary pigment cells are distributed between the rhabdom and the crystalline cone. In the compound eyes of adults, the nuclei of secondary pigment cells are distributed around the proximal center of crystalline cone. In the dark brown compound eyes of the last instar nymphs, the pigment granules of the primary pigment cells and retinula cells are evenly distributed, whereas those in the compound eyes of adults are mainly distributed around the rhabdoms.【Conclusion】 The compound eyes of the last instar nymphs and adults of M. mongolica show significant differences in the development of ommatidia, the distribution of pigment cells, and the presence or absence of sensilla. The major developmental period of compound eyes of cicadas is during the last instar nymphal stage when the compound eyes are red in color. The results suggest that the compound eyes of cicada nymphs during the earlier developmental stage of the last instar as well as the earlier instars do not have visual ability, but can function to sense mechanical pressure or other environmental signals underground. This should be an adaptation to the niche shift during the ontogeny of cicadas which live underground for a very long time in the nymphal stage and have a short adult lifespan above ground. The findings of morphological, ultrastructural and functional characteristics of compound eyes of the last instar nymphs and adults of this species improve our understanding of the changes in the development and function of compound eyes of insects and their association with habitats, and provide new information for further exploring the phylogenetic relationship between Cicadoidea and allies in Hemiptera.
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Division of larval instars of Wiebesia pumilae (Hymenoptera: Agaonidae)
BAO Tian-Tian, CHEN You-Ling, WU Wen-Shan, WU Ting-Ting, ZHANG Xing-Tan, CHOU Lien-Siang
Acta Entomologica Sinica    2020, 63 (12): 1508-1515.   DOI: 10.16380/j.kcxb.2020.12.009
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【Aim】 Ficus pumila. var. pumila is not only a key tree species of tropical and subtropical plant ecosystems, but also an important tree species of urban vertical greening. Wiebesia pumilae is the obligate pollinator of F. pumila var. pumila. Fig wasp larvae live in the gall of syconium for a long time, with special features of small body, long stadium, and difficulty for observation. The purpose of this study is to establish the classification criteria of larval instars of fig wasps, and to explore a reliable method for determining the number and stadia of larval instars of recessive insect larvae. 【Methods】 A total of 447 larvae of W.pumilae at different developmental stages were collected, and the morphological indicators including head capsule width, width of the 3rd segment, body length and girth were measured. The larval instars were determined by frequency distribution method, and the larval stadia were calculated by the population median stadium measurement method. 【Results】 The frequency distribution of larval head capsule width, width of the 3rd segment, body length and girth all showed five peaks, suggesting that there are five larval instars. Since the coefficient of variation in the width of the 3rd segment was greater than 20%, it was not suitable as the index of instar division. Regression analysis showed that the head capsule width, body length and girth were extremely significantly correlated with the larval instars (P<0.01). Because the coefficient of determination (R2) of regression curve for body length measurement and larval instars was the highest, the body length was the optimal measurement index of instars, and the regression equation was y=0.14e0.55x(P<0.01, R2=0.97). The stadia of the 1st-5th instar larva of W. pumilae was 429, 13.19, 6.27, 24.46 and 8.90 d, respectively, and the total larval stadium was 57.11 d. 【Conclusion】 This study determined the number and stadia of larval instars of W. pumilae and screened out the optimal age index of larval fig wasps, providing a foundation for studying fig-wasp co-evolution.
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Research progress of the influence of microorganisms on insect behavior
CHENG Dai-Feng, LI Hui-Jing, LU Yong-Yue
Acta Entomologica Sinica    2021, 64 (6): 743-756.   DOI: 10.16380/j.kcxb.2021.06.010
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 In the long evolutionary process, a variety of forms of interaction between microorganisms and insects have been formed. The wide distribution of microorganisms provides background conditions for their contact with insects and influence on insect behavior. To further explore the phenomenon and mechanism of microorganisms influencing insect behaviors, the research progress of the influence of microorganisms on insect behaviors was reviewed in this article. Host location and selection of insects can be influenced by chemical signal substances produced by microorganisms, and the synthesis of semiochemicals in insects and host plants can also be influenced by microorganisms. Microorganisms have also been found to play important roles in intraspecific and interspecific relationships of insects. Microorganisms can even influence the reproductive behavior of insects by altering, for example, insect sex pheromones. Besides, social and aggregation behaviors of insects are also found to be influenced by semiochemicals synthesized by microorganisms. Considering the current research status of the influence of microorganisms on insect behaviors, the following aspects are suggested to be further investigated: (1) How are semiochemicals that influence insect behavior synthesized in the processes of influencing insect behavior by microorganisms? (2) Do microorganisms involve more interspecies interaction in influencing insect behavior? (3) How do host insects acquire and maintain symbiotic microoganisms that influence insect behavior at certain stages?
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Changes in the mRNA levels and activities of carbohydrate metabolism-related enzymes in the diapause-destined, non-diapause-destined and immediately acid-treated eggs of the bivoltine silkworm (Bombyx mori)
XU Jin, JIANG Tao, XUE Peng, SHEN Guang-Sheng, HUANG Jing-Yi, ZHU Juan, WANG Mei-Xian, TANG Shun-Ming, SHEN Xing-Jia
Acta Entomologica Sinica    2020, 63 (12): 1431-1440.   DOI: 10.16380/j.kcxb.2020.12.001
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【Aim】 To clarify the relationship between carbohydrate metabolism and diapause during the earlyembryonicdevelopmentalstageinBombyx mori. 【Methods】 Theactivatedhibernating eggs of bivoltine B. mori strain Qiufeng were used as materials. Some were incubated under 17℃ in darkness and fed normally to produce the non-diapause-destined eggs (ND), and others were incubated under 25℃ in natural circadian rhythm and fed normally to produce the diapause-destined eggs (DD), part of which were further treated with HCl solution to produce the immediately acid-treated eggs (IA). Samples were collected at 0, 2, 6, 12, 24, 48, 72, and 96 h, respectively, after oviposition or acid treatment. The mRNA levels of genes of five carbohydrate metabolism-related enzymes including hexokinase (HK), phosphofructokinase (PFK), sorbitol dehydrogenase (SDH), phosphoenolpyruvate carboxykinase (PEPCK) and trehalase (TRE) in B. mori eggs were detected by qRT-PCR, and the activities of the five enzymes in B. mori eggs were assayed by ultraviolet visible spectrophotometry. 【Results】 The overall mRNA levels and activities of the key enzymes of glycolysis,hexokinase (BmHK) and phosphofructokinase (BmPFK), and the key enzymes of glycometabolism, sorbitol dehydrogenase (BmSDH-1) and trehalase (BmTRE), in ND and IA of B. mori were higher than those in DD, while the mRNA levels and activities of phosphoenolpyruvate carboxykinase (BmPEPCK) related to gluconeogenesis in DD were higher than those in ND and IA. 【Conclusion】 The results suggest that in the early embryonic developmental stage of DD, carbohydrate metabolism is mainly towards energy and material storage for diapause, while in ND and IA, carbohydrate metabolism is mainly towards material catabolism due to fast embryonic development. This study has preliminarily uncovered the relationship between carbohydrate metabolism and diapause of B. mori, being beneficial to better understand the molecular mechanisms of diapause of B. mori.
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Differences in the accumulation and allocation of nutrients in the long-winged and short-winged male adults of Velarifictorus aspersus (Orthoptera: Gryllidae)
ZENG Yang, ZHANG Bin, HE Yi-Yuan, ZHAO Xin, ZHU Dao-Hong
Acta Entomologica Sinica    2021, 64 (1): 1-9.   DOI: 10.16380/j.kcxb.2021.01.001
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 【Aim】 For wing polymorphic insects the energy investment of males in reproduction is different from that of females, and this difference may lead to changes in the physiological mechanism of the trade-off between flight and reproduction. To test this speculation, we investigated the accumulation and allocation of nutrients in wing-dimorphic male adults of Velarifictorus aspersus. 【Methods】 We compared the temporal changes in body weight between the long-winged (LW) and short-winged (SW) male adults of V. aspersus with similar head width within 7 d after emergence. The contents of protein, total lipids and glycogen in the whole body, and flight and reproductive organs in the wing-dimorphic male adults were determined by spectrophotometry. 【Results】 The body weight of the LW male adults of V. aspersus remained unchanged within the first 7 d after emergence, while that of the SW male adults increased significantly. The protein contents in the LW males and SW males on the day of adult emergence were 46.5±2.7 and 42.3±1.9 mg, respectively, showing significant difference between the two wing morphs. In addition, the protein content increased faster in the LW male adults than in the SW male adults thereafter. The contents of total lipids in the LW and SW males showed no significant difference on the day of adult emergence, but the contents of total lipids in the SW males were significantly higher than those in the LW males on the 5th and 7th days after adult emergence. Within 7 d after adult emergence, the contents of proteins and total lipids in flight muscles of the LW males were significantly higher than that of the SW males. On the day of adult emergence, there was no significant difference in the contents of proteins and total lipids between the accessory glands of the LW and SW males. However, the contents of proteins and total lipids in the accessory glands of the SW males were significantly higher than those of the LW males from the 3rd day after adult emergence. 【Conclusion】 The results suggest that the trade-off between flight and reproduction can influence the accumulation and allocation of nutrients in wing-dimorphic male adults of V. aspersus.
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Research progress of pheromone receptors in moths
CAO Song, LIU Yang, WANG Gui-Rong
Acta Entomologica Sinica    2020, 63 (12): 1546-1568.   DOI: 10.16380/j.kcxb.2020.12.013
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 As a vital element in male moth for reception of sex pheromone components emitted by the sex gland of the conspecific female moth, pheromone receptor (PR) determines the selectivity and specificity of male odorant receptor neuron (ORN) sensing sex pheromones. Since the first PR gene in moths was identified from Heliothis virescens, PR genes have been identified in more than 60 moth species with the development of high-throughput sequencing techniques combined with sequence homology analysis. Subsequent studies proved that unlike ordinary odorant receptor (OR) genes, PR genes in moths are relatively conserved in evolution, and they cluster into a unique group in the phylogenetic tree, forming the so-called traditional PR subfamily. The expression profile and in situ hybridization results demonstrate that PR genes are mainly specifically or biased expressed in male antennae, and in the studied moth species, PRs are restrictedly expressed in the long sensilla trichodea of male antennae. In recent years, the PRs of 30 moth species have been functionally characterized by using in vitro expression system, transgenic Drosophila and other methods. As an increasing number of PRs in moths have been identified and functionally studied, researchers found other PR clades separated with the traditional PR clade in moths, which also function to recognize moth pheromone components, giving us a new understanding of the evolutionary relationships of PRs in moths and the relationship between PR evolution and species differentiation. In this article, we reviewed the new research advances of PRs in moths from aspects including PR identification, expression patterns, functional characterization and evolution, and proposed the following important directions for further research: (1) Identifying and deorphanizing more PR genes in moths that do not use type I pheromone, to promote the understanding of the evolution of PR genes; (2) Characterizing the function of special PRs, to broaden our knowledge of the function of PRs; (3) Paying more attention to PRs in the new PR clades, especially for moth species whose PR genes belonging to the traditional PR clade have not been identified; (4) Studying the interactions between PR and other olfactory-related proteins, especially PBPs and SNMP1, to further understand how PR works; (5) Illuminating the structure of PR and Orco complex, to reveal the relationship between PR structure and function, and the relationship between the differentiation of PR function and the evolution of moth species; (6) Designing efficient and environmentally friendly measures to control moth pests based on the identified PRs.
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Dynamic distribution of histone H3 Ser10 phosphorylation during meiosis in spermatocytes of the silkworm, Bombyx mori
ZHANG Bing, QIU Reng, KAN Yun-Chao
Acta Entomologica Sinica    2021, 64 (3): 302-308.   DOI: 10.16380/j.kcxb.2021.03.002
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【Aim】 To explore the function of histone H3 Ser10 phosphorylation  (H3Ser10ph) during meiosis in spermatocytes of the silkworm, Bombyx mori. 【Methods】 The testis tissues of B. mori from the 4th instar larval stage to pupal stage weredissected and separated. The slides of testis tissues at different stages of meiosis in spermatocytes were prepared by encapsulation of acrylamide gel and then the localization characteristics of the H3Ser10ph antibody was detected by using immunofluorescence staining. 【Results】 The phosphorylation of histone H3 Ser10 occurred in the specific position of pachytene chromosomes during meiosis in eupyrene spermatocytes of B. mori. The signal of H3Ser10ph was gradually weakened in the diplotene, and no phosphorylation signal was detected in the chromosomes at the diakinesis. With the progress of cell cycle, phosphorylation signal began to increase, and reached the highest level at the metaphase of the first meiosis. When the cells entered the prometaphase II of meiosis, the signal of H3Ser10ph on the chromosome arm disappeared, and there was holo-shaped fluorescent signal of the H3Ser10ph antibody close to the spindle microtubulin attaching side. At the end of meiosis II, a weak H3Ser10ph signal remained in the specific position of chromatin. During apyrene spermatocyte meiosis, the signal of H3Ser10ph was aggregated uniformly along the whole chromosomes from metaphase I to telophase I, and the spindle microtubules were parallel to the equatorial plane at anaphase I. 【Conclusion】 Histone H3 Ser10 phosphorylation is correlated with the dynamic changes of chromatin in eupyrene spermatocytes and apyrene spermatocytes of the silkworm. 
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Feeding preference and adaptability of Spodoptera frugiperda (Lepidoptera: Noctuidae) on different wheat cultivars in relation to leaf biochemical contents#br#
LIU Huan, ZHANG Yong, CHEN Ju-Lian
Acta Entomologica Sinica    2021, 64 ( 2): 230-239.   DOI: 10.16380/j.kcxb.2021.02.010
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【Aim】 This study aims to determine the feeding preference and adaptability of the fall armyworm (FAW), Spodoptera frugiperda, on different wheat cultivars collected from the Huang-Huai-Hai wheat growing areas in North China, to explore the relationship between insect pest damage and wheat cultivars, and to clarify the resistance levels of different wheat cultivars to FAW, so as to provide a theoretical basis for the arrangement of insect resistant cultivars and integrated pest management in wheat fields. 【Methods】 The feeding preference of the 1st and 3rd instar larvae of FAW to 15 wheat cultivars was compared. Four wheat cultivars with high feeding preference and two wheat cultivars with low feeding preference for the 1st and 3rd instar larvae of FAW were screened out, the contents of biochemical components (total protein, soluble sugar, total phenol and tannin) in leaves of these six wheat cultivars were determined, the relationship between the feeding preference of the larvae and the contents of biochemical components in leaves of these six wheat cultivars was clarified by Pearson correlation analysis, and the adaptability of FAW on the six wheat cultivars was observed. 【Results】 The results revealed that the 1st and 3rd instar larvae of FAW preferred to feeding on Huaimai 46, Huaimai 33, KOK1679 and Tongmai 6, but not on Luyuan 502 or Yannong 21. The feeding preference of the 1st and 3rd instar larvae to leaves of different wheat cultivars was positively correlated with the total protein content and the soluble sugar content in wheat leaves, but negatively correlated with the total phenol content and the tannin content. The development of FAW was significantly different on leaves of different wheat cultivars, with the highest host suitability index and the shortest larval duration on Tongmai 6, and with the longest larval duration on Yannong 21. 【Conclusion】 The FAW showed different adaptability to different wheat cultivars, and Yannong 21 showed higher resistance to this insect pest. The FAW larvae prefer wheat leaves with higher contents of total protein and soluble sugar, while the feeding can be inhibited by high contents of total phenol and tannin in wheat leaves. It is speculated that the difference of resistance levels of different wheat cultivars to FAW is resulted from the coordination of nonpreference and antibiosis in wheat by the metabolism of nutrients and secondary substances.
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Identification of salivary gland proteins of nymphs and male and female adults of Sogatella furcifera (Hemiptera: Delphacidae)
LI Fei, YI Jin-Yu, LIU Yu-Di, HOU Mao-Lin
Acta Entomologica Sinica    2021, 64 (3): 281-301.   DOI: 10.16380/j.kcxb.2021.03.001
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 【Aim】 To identify and functionally annotate the salivary gland proteins of Sogatella furcifera and to explore their differences and correlation between different developmental stages and sexes. 【Methods】 Proteins were extracted from salivary gland tissues dissected from anesthetized nymphs, and male and female adults of S. furcifera, subjected to reductive alkylation and enzymolysis, and identified by liquid chromatography tandem mass spectrometry. Then, the salivary gland proteins were functionally annotated and classified by comparison with the Unigene protein database and KOG analysis. 【Results】There were 385, 168 and 82 salivary gland proteins specifically associated with nymphs, and male and female adults of S. furcifera, respectively. In total 319 salivary gland proteins were shared by nymphs and male adults, 60 by nymphs and female adults, and 60 by male and female adults. The KOG functional annotation showed the highest number of proteins associated with cellular processes and signaling, with 81, 22, and 70 proteins of this type specifically associated with nymphs, male adults, and female adults, respectively, and 19, 21 and 12 proteins of this type shared by nymphs and male adults, nymphs and female adults, and male and female adults, respectively, and these proteins are principally involved in post-translational modification, protein turnover, chaperones, intracellular trafficking, secretion and vesicular transport, and signal transduction.【Conclusion】 The salivary gland proteins of S. furcifera are quite active in signal transduction, which may be connected to its piercing and sucking damage to host plants.

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Genetic differentiation and phylogeography of the pierid butterfly Colias fieldii (Pieridae: Coliadinae) in China based on mitochondrial gene sequences(In English)
CHEN Ke-Ke, LUO A-Rong, WANG Yun-Liang, SI Cheng-Cai, HUANG Dun-Yuan, SU Cheng-Yong, HAO Jia-Sheng, ZHU Chao-Dong
Acta Entomologica Sinica    2020, 63 (12): 1525-1535.   DOI: 10.16380/j.kcxb.2020.12.011
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【Aim】This study aims to explore the genetic diversity, genetic differentiation, and phylogenetic relationships among populations of the pierid butterfly Colias fieldii in China, to infer their origin and divergence time, and to preliminarily clarify the causes of their spatiotemporally evolutionary history. 【Methods】 The four mitochondrial gene (COI, Cytb, NDI and ND5) sequences of 115 individuals from 23 geographic populations of C. fieldii in China collected in 2006-2018 were amplified by PCR and sequenced. The genetic diversity and genetic differentiation were analyzed using MEGA v.7.0, DnaSP v.5.0, Arlequin v.3.5.1 and other genetic analysis software. Using the closest relatives as the outgroups, the phylogenetic trees and haplotype median-joining network of C. fieldii were reconstructed with such analytical software as IQ-TREE, MrBayes v.3.1.2, Network v.4.6 and BEAST v.1.8.3, and the origin and divergence time of C. fieldii were estimated by using relaxed molecular dating method and calibrations of the previous studies. Based on the present biogeographic distribution of C. fieldii and the main earth environmental events since the Quaternary Period, the spatio-temporal pattern of its biogeographic distribution and the underlying earth environmental factors were tentatively inferred. 【Results】 The aligned sizes of mitochondrial gene segments of COI, Cytb, NDI and ND5 of C. fieldii populations are 648, 699, 393 and 777 bp, respectively, and the concatenated sequence of the four genes is 2 517 bp in length, which was shown to be significantly AT biased. In total, 18 haplotypes based on the four mitochondrial gene sequences were found in 115 individuals of 23 geographic populations of C. fieldii, with the haplotype diversity (Hd) of 0.677±0.048 and nucleotide diversity ) of 0.00066±0.00007 of the total population, showing a relatively high level of haplotype diversity and a low level of nucleotide diversity. The phylogenetic analysis showed that 18 haplotypes of C. fieldii populations were distinctly categorized into two large clades (clade I and II). Clade I included 13 haplotypes of populations from Shaanxi, Henan, Gansu, Anhui, Hubei, Sichuan, Qinghai, and some regions of Yunnan. Clade II included five haplotypes of populations from some regions of Yunnan and Tibet. The results of reconstructed haplotype median-joining network were generally consistent with those of the phylogenetic tree. AMOVA analysis indicated that a larger level of population differentiation (64.36%) occurred between the two haplotype clades and a subtle genetic differentiation (35.64%) existed within each haplotype clade of C. fieldii. The analysis results of neutrality tests and mismatch distribution indicated that populations with haplotypes in clade I did not experience a population expansion event, whereas those with haplotypes in clade II probably had a sudden demographic expansion at about 0.085 Ma in the late Pleistocene, a little earlier than the Last Glacial Maximum event, which may be caused by the warm and humid plateau climate in the interglacial period, the expansion of forest grassland and the decreased heavy rainfall on the core plateau. 【Conclusion】 The genetic differentiation of C. fieldii populations is correlated significantly with the geographical distance. In addition, we proposed that C. fieldii populations originated at about 0.48 Ma in southwestern areas of China (presently the Hengduan Mountains and adjacent areas), and began to diversify into two clades and later dispersed into other low-latitude areas due to the Quaternary glacial-interglacial cycle events, Southeast Asia monsoon and different habitat environments.
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Biological characteristics of Asobara japonica (Hymenoptera: Braconidae) parasitizing Drosophila melanogaster and the effects of its parasitization on the host growth and immune responses
ZHANG Xian, ZHOU Si-Cong, CHEN Jia-Ni, PANG Lan, ZHANG Qi-Chao, WANG Ying, SHI Min, CHEN Xue-Xin, HUANG Jian-Hua
Acta Entomologica Sinica    2021, 64 (1): 70-79.   DOI: 10.16380/j.kcxb.2021.01.008
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【Aim】 This study aims to investigate the biological characteristics of the parasitoid Asobara japonica parasitizing Drosophila melanogaster and to determine the effects of its parasitization on the host growth and immune responses. 【Methods】The developmental duration and morphological characterastics of A. japonica at different developmental stages, the parasitism rate and emergence rate of A. japonica after parasitization on the 2nd instar larvae of D.melanogaster, the changes of host pupation time and transcriptional levels of 15 important genes of host in different immune pathways (SPE, Toll, Myd88, Dif and Drosomycin involved in Toll pathway, PGRP-LE, PGRP-LC, imd, Relish and Diptericin involved in Imd pathway and Spn27A, MP2, yellow-f2, DoxA2 and PPOinvolved in PO pathway) in the parasitized 2nd instar larvae of D. melanogaster were investigated and analyzed by anatomical imaging and qRT-PCR technology. 【Results】 The developmental duration of egg stage, larval stage and pupal stage of A. japonica under the conditions of 25±1℃, 50%±1% relative humidity and photoperiod of 16L∶8D was 2.38±0.01, 5.36±0.07 and 8.30±0.04 d, respectively. After parasitizing the 2nd instar larvae of D. melanogaster, the parasitism rate and wasp emergence rate of A. japonica were 94.9%±4.0% and 64.3%±7.1%, respectively. Moreover, after being parasitized by A. japonica, the pupation time of D. melanogaster with 50% pupation rate was prolonged by 0.5 d, and the transcriptional levels of the antibacterial peptide genes Drosomycin and Diptericin were up-regulated significantly, whereas that of the prophenoloxidase gene PPOwas down-regulated as compared to the unparasitized control. 【Conclusion】 A. japonica can successfully and effectively parasitize D. melanogaster larvae through triggering host developmental delay and suppressing host melanization immune responses. The results provide some useful information for the massive release programs of the agriculturally important parasitoid species A. japonica. 
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Investigation and evaluation of insect diversity in Xingshan County, Three Gorges Reservoir Area, Hubei Province, central China
ZHENG Xiao-Xu, XIAO Neng-Wen, ZHAO Mu-Hua, WEN Dong, HE Shuai-Jie, LI Xue-Mei, YANG Feng-Lian, WU Gang
Acta Entomologica Sinica    2020, 63 (12): 1497-1507.   DOI: 10.16380/j.kcxb.2020.12.008
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【Aim】 The unique geographical environment and climatic conditions of Xingshan County in the Three Gorges Reservoir Area of Hubei Province, central China have created abundant insect resources. This study aims to investigate and evaluate the insect diversity in Xingshan County, Hubei Province, to explore the threat factors to insects and to put forward the suggestions for protecting the insect diversity in Xingshan County. 【Methods】 The Xingshan County was divided into 28 grids using GPS positioning system, and each was 10 km both in length and width. The diversity of insects in Xingshan County during the occurrence period (mid-June), outbreak period (mid-August) and overwintering period (mid-October) of insects were investigated and evaluated in 2017 using the line transect survey method. 【Results】 A total of 612 insect species of 136 families in 16 orders were collected and identified from Xingshan County, Hubei Province in 2017. The dominant orders are Lepidoptera (47.39%), Coleoptera (22.88%) and Hemiptera (11.44%). The low-altitude grids in Xingshan County had low insect diversity index and richness, while the high-altitude grids in Xingshan County had high insect diversity index and richness. The highest Shannon-Wiener diversity index (H′) and Margalef richness index (ds) of insects were found in Huangbo Plain (Longmenhe Forest Farm). 【Conclusion】 The dominant groups of insects in Xingshan County, Hubei Province are Lepidoptera, Coleoptera and Hemiptera. Habitats in the grids of such as woodland and woodland-shrubland mixture are more suitable for insects. The insect diversity index in the high-altitude habitats with rich vegetation (mainly woodlands and bushes) is significantly higher than that in the low-altitude habitats with a single habitat composition (mainly grasslands, underbushes, vegetable gardens and orchards). The threat factors to insect diversity in Xingshan County mainly include habitat destruction, climate change and pesticide abuse. We recommend that Longmenhe Forest Farm be listed as a key protection and research object based on the investigation of insect diversity in Xingshan County.
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Effects of silencing insulin like peptide (Ilp) genes on the wing and ovarian development and trehalose metabolism in Nilaparvata lugens (Hemiptera: Delphacidae)
YU Wei-Dong, LIU Yong-Kang, LUO Yu-Jia, HUANG Zhen, ZHOU Tai, YE Lin, TANG Bin, WANG Shi-Gui
Acta Entomologica Sinica    2021, 64 (4): 428-438.   DOI: 10.16380/j.kcxb.2021.04.002
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【Aim】 Insulin-like peptide (Ilp) is located in the upstream of insulin signaling pathway and plays a key role in regulating sugar metabolism. This study aims to explore the function of Ilp in regulating the trehalose metabolism balance in the brown planthopper, Nilaparvata lugens. 【Methods】 The expression of Ilp genes in the 5th instar nymphs of N. lugens was inhibited by RNAi technology, and the phenotype and ovarian development of the female adults of N. lugens were observed after RANi.The changes in the contents of trehalose, glycogen and glucose and the trehalase activity were determined by biochemical methods, and the changes in the expression levels of trehalase genes (TRE1-1, TRE1-2 and TRE2) and trehalose synthase genes (TPS1 and TPS2) in the 5th instar nymphs of N. lugens at 48 h and 72 h after RNAi were tested by qPCR. 【Results】 The dsRNA effectively inhibited the expression of Ilp genes and resulted in abnormal wing in N. lugens, and the injection of dsIlp3 and dsIlp1+dsIlp2+dsIlp3+dsIlp4 caused underdeveloped ovaries in the 2-day-old female adults of N. lugens. The glucose levels were significantly increased at 48 h after injection of dsIlp1-4 and dsIlp1+dsIlp2+dsIlp3+dsIlp4, respectively, and the glycogen levels were also significantly increased at 48 h after injection of dsIlp2, dsIlp3 and dsIlp4, respectively. The trehalose content was significantly increased at 48 h after injection of dsIlp3 and dsIlp4, respectively, while the trehalose content decreased significantly at 72 h after inhibition of Ilp2 and Ilp4, respectively, and increased significantly both at 48 h and 72 h after injection of dsIlp1+dsIlp2+dsIlp3+dsIlp4. In addition, the soluble trehalase activity increased significantly at 72 h after injection of dsIlp1+dsIlp2+dsIlp3+dsIlp4, while the membrane-bound trehalase activity decreased significantly at 72 h after injection of dsIlp3, dsIlp4 and dsIlp1+dsIlp2+dsIlp3+dsIlp4, respectively. The expression levels of TRE1-1, TRE1-2, TRE2, TPS1 and TPS2 were significantly decreased after injection of dsIlp1, dsIlp2 and dsIlp4, respectively. 【Conclusion】 Silencing of Ilp genes has an inhibitory effect on the development and reproduction of N. lugens, and can increase the contents of glucose and glycogen, downregulate the expression of genes of trehalase and trehalose synthase, and increase the soluble trehalase activity, thus regulating the trehalose metabolism in N. lugens.
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Expression, purification and characterization of the cuticular proteins TdCPR12611 and TdCPR7854 from Trypoxylus dichotomus (Coleoptera: Scarabaeidae)
YE Chang-Qing, BAO Han, LIU Tian, YANG Qing
Acta Entomologica Sinica    2021, 64 (1): 19-29.   DOI: 10.16380/j.kcxb.2021.01.003
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【Aim】 To explore the sequence characteristics and biochemical properties of cuticular proteins from Trypoxylus dichotomus. 【Methods】 RT-PCR was used to clone cuticular protein genes of T. dichotomus, and the structural features and phylogeny of cuticular proteins were analyzed by bioinformatics methods. Recombinant cuticular proteins of T. dichotomus were expressed in Escherichia coli expression system, and purified by metal-chelating affinity chromatography. In vitro binding experiments were performed to detect the binding ability of cuticular proteins of T. dichotomus with chitins including α-chitin, β-chitin, chitosan and colloidal chitin. Liquid-liquid phase separation (LLPS) was observed and determined. 【Results】 Two cuticular protein genes TdCPR12611 (GenBank accession no.: MT813021) and TdCPR7854 (GenBank accession no.: MT813022) of T. dichotomus were cloned and obtained. Phylogenetic analysis results showed that TdCPR12611 is closely related to OtCP-1 from Onthophagus taurus, while TdCPR7854 is closely related to OtCP-acp20-1, OtCP-acp20-2, and OtCP-acp20-3 from O. taurus, all of which belong to the CPR_RR-2 family. Recombinant TdCPR12611 and TdCPR7854 proteins were expressed by prokaryotic expression and purified. The two recombinant proteins had different binding abilities with four types of chitins, among which 41.4% of TdCPR12611 could bind with chitosan, while 62.3% of TdCPR7854 could bind with β-chitin. TdCPR12611 was predicted to have a relatively disordered structure and could spontaneously coacervate at room temperature to form liquid-liquid phase separation, while TdCPR7854 could not. 【Conclusion】 In this study we assayed and analyzed the sequence characteristics and chitin binding properties of two RR-2 cuticular proteins, TdCPR7854 and TdCPR12611 of T. dichotomus. The results not only deepen our understanding of insect cuticle assembly mechanism, but also provide ideas for the development of protein biomimetic materials.
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Electroantennographic and behavioral responses of female adults of Maladera orientalis (Coleoptera: Scarabaeidae) to peach tree volatiles
ZHANG Meng-Meng, CHEN Hong-Hao, WANG Wen-Kai, CHEN Li
Acta Entomologica Sinica    2020, 63 (12): 1482-1489.   DOI: 10.16380/j.kcxb.2020.12.006
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【Aim】 Maladera orientalis is an important garden pest in China. This study aims to identify peach tree volatile compounds attracting M. orientalis, so as to provide a theoretical basis for developing attractants of plant origin. 【Methods】 Leaf volatiles were collected from peach tree by dynamic headspace adsorption. The electrophysiologically active compounds for M. orientalis adults were identified from peach tree volatiles by gas chromatographymass spectrometry (GC-MS) and coupled gas chromatography electroantennogram detection (GC-EAD) techniques. The electrophysiological and behavioral responses of female adults of M. orientalis to the identified active compounds were tested by using electroantennography (EAG) and Y-tube olfactometer, respectively, using hexane as a control. 【Results】 Six active compounds were identified from peach tree volatiles, including E-2-hexenal, Z-3-hexenyl acetate, E-3-hexenol, Z-3-hexenol, methyl salicylate and an unknown compound. E-2-Hexenal triggered significantly greater EAG response in female adults of M. orientalis than the other four compounds at the doses of 1 and 10 μg. When the dose was increased to 100 μg, Z-3-hexenyl acetate triggered significantly lower EAG response in female adults of M. orientalis than the other four compounds. In behavioral response experiments, the female adults of M. orientalis were significantly attracted to E-3-hexenol and Z-3-hexenol at the dose of 100 μg. 【Conclusion】 Among peach tree volatile compounds, E-3-hexenol and Z-3-hexenol are significantly attractive to female adults of M. orientalis.
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Effects of 5 h LED light of different wavelengths in scotophase on the growth, development and reproduction of Spodoptera litura (Lepidoptera: Noctuidae)
ZHONG Chun-Lan, CHEN Su-Hong, ZHOU Xiao-Mei, WENG Ru-Yu, XIAO Guo-Quan, ZHENG Huan-Huan, TU Xiao-Yun
Acta Entomologica Sinica    2020, 63 (12): 1490-1496.   DOI: 10.16380/j.kcxb.2020.12.007
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【Aim】 To understand the effects of exposure to 5 h LED light of different wavelengths in scotophase on the growth, development and reproduction of Spodoptera litura. 【Methods】 Various developmental stages of S. litura in treatment groups were exposed to LED light of different wavelengths [red (620-625 nm), yellow (580-585 nm), blue (465-467 nm), green (520-523 nm) and white (composite) light], respectively, for 5 h in scotophase, expressed as the photoperiods of 14L∶5R∶5D, 14L∶5Y∶5D, 14L∶5B∶5D, 14L∶5G∶5D and 14L∶5W∶5D, respectively, with those in the control group cultured under a photoperiod of 14L∶10D, and the egg hatching rate, pupation rate, adult eclosion rate, developmental duration and adult longevity were observed and calculated. Pupae were weighted and the number of eggs laid per female counted. 【Results】 Exposure to LED light of different wavelengths for 5 h in scotophase had no significant effect on the egg hatching rate, pupation rate, adult eclosion rate of S. litura. The egg duration of the yellow light group (4.0 d) was significantly prolonged as compared to that of the control (3.0 d), while the pupal duration of various treatment groups was significantly shortened as compared to that of the control, and the male pupal duration was significantly longer than the female pupal duration. The pupal weight of various treatment groups was reduced as compared to that of the control, but there was no significant difference in the pupal weight between female and male. The adult longevity of various treatment groups was extended as compared to that of the control. The male adult longevity of the red light group (18.4 d) was significantly longer than the female adult longevity of this group (15.0 d), and the pre-oviposition period of the red light group (3.0 d) was the shortest and the number of eggs laid per female (1 346.0 eggs) of this group was the highest. 【Conclusion】 Under certain conditions, 5 h LED light of different wavelengths in scotophase mainly affects the pupal duration, pupal weight and adult longevity of S. litura, but the effects vary with the wavelength.
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Host adaptability of Bemisia tabaci on tomato plants with ToCV single infection and TYLCV&ToCV co-infection and the changes in the nutrient contents and defense responses of host plants
DING Tian-Bo, ZHOU Xue, YANG Nan, YANG Yang, TANG Yao, CHU Dong
Acta Entomologica Sinica    2021, 64 (3): 384-391.   DOI: 10.16380/j.kcxb.2021.03.010
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【Aim】 This study focuses on two important tomato viruses, tomato chlorosis virus (ToCV) and tomato yellow leaf curl virus (TYLCV). It aims to explore the effects of ToCV single infection and TYLCV&ToCV co-infection on the host adaptability of Bemisia tabaci MED, and to reveal the physiological mechanism from the perspective of host plant nutrient contents and defense responses. 【Methods】 After ToCV single infection andTYLCV&ToCV co-infection, the survival rates and number of eggs laid of B. tabaci MED adults on the infected tomato plants, and the contents of amino acids and total sugar in the infected tomato plants were detected. Additionally, the expression profiles of key genes related to jasmonic acid (JA) (FAD7 and PI II) and salicylic acid (SA) (NPR1 and PR1) signal pathways in tomato plants in response to ToCV single infection and TYLCV&ToCV co-infection were analyzed by RT-qPCR. 【Results】 The survival rate and number of eggs laid of B. tabaci MED adults feeding on the tomato plants infected by tomato viruses decreased significantly compared to those feeding on the healthy tomato plants. Feeding onthe TYLCV&ToCV coinfected tomato plants could reduce the survival rate and number of eggs laid of B. tabaci MED adults to the lowest level. The contents of total amino acids, fourteen types of hydrolyzed amino acids and total sugar in the TYLCV&ToCV co-infected tomato plants were significantly lower than those in the ToCV-singly infected tomato plants. The expression levels of FAD7 and PI II were significantly downregulated in tomato plants infected by ToCV and TYLCV&ToCV, and the expression levels of these two genes were the lowest in the TYLCV&ToCV co-infected tomato plants. However, the expression levels of NPR1 and PR1 were upregulated in tomato plants infected by ToCV and TYLCV&ToCV. The expression levels of NPR1 in the TYLCV&ToCV co-infected tomato plants and PR1 in the ToCV-singly infected tomato plants were all significantly higher than those in the healthy tomato plants. 【Conclusion】 Both ToCV single infection and TYLCV&ToCV co-infection could decrease the host adaptability of B. tabaci MED on tomato plants, and TYLCV&ToCV co-infection brought a more adverse effect on the survival of B. tabaci MED. Compared with the healthy tomato plants, the nutritional condition and defense system in ToCV-singly infected and TYLCV&ToCV co-infected tomato plants have obviously altered and are different between both. The results provide a basis for revealing the interaction between B. tabaci and plant viruses.

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Regulation of histone methylation modification in 20E signaling transduction detected by CRISPR/Cas9 system in Drosophila cells
ZHANG Wen-Hao, LONG Shi-Hui, NI Yi-Lu, ZHANG Jia-Hui, LI Sheng, LI Kang
Acta Entomologica Sinica    2021, 64 (5): 549-557.   DOI: 10.16380/j.kcxb.2021.05.001
Abstract211)      PDF(pc) (3507KB)(111)    PDF(mobile) (3507KB)(20)    Save
【Aim】 To screen and detect the regulation of histone methylation modification in 20-hydroxyecdysone (20E) signaling transduction by CRISPR/Cas9 knockout system in Drosophila melanogaster cells. 【Methods】 Histone methyltransferases of D. melanogaster and their modification sites were analyzed and summarized from Flybase website. After transfection of the constructed knockout vector pAc-sgRNA-Cas9 inserted with the sgRNA of five histone methyltransferase genes [trr, Mes-4, ash1, Su(var)3-9 and egg] into Kc cells of D. melanogaster, the gene mutation was detected by TA cloning and sequencing. Taking Trr inducing 20E primary response gene Br-C as a positive control, qRT-PCR and Western blotting were used to test the validity of pAc-sgRNA-Cas9 knockout system. The dual luciferase assay of 20E response element (EcRE) was used to determine whether trr, Mes-4, ash1, Su(var)3-9 and egg participate in 20E signaling transduction. 【Results】 Drosophila histone methylation modification occurs mainly on histone H3 lysine residues and less on H3 arginine residues. Besides, there was less methylation modification on histone H4. After the transfection of its knockout vector pAc-trr-sgRNA-Cas9 in Kc cells, trr was mutated successfully, the tri-methylation level of H3K4 was reduced, and the 20E-induced transcriptional level of its primary response gene Br-C was inhibited, indicating the validity of this knockout system. Besides trr, Mes-4 and egg knockouts also reduced the dual luciferase activity of EcRE. 【Conclusion】 The pAc-sgRNA-Cas9 knockout system inserted with sgRNA of target gene is effective and fast for gene mutation in Drosophila Kc cells. Using this knockout system, in addition to Trr, Mes-4 and egg were found to participate in the regulation of 20E signaling transduction via manipulating the activity of EcRE. This study lays the theoretical basis and work foundation for CRISPR/Cas9-mediated gene knockout in insect cells and further studying histone methylation modification involved in regulating 20E signaling transduction.
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Distribution and structure of wax glands in various body forms of the horned gall aphid, Schlechtendalia chinensis (Hemiptera: Aphididae)
WEI Hong-Yuan, FENG Guo-Rui, XU Xin, SHAO Shu-Xia, CHEN Xiao-Ming, YANG Zi-Xiang
Acta Entomologica Sinica    2021, 64 (4): 490-497.   DOI: 10.16380/j.kcxb.2021.04.008
Abstract210)      PDF(pc) (6961KB)(49)    PDF(mobile) (6961KB)(14)    Save
【Aim】 The horned gall aphid, Schlechtendalia chinensis, is the major productive species of Chinese gallnuts. S. chinensis exhibits complex life cycles, including alteration of sexual and asexual reproduction via host switch between Chinese sumac (Rhus chinensis) and mosses. There are six body forms, i.e., fundatrix, fundatrigeniae, fundatrispuriae, apterae, sexuaparae, and sexual forms, alternating inside and outside a gall to finish its whole life cycle. 【Methods】 Specimens of various body forms of S. chinensis were obtained from S. chinensis reared in the laboratory based on the original population of S. chinensis collected from the fields in Yanjin, Yunnan, southwestern China. The distribution, morphology, and structure of wax glands in S. chinensis were investigated using light microscope, electron microscope, and technique of ultrathin section. 【Results】 All body forms of S. chinensis except the 1st instar fundatrix possess wax glands on the dorsum, including 2 columns and 2 rows on the head, 4 columns and 1 row on the thorax, 6 columns and 8 rows on the abdomen, with a total of 56 wax glands. No wax glands appear on the dorsum of the 1st instar fundatrix (living outside a gall), but there are 2 columns and 8 rows of wax glands (in total 16 wax glands) on the dorsum of the 2nd instar fundatrix (living inside a gall). Each wax gland is composed of 2-22 polygonal depressions, and the number and morphology of depressions are obviously different among various body forms. The wax glands of fundatrigeniae, sexuaparae, and fundatrispuriae are the most complicated, and followed by apterae. Furthermore, the structure of wax glands in different parts of the sexual forms is significantly different: the wax gland structure of the dorsal plate and dorsolateral plate near the midline is simple, while that near the lateral line is complex. The wax gland structure of the 2nd instar fundatrix is simple as well. Corresponding to the distribution of wax glands, waxes appear on the body surface of all body forms except the 1st instar fundatrix. Fundatrigeniae, sexuaparae, and fundatrispuriae secrete a large amount of wax, followed by apterae. There is a small amount of wax on the body surface of the sexual forms and the 2nd instar fundatrix, whereas no wax exists on the body surface of the 1st instar fundatrix. 【Conclusion】 The number, arrangement, and development of wax glands on various body forms of S. chinensis are different. These differences are closely related to the microenvironments in which they live and the biological characteristics of each stage, being probably the long-term adaptation of the horned gall aphid to environmental conditions.
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Postembryonic development of the thoracic muscles of Apis mellifera ligustica (Hymenoptera: Apidae)
LI Zhao-Ying, SUN Jing, ZHANG Min, XIE Shu-Qin, TONG Li-Juan, WANG Shu-Wen
Acta Entomologica Sinica    2020, 63 (12): 1452-1460.   DOI: 10.16380/j.kcxb.2020.12.003
Abstract209)      PDF(pc) (2044KB)(52)    PDF(mobile) (2044KB)(11)    Save
【Aim】 This study aims to understand the postembryonic development of the thoracic muscles of the Italian honey bee, Apis mellifera ligustica, so as to provide some theoretical support for the study of animal muscle development and pathology, and to present a valuable insect model system to investigate muscle regeneration in vivo. 【Methods】 The postembryonic developmental process and structure characteristics of the thoracic muscles of A. m. ligustica were observed and compared by using hematoxylin and eosin (HE) staining and 5-bromo-2-deoxyuridine (BrdU) incorporation. 【Results】 The thoracic muscles of A. m. ligustica larvae are derived from myoblasts formed during embryonic development. Muscle development is accomplished through the fusion of fusion-competent myoblasts (FCMs) and founder cells (FCs), and the division of myoblasts and muscle cells. In the early stages of the development of A. m. ligustica pupa, most of the larval muscles degenerate and disappear. Adult thoracic muscles originate from three myoblast nests of two muscle cell groups, of which dorsal longitudinal muscles (DLMs) mainly originate from one myoblast nest of dorsal fan-shaped cell group, and other muscles originate from two myoblast nests of the ventral spindle cell group. Adult myoblasts of the myoblast nest continuously divide and proliferate. These divided cells expand and extend in specific directions. These cells become muscle fibers after the myofilaments are formed, and then multiple cells form a muscle bundle. These muscle bundles make up the entire muscles of the adult thorax. 【Conclusion】 The developmental modes of muscles of A. m. ligustica are different between its larvae and adults, and also different from those of Drosophila.
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Research progress of integrated coding of peripheral olfactory signals in the central nervous system of insects
LIU Wei, WANG Gui-Rong
Acta Entomologica Sinica    2020, 63 (12): 1536-1545.   DOI: 10.16380/j.kcxb.2020.12.012
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A sensitive and complex sense of smell is essential for the survival and reproduction of insects. Antennae are the main olfactory organs of insects and covered with a large number of different kinds of olfactory sensilla on their surface. Volatile chemicals in the environment are detected by these sensilla and subsequently transformed into electrical signals. Those signals are then transmitted through olfactory receptor neurons to the primary olfactory center of the brain, the antennal lobe, for the early processing. After processing, the signals are sent to the lateral horn which is thought to mediate innate behaviors as well as the mushroom body which is required for memory storage and retrieval. In this article, we reviewed the research progress of integrated coding of insect olfactory central system for signals from the peripheral olfactory system with focus on the Drosophila model and lepidopterans. Significant progress has been made in Drosophila melanogaster due to its advantages in genetic manipulation technology, including the propagation of information within the primary olfactory center and the subsequent processing in higher olfactory centers. Compared to Drosophila, the studies on olfactory coding of the central nervous system of other insects make tardy progress and are limited to recording and identification of the neurons in the antennal lobe of lepidopterans because of lacking genetic manipulation technology. The following aspects are suggested to be investigated in the future: (1) Olfactory coding in the lateral horn should be thoroughly investigated in the Drosophila model to further explain the neurological mechanism of olfactory behaviors; (2) Genetic tools should be developed and combined with two-photon imaging in none-model insects to elucidate the mechanism of perception, interaction, and behavioral consequence of the critical odorants.
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Morphology and distribution of antennal sensilla of Thrips hawaiiensis (Thysanoptera: Thripidae)
LIU Yu-Yan, WANG Liang, ZHANG Xiang-Qin, CHEN Yi-Xin, TIAN Hou-Jun, LIN Shuo, LI Heng, YU Yun, LIN Ling-Hong, ZHANG Jie, CHEN Yong, WEI Hui
Acta Entomologica Sinica    2021, 64 (4): 498-509.   DOI: 10.16380/j.kcxb.2021.04.009
Abstract207)      PDF(pc) (17457KB)(85)    PDF(mobile) (17457KB)(19)    Save

【Aim】 The aim of this study is to determine the type, morphology and distribution of antennal sensilla of Thrips hawaiiensis (Thysanoptera: Thripidae) at various developmental stages. 【Methods】 The antennal morphology and structure and the type, morphology and distribution of antennal sensilla of female and male adults, nymphs, pre-pupae and pupae of T. hawaiiensis were observed by using the scanning electron microscopy (SEM). 【Results】 The antenna of adult T. hawaiiensis is composed of scape, pedicel and a long flagellum with five flagellomeres (I-V). The mean length of antennae of female and male adults is 263.70±5.78 and 225.79±8.92 μm, respectively. The length of antennae increases with the growth of T. hawaiiensis. There are seven types of sensilla, i.e., Bhm bristles (BB), sensilla campaniformia (SCa), sensilla chaetica (SChI, SChII), sensilla trichodea (ST), sensilla basiconica (SBI, SBII, SBIII), sensilla coeloconica (SCo) and sensilla cavity (SCav), and two cuticular structures, i.e., microtrichia (mt) and cuticular denticles (cd), on the antenna of adults. The antenna of prepupae is conical, has no distinct segmentation and can move freely, with the mean length of 138.81±6.29 μm. The pupal antenna is cylindricalshaped pressing against the cephalo-thoracic back and immobile, and has no obvious segmentation, with a mean length of 213.07±6.30 μm. The antenna of the 1st and 2nd instar nymphs is composed of scape, pedicel and a flagellum with four flagellomeres (I-IV), with the mean length of 122.48±1.72 and 134.58±3.75 μm, respectively. There are eight types of sensilla [BB, SCa, SCh (SChI, SChII), SB (SBI, SBII), SCo, SCav, ST, and unusual sensillum (US)] and two cuticular structures [cd and ligulate structure (LS)] on the antenna of the 1st instar nymphs. There are seven types of sensilla [BB, SCa, SCh (SChI, SChII), ST, SB (SBI, SBII), SCo, and SCav] and one cuticular structure (cd) on the antenna of the 2nd instar nymphs. 【Conclusion】 In this study, the antenna and the morphology and distribution of antennal sensilla of T. hawaiiensis at various developmental stages were observed and comprehensively described, and the functions of antennal sensilla were inferred. The results lay a theoretical foundation for further research on the physiological functions of antennal sensilla of thrips.

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Identification and expression profiling of odorant receptor genes based on the transcriptomes of two sibling species of Eucryptorrhynchus (Coleoptera: Curculionidae) at different developmental stages
LU Yi, WANG Qian, WEN Jun-Bao
Acta Entomologica Sinica    2021, 64 (6): 655-665.   DOI: 10.16380/j.kcxb.2021.06.001
Abstract206)      PDF(pc) (4365KB)(160)    PDF(mobile) (4365KB)(33)    Save
【Aim】 To complement the odorant receptor (OR) information of two sibling species of Eucryptorrhynchus, E. scrobiculatus and E. brandti, and to provide theoretical basis for subsequent functional studies by the identification and expression profile analysis of OR genes based on the transcriptome data of these two weevil species at different developmental stages. 【Methods】 The OR gene sequences were screened from the transcriptome databases of different developmental stages of E. scrobiculatus (larva, pupa and adult) and E. brandti (egg, larva, pupa and adult). Interspecific and intraspecific sequence alignments of the screened ORs were performed based on the antennal transcriptome data of the two weevil species. Phylogenetic analysis of the newly identified OR genes was performed using the maximum likelihood method. Expression abundance analysis was performed according to FPKM (fragments per kilobase per million mapped fragments) values of the newly identified OR genes in the transcriptomes of two weevil species at different developmental stages. The expression profiles of the newly identified OR genes at different developmental stages and in different adult tissues of E. scrobiculatus and E. brandti were detected by qPCR. 【Results】 Six and four OR genes with complete open reading frames were identified from the transcriptome databases of different developmental stages of E. scrobiculatus and E. brandti, respectively. According to the sequence alignment results with the antennal transcriptome data, four (EscrOR50-53) and two (EbraOR46-47) OR genes were newly identified from E. scrobiculatus and E. brandti, respectively. We also found a pair of potential homologous genes, EscrOR53 and EbraOR45, between the two weevil species. The phylogenetic tree indicated that the six newly identified ORs belong to the subfamilies 2B and 7 of coleopteran ORs. According to the FPKM data of the transcriptomes of different developmental stages of the two weevil species and the spatiotemporal expression profiles detected by qPCR, three of the newly identified OR genes of E. scrobiculatus and the two newly identified OR genes of E. brandti were all highly expressed in non-olfactory tissues of adults, and also expressed in egg, larval or pupal stages. 【Conclusion】 In this study, the OR genes and their spatiotemporal expression profiles in the transcriptomes of different developmental stages of E. scrobiculatus and E. brandti have been clarified, and it is found that non-olfactory functional ORs possibly exist in the reproductive organs, suggesting that they may play a role in the early stage of development.
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Effects of RNAi-mediated suppression of Br-Z2/3 gene on the expression of downstream apoptotic genes and pupation in the diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae)
HU Xiao-Han, TIAN Su-Fen, LI Ya-Qing, LIN Shuo, CHEN Yi-Xin, WEI Hui, GU Xiao-Jun, HUANG Jing-Fei, WANG Xi-Ying, LI Zhi-Hua
Acta Entomologica Sinica    2021, 64 ( 2): 141-148.   DOI: 10.16380/j.kcxb.2021.02.001
Abstract206)      PDF(pc) (3456KB)(163)    PDF(mobile) (3456KB)(49)    Save
【Aim】 This study aims to establish an in vitro prokaryotic expression system for synthesis of the dsRNA of pupa-specific expression gene Br-Z2/3 from the diamondback moth, Plutella xylostella, and to find out the effects of RNAi-mediated suppression of Br-Z2/3 on the expression of Br-Z2/3 and apoptotic genes and the pupation of the moth. 【Methods】 The L4440-Br-Z2/3 recombinant vector was constructed and transformed into competent cells of Escherichia coli HT115. The dsRNA of Br-Z2/3 was obtained after induction with IPTG, and RNAi was carried out by microinjecting Br-Z2/3 dsRNA into the 4th instar larvae of P. xylostella. At 12 and 24 h after RNAi of Br-Z2/3, the expression levels of Br-Z2/3 and its downstream apoptotic genes, reaper, caspase-9 and Gadd45g, were detected through qPCR. Simultaneously, the pupation rate, average pupation time, rate of deformed pupae and larval mortality of the 4th instar larvae of P. xylostella after RNAi of Br-Z2/3 were observed and recorded. 【Results】 The dsRNA of Br-Z2/3 was successfully expressed in the prokaryotic system. The qPCR results showed that the expression levels of Br-Z2/3 and the related apoptotic gene reaper decreased significantly, but those of caspase-9 and Gadd45g increased significantly after RNAi of Br-Z2/3 in the 4th instar larvae of P. xylostella. Compared with the control group, the dsBr-Z2/3-injected group showed significantly lower pupation rate with delayed pupation peak, and had significantly higher mortality and higher rate of deformed pupae. 【Conclusion】 An in vitro prokaryotic expression system for synthesis of the dsRNA of Br-Z2/3 from P. xylostella was successfully established. RNAi of Br-Z2/3 by microinjection demonstrates that Br-Z2/3 plays a crucial role in the pupation of P. xylostella.
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Analysis of the functions of allatostatin genes in the reproductive diapause preparation of Colaphellus bowringi (Coleoptera: Chrysomelidae)
TIAN Zhong, LIU Xi, ZHU Li, LIU Wen, ZHU Fen, WANG Xiao-Ping
Acta Entomologica Sinica    2021, 64 (1): 30-40.   DOI: 10.16380/j.kcxb.2021.01.004
Abstract204)      PDF(pc) (4340KB)(83)    PDF(mobile) (4340KB)(8)    Save
【Aim】 This study aims to clarify the molecular characteristics and expression difference of allatotropin (AT) and allatostatin (AST) genes during the reproduction and diapause of Colaphellus bowringi, so as to further reveal the functions of these genes in the reproductive diapause preparation of C. bowringi. 【Methods】 We identified the sequences of CbAT and CbASTs in C. bowringi based on the previously constructed transcriptome database, cloned their ORFs and carried out the bioinformatics analysis of their sequences. Then, we assayed the expression patterns of CbAT and CbASTs in the heads of the diapause-destined and non-diapause-destined female pupae and female adults via RT-PCR. RNAi experiments of CbAST-B, CbAST-C and CbAST-B+CbAST-C were done in the diapause-destined 2-day-old female pupae, and the expression levels of juvenile hormone (JH) signaling genes and vitellogenin genes in 4-day-old female adults were detected. 【Results】 One allatotropin gene CbAT (GenBank accession no.: MT977128) and two allatostatin genes CbAST-B (GenBank accession no.: MT977126) and CbAST-C (GenBank accession no.: MT977127) of C. bowringi were identified and cloned, with the ORFs of 408, 600 and 303 bp, respectively. By amino acid sequence alignment and phylogenetic analysis, we found that CbAT, CbAST-B and CbAST-C respectively cluster into the same branch with homologous proteins of Coleoptera insects, showing high conservation in the evolution. The qRT-PCR assay indicated that CbAT had no significant expression differences in the female heads between the diapause-destined and non-diapause-destined individuals from the 2-day-old pupal stage to the 4-day-old adult stage. CbAST-B and CbAST-C were significantly highly expressed in the heads of diapause-destined female adult since the 1- and 2-day-old, respectively, and the differential expression state between the diapause-destined and non-diapause-destined individuals lasted until the end of diapause preparation period. After knocking down CbAST-B, CbAST-C and CbAST-B+CbAST-C in the diapause-destined 2-day-old female pupae, the expression of CbAST-B and CbAST-C in the heads of 4-day-old female adults was significantly suppressed with the RNAi efficiency over 70%. Further study revealed that the expression of JH biosynthesis genes AACT, FPPS and JHAMT, in the head without antenna, JH-responsive genes Kr-h1 and JHE1 in the fat body, and vitellogenin genes Vg1 and Vg2 in the fat body was all significantly up-regulated in the treatment groups of dsCbAST-B, dsCbAST-C and dsCbAST-B+dsCbAST-C【Conclusion】 CbAT may be not a key regulator mediating the differences of JH signaling between the preoviposition period and diapause preparation phase in C. bowringi. However, CbAST-B and CbAST-C suppress the synthesis of JH and further down-regulate the expression of Vg genes in C. bowringi during the reproductive diapause preparation phase, thereby promoting the occurrence of diapause. This study further reveals an upstream regulation mechanism for the JH signaling in insects during the reproductive diapause preparation phase, being helpful to understand the seasonal adaptation strategy of insects and providing new guidance and reference for potential target genes for pest control.
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Immunolocalization and ligand binding characteristics of the odorant binding protein AplaOBP3 of Agrilus planipennis(Coleoptera: Buprestidae)
SONG Xuan, WANG Ze-Hua, SHAN Shuang, ZHANG Yong-Jun, WANG Shan-Ning
Acta Entomologica Sinica    2021, 64 (1): 10-18.   DOI: 10.16380/j.kcxb.2021.01.002
Abstract202)      PDF(pc) (12948KB)(87)    PDF(mobile) (12948KB)(22)    Save
【Aim】 The aim of this study is to explore the localization of the odorant binding protein AplaOBP3 in antennae of Agrilus planipennis and its ligand binding characteristics, and to compare the function of AplaOBP3 with the reported AplaOBP1 and AplaOBP2. 【Methods】 The recombinant AplaOBP3 of A. planipennis was expressed in prokaryotic expression system. The polyclonal antibody against AplaOBP3 was prepared and detected by Western blot assay. Immunocytochemical technique was used to localize the expression of AplaOBP3 in antennal sensilla of A. planipennis. Its binding characteristics to 58 compounds were determined by fluorescence competitive binding assay, and compared with those of AplaOBP1 and AplaOBP2 previously reported. 【Results】 The recombinant protein AplaOBP3 was successfully expressed in prokaryotic expression system. The immunolocalization results showed that AplaOBP3 was expressed in the antennal sensilla basiconica type I. The results of fluorescence competitive binding assay showed that AplaOBP3 had strong binding capabilities with trans-2-hexenal, benzaldehyde, 4′-ethylacetophenone, β-ionone and ocimene, with the dissociation constant KD values of 6.20, 4.03, 5.24, 1.72 and 4.83 μmol/L, respectively. AplaOBP3 and AplaOBP2 had similar expression and ligand binding properties. 【Conclusion】 AplaOBP3 and AplaOBP2 have similar expression and ligand binding properties, and both may be involved in olfactory perception together and play a role in host localization in A. planipennis.
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Parasitism and development performance of bisexual and thelytokous lines of Trichogramma dendrolimi (Hymenoptera: Trichogrammatidae) on eggs of different strains of Antheraea pernyi (Lepidoptera: Saturniidae)
ZHOU Jin-Cheng, HE Yue, ZHAO Qian, DONG Hui
Acta Entomologica Sinica    2021, 64 (1): 80-89.   DOI: 10.16380/j.kcxb.2021.01.009
Abstract201)      PDF(pc) (1781KB)(53)    PDF(mobile) (1781KB)(7)    Save
【Aim】 To determine the parasitism and development performance of bisexual and thelytokous lines of Trichogramma dendrolimi on eggs of different strains of Antheraea pernyi, so as to provide the basis for the mass-rearing of richogrammatid parasitoids using A. pernyi eggs as the alternative host. 【Methods】 The biological parameters including parasitism rate, brood size of offspring (number of offspring wasps emerged from a single brood), and proportion of female offspring of bisexual and thelytokous lines of T. dendrolimi on eggs of six strains of A. pernyi [Kangda (KD), Dasi (DS), Gaoxin (GX), 988 (NEE), Qingda (QD), and Teda (TD)] were detected, and the quality parameters of eggs of the six strains of A. pernyi (wet weight per egg, protein content, total carbohydrate content and triglyceride content) were determined. The correlation between the quality parameters of A. pernyi eggs and the biological parameters of T. dendrolimi was detected by principal component analysis. 【Results】 The body size of offspring of the bisexual wasps of T. dendrolimi developed on eggs of the NEE strain of A. pernyi were significantly higher than those developed on eggs of the other strains of A. pernyi except for the KD and QD strains. The parasitism rate, brood size of offspring, and proportion of female offspring of the bisexual wasps of T. dendrolimi were not significantly influenced by the strain of A. pernyi. The brood size of offspring of thelytokous wasps of T. dendrolimi developed on eggs of the KD strain of A. pernyi was the highest, being significantly higher than those developed on eggs of the NEE and QD strains of A. pernyi, but without significant difference with those developed on eggs of the DS, GX, and TD strains of A. pernyi. The parasitism rate and brood size of offspring of the thelytokous wasps of T. dendrolimi were not significantly influenced by the strain of A. pernyi. The protein content in eggs of the TD strain of A. pernyi was the highest, significantly higher than those in eggs of the other strains of A. pernyi except for the GX strain. The total carbohydrate content in eggs of the KD strain of A. pernyi and the triglyceride content in eggs of the QD strain of A. pernyi were the highest, significantly higher than those in eggs of the other strains, respectively. The principal component analysis showed that the parasitism rate of bisexual wasps of T. dendrolimi, and the protein content and triglyceride content in A. pernyi eggs were negatively correlated with the total carbohydrate content in A. pernyi eggs. The body size of offspring of the bisexual wasps of T. dendrolimi was negatively correlated with the wet weight per egg of A. pernyi. The parasitism rate of the thelytokous wasps of T. dendrolimi and the triglyceride content in A. pernyi eggs were negatively correlated with the brood size of offspring of T. dendrolimi, the total carbohydrate content, and the wet weight per egg of A. pernyi. The body size of offspring of thelytokous wasps of T. dendrolimi and the total carbohydrate content in A. pernyi eggs were negatively correlated with the protein content in A. pernyi eggs. 【Conclusion】 This study determined the suitable strain of A. pernyi eggs for the mass-rearing procedure of bisexual and thelytokous lines of T. dendrolimi and revealed the relationship between the nutritive indexes of A. pernyi eggs and the biological parameters of T. dendrolimi. The results provide basic data for improving mass-rearing of T. dendrolimi.
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Diversity and sequence characteristics of the OCT family genes in Anopheles sinensis (Diptera: Culicidae)
LEI Dan, YAN Zhen-Tian, ZHANG Xiao-Xiao, CHEN Bin
Acta Entomologica Sinica    2021, 64 (3): 337-347.   DOI: 10.16380/j.kcxb.2021.03.006
Abstract200)      PDF(pc) (7078KB)(60)    PDF(mobile) (7078KB)(7)    Save

 【Aim】 To identify, classify and name genes of the organic cation transporter (OCT) family of Anopheles sinensis at the whole-genome level, so as to provide an information frame for the OCT genes of insects. 【Methods】 The amino acid sequences encoded by OCT genes in An. gambiae, Drosophila melanogaster and Caenorhabditis elegans were downloaded from NCBI, VectorBase and EMBL databases, and used as queries to search for the OCT genes in An. sinensis genome using the local Blast program based on the genome and transcriptome sequencing data of An. sinensis. The characteristics of these OCT genes in An. sinensis, including the gene structure, Scaffold location and conservative motifs, were predicted using bioinformatics methods. The phylogenetic tree of OCT genes of An. sinensis was constructed by the maximum likelihood method. 【Results】 The An. sinensis genome contains 54 OCT genes, which are divided into three subfamilies, OCTA, OCTB and OCTC, with 33, 15 and 6 genes, respectively. The encoded OCT amino acid sequences, except for AsOCTA20 and AsOCTB2, have MFS_1 and Sugar_tr transmembrane domains (TMDs), and most of the OCT amino acid sequences have 12 TMDs. All the OCT amino acid sequences have three conserved motifs, GRK-(PT)-VL, PES-(APVS) and EQFPT-(VI)-RN, and each subfamily has its own conservative motifs. Four pairs of genes (AsOCTB4a and AsOCTB4b, AsOCTA10a and AsOCTA10b, AsOCTA19a and AsOCTA19b, and AsOCTA23a and AsOCTA23b) are derived from gene duplication events. AsOCTC genes are relatively primitive, while AsOCTB genes are relatively evolved, both forming an obvious monophyly. AsOCTA genes are between AsOCTC genes and AsOCTB genes in evolution, with more complicated evolutionary relationship. 【Conclusion】 This study enriches the genome data of An. sinensis, and lays the foundation for the subsequent research of the functions of OCT genes in An. sinensis, especially their functions in insecticide resistance mechanism.

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Advances and perspectives of epigenetic regulation of insect diapause
AN Hao-Min, LIU Wen, WANG Xiao-Ping
Acta Entomologica Sinica    2021, 64 (4): 510-522.   DOI: 10.16380/j.kcxb.2021.04.010
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 Diapause is a strategy for insect to avoid harsh environment, and has a great significance for continuation of insect population. In particular, facultative diapause can be affected by periodic seasonal changes, in which epigenetics may play critical roles. Epigenetics refers to heritable variations independent of DNA sequence, including various modifications at DNA, RNA, protein and chromatin levels, and may be involved in development plasticity. The research of epigenetic regulation of insect diapause mainly focuses on two aspects: one is how epigenetic regulation respond to environmental signals, and the other is how environmental signal induces epigenetic regulation in insect diapause. Although it has been reported that DNA methylation can respond to photoperiodic signal and histone acetylation can be coupled with endocrine systems, the detail mechanisms of epigenetic regulations of insect diapause, however, are not completely revealed. Regulations of diapause induced by epigenetics have been reported in multiple types of insect diapause. For the same diapause process, there may be co-regulation between different epigenetic mechanisms. However, how this synergy responds to environmental signals and how it precisely regulates insect diapause are still unknown. In conclusion, the previous research only indicated the possibility of epigenetic regulation in insect diapause, but the molecular mechanisms are scarcely known and need further study. Especially the following aspects might be critical in the future research: the molecular mechanism of epigenetics responding to diapause-induced environmental signals, the molecular mechanism of epigenetics coupled with endocrine regulation, the cell signal transduction of epigenetic regulation, and the synergetic regulation of epigenetics in insect diapause.
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Construction and annotation of the full-length transcriptome of Nosema ceranae based on the third-generation nanopore sequencing technology
CHEN Hua-Zhi, DU Yu, FAN Xiao-Xue, ZHU Zhi-Wei, JIANG Hai-Bin, WANG Jie, FAN Yuan-Chan, XIONG Cui-Ling, ZHENG Yan-Zhen, FU Zhong-Min, XU Guo-Jun, CHEN Da-Fu, GUO Rui
Acta Entomologica Sinica    2020, 63 (12): 1461-1472.   DOI: 10.16380/j.kcxb.2020.12.004
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【Aim】 This study aims to assemble and annotate a high-quality full-length transcriptome of Nosema ceranae using Oxford Nanopore sequencing technology. 【Methods】 The transcriptome of clean spores of N. ceranae was sequenced using Nanopore PromethION system. Full-length transcripts were identified by recognizing primers at both ends of every clean read. Full-length transcripts were aligned to Nr, Swiss-Prot, KOG, eggNOG, Pfam, GO and KEGG databases to gain the corresponding annotations. Protein domain analysis methods including CPC, CNCI, CPAT and Pfam were used to predict long noncoding RNAs (lncRNAs), and the intersection was determined to be high-reliability lncRNAs. The expression level of each full-length transcript was calculated using CPM (counts per million) method. 【Results】 A total of 6 988 795 raw reads were obtained by Nanopore PromethION sequencing system, and 6 953 469 clean reads were gained after quality control, including 5 143 999 full-length transcripts. Besides, 10 243 non-redundant fulllength transcripts were identified, with the N50, the average length and the maximum length of 1 042, 894 and 4 855 bp, respectively. Furthermore, 9 342, 4 038, 4 283, 2 569, 4 859 and 3 450 full-length transcripts were annotated to Nr, KOG, eggNOG, Pfam, GO and KEGG, respectively. Additionally, the majority of full-length transcripts were annotated to N. ceranae, Nosema apis and Nosema bombycis. Totally, 87 high-reliability lncRNAs were identified, including 49 sense lncRNAs, 25 antisense lncRNAs and 13 intergenic lncRNAs. The sequencing depth in this study was enough to detect all expressed fulllength transcripts with the expression level (CPM) ranging from 0.1 to more than 10 000. 【Conclusion】 The high-quality full-length transcriptome of N. ceranae was constructed and annotated in this study, laying a key foundation for comparative transcriptome analysis, investigation of alternative splicing and alternative adenylation of transcripts, identification of simple sequence repeat (SSR) loci, optimization of gene structure, and full-length sequence cloning and functional study of genes.
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Cloning, prokaryotic expression and tissue expression analysis of the silk gland transcription factor gene AaSGF-1 of Antheraea assama (Lepidoptera: Saturniidae)
LI Qiong-Yan, CHEN An-Li, XUN Li-Jie, LIU Zeng-Hu, LIAO Peng-Fei, YANG Wei-Ke, DONG Zhan-Peng
Acta Entomologica Sinica    2021, 64 (1): 41-50.   DOI: 10.16380/j.kcxb.2021.01.005
Abstract194)      PDF(pc) (4602KB)(82)    PDF(mobile) (4602KB)(9)    Save
【Aim】 This study aims to clone the silk gland transcription factor gene AaSGF-1 from Antheraea assama, to analyze its sequence features and expression pattern, and to obtain the polyclonal antibody, so as to lay a basis for further studying the function of this gene. 【Methods】 The cDNA sequence of AaSGF-1 was cloned from the silk gland of A. assama by RT-PCR and RACE techniques and subjected to bioinformatical analysis. qPCR was employed to analyze the expression profile of this gene in different tissues (head, midgut, fat body, silk gland, hemolymph, and cuticle) of the day-4 5th instar larvae of A. assama. Prokaryotic expression plasmid was constructed and the fusion protein AaSGF-1 was expressed in Escherichia coli BL21. The polyclonal antibody with high serum titer was prepared using the purified fusion protein to immunize New Zealand rabbit. The protein level of AaSGF-1 in the silk gland and cuticle of the newly hatched larva and the silk gland of the 4th instar larva of A. assama was detected by immunofluorescence technique. 【Results】 The cDNA sequence of AaSGF-1 (GenBank accession no.: MK889510.1) of A. assama was cloned. The ORF of AaSGF-1 is 1 050 bp in length, encoding a polypeptide of 349 amino acids with the molecular weight of 38.8 kD and the isoelectric point (pI) of 8.74. The qPCR analysis results showed that AaSGF-1 was highly expressed in the silk gland of the 5th instar larvae of A. assama, especially in the posterior silk gland, but hardly expressed in other tissues. Immunofluorescence assay showed that AaSGF-1 was expressed in silk glands of the newly hatched larva and 4th instar larva of A. assama. 【Conclusion】 In this study AaSGF-1 was expressed by prokaryotic expression system, the polyclonal antibody was prepared, and AaSGF-1 was confirmed to be highly expressed in the silk gland of A. assama larva, providing a basis for further studying its roles in silk gland development and silk protein synthesis in A. assama.
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Expression of the fungal-resistance factor BmSPI39 in Bombyx mori in response to Beauveria bassiana invasion
LI You-Shan, LU Qing-Jun, YANG Xi, ZHANG Jie, LUO Zhu-Xing, XIA Qing-You, ZHAO Ping
Acta Entomologica Sinica    2021, 64 (1): 61-69.   DOI: 10.16380/j.kcxb.2021.01.007
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【Aim】 To prepare polyclonal antibody of the fungal-resistance factor BmSPI39 in Bombyx mori and to analyze its expression in response to Beauveria bassiana invasion. 【Methods】 Prokaryotic expression and immobilized nickel ion affinity chromatography were used to obtain the highly-purified recombinant BmSPI39 protein. The inhibitory activity of the recombinant BmSPI39 protein to Bacillus subtilis protease A was detected by in-gel activity staining. Polyclonal antibodies to BmSPI39 were prepared by immunizing New Zealand white rabbit with the recombinant BmSPI39 protein. Based on the open database SilkDB 3.0 of B. mori, the spatiotemporal expression characteristics of BmSPI39 in the immune-related tissues, integument, midgut and fat body, of B. mori from the day-3 4th instar larva to 1 day-old pupa were analyzed. And the expression of BmSPI39 in response to B. bassiana invasion in the integument of B. mori larvae and pupae at different time after B. bassiana infection was assayed using the prepared BmSPI39 antibodies by Western blot. 【Results】 The results of in-gel activity staining showed that the recombinant BmSPI39 protein could strongly inhibit the activity of subtilisin A. Enzyme-linked immunosorbent assay (ELISA) showed that the antibody titer of BmSPI39 antiserum reached 1∶128 000. The results of Western blot and in-gel activity staining showed that the purified BmSPI39 antibody could effectively bind to the BmSPI39 antigen protein in different multimerization states. The heat map analysis of the expression data of BmSPI39 showed that BmSPI39 was expressed in the integument of B. mori at the wandering stage and the pre-pupal stage, with a higher expression level in the fat body at the pre-pupal stage, but not in the midgut at all the tested developmental stages. The results of Western blot showed that the expression of BmSPI39 in the integument of B. mori larvae was up-regulated at 84 h and down-regulated at 108 and 120 h after B. bassiana infection. 【Conclusion】 Polyclonal antibody to BmSPI39 was successfully prepared in this study. The expression of BmSPI39 protein can respond to B. bassiana invasion and may be involved in the process of resisting fungal invasion in the pupal stage of B. mori.
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Resistance selection and biological characteristics of Neoseiulus barkeri (Acari: Phytoseiidae) to chlorpyrifos
WANG Li, CHENG Ming-Ming, FU Yun-Mei, FANG Yun-Hong, WEI Zhi-Tang, CHENG Lu-Yan, LEI Shuang, DING Li-Li, YU Shi-Jiang, CONG Lin, RAN Chun
Acta Entomologica Sinica    2021, 64 (3): 374-383.   DOI: 10.16380/j.kcxb.2021.03.009
Abstract191)      PDF(pc) (1652KB)(32)    PDF(mobile) (1652KB)(10)    Save

 【Aim】 To clarify the effects of acquired chlorpyrifos resistance in the predatory mite Neoseiulus barkeri on its biological characteristics, so as to provide a theoretical basis for its field application. 【Methods】 The residual film method was adopted to test the toxicity of chlorpyrifos to N. barkeri in the laboratory, and the lethal medium concentration (LC50) was chosen as the selection pressure. Life table was used to assess the effects of acquired chlorpyrifos resistance on the relative fitness (Rf)of N. barkeri. In addition, Holling type II model was used to analyze the differences in the predation functional response of the chlorpyrifos-susceptible and resistant strains of N. barkeri to eggs and female adults of Panonychus citri at different temperatures.【Results】 A chlorpyrifos-resistant strain of N. barkeri with the resistance ratio of 34.77-fold was obtained after selection of 21 generations. The acquired resistance had no significant effects on the developmental duration, proportion of female offspring and predation functional response of N. barkeri, but shortened the female adult longevity and oviposition period. The susceptible and resistant strains of N. barkeri could not complete the whole generation at 15℃. The fecundity (number of eggs laid per female) of the susceptible and resistant strains of N. barkeri was the highest at 25℃, and the fecundity was significantly higher for the susceptible strain (41.64±1.04 eggs/female) than for the resistant strain (38.80±0.93 eggs/female). Likewise the oviposition periods of the susceptible and resistant strains of N. barkeri were the longest at 25℃, and the oviposition period was significantly longer for the susceptible strain (24.82±1.50 d) than for the resistant strain (21.34±1.26 d). Further, the shortest developmental duration of the susceptible and resistant strains of N. barkeri was recorded at 30℃, being 6.62±0.23 d for the susceptible strain and 6.53±0.13 d for the resistant strain. Meanwhile, the strongest predation ability of the susceptible and resistant strains of N. barkeri to eggs or female adults of P. citri was recorded at 30℃, with the daily maximum predation amounts of the susceptible and resistant strains of N. barkeri to P. citri eggs being 156.25 and 140.85, respectively, and to female adults of P. citri being 23.10 and 22.32, respectively. The relative fitness (Rf) of the resistant strain of N. barkeri was lower than that of the susceptible strain at 25 and 30℃, but slightly higher than that of the susceptible strain at 20 and 35℃. 【Conclusion】 Generally, chlorpyrifos resistance in N. barkeri has little effect on its biological characteristics under different temperatures. The study results provide a theoretical basis for screening resistant strains of N. barkeri and their field application.

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Research progress of insect miRNAs
YANG Jie, XIE Miao, XU Xue-Jiao, BAI Jian-Lin, YOU Min-Sheng
Acta Entomologica Sinica    2021, 64 ( 2): 259-280.   DOI: 10.16380/j.kcxb.2021.02.013
Abstract186)      PDF(pc) (2714KB)(122)    PDF(mobile) (2714KB)(15)    Save
 MicroRNAs (miRNAs) are a class of 19-24 nt endogenous non-coding small RNAs universally existing in various organisms. MiRNAs can regulate gene expression mainly by the combination between its seed region and the ORF or 3′UTR of target genes at the posttranscriptional level, and play critical roles in multiple biological processes including cell differentiation, proliferation and apoptosis. As miRNAs have eventually become a research hotspot in life sciences, great advances have been made in the study of insect miRNAs. The development of highthroughput sequencing and bioinformatics have accelerated the identification of miRNAs in different species, providing the theoretical basis for subsequent research. MiRNAs can be identified with direct cloning, bioinformatics prediction and highthroughput sequencing while their expression levels can be detected through miRNA gene chip analysis, Northern blot and realtime quantitative PCR (RT-qPCR). Inhibition of expression or overexpression of miRNAs can be applied to further reveal their biological functions. MiRNAs exert significant influence on insect growth and reproduction process by participating in ecdysone pathway and regulating the expression of some related genes such as genes related to ecdysone receptor, sex differentiation, wing development, lipid metabolism and ovarian development. MiRNAs are also involved in the circadian rhythm, memory formation and leaning capacity of some insects. In the progress of insect-virus interaction, a number of virus-encoded miRNAs disturb the immune response of host insects by regulating target gene expression while the insectencoded miRNAs influence virus replication. MiRNAs have also been found to participate in insect immune response against exogenous pathogens via regulating the expression of immunerelated genes, which affect innate immune response of insects. Additionally, it has been reported that miRNAs develop or enhance insecticide resistance by negatively regulating the expression of detoxification-related genes, altering insect susceptibility to insecticides and contributing to insecticide resistance in insects. This review provides a theoretical basis for further understanding of insect miRNAs and their potential applications in integrated pest management.
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Selection of reference genes for real-time fluorescence quantitative PCR of miRNAs in Sitobion avenae (Hemiptera: Aphididae)
YANG Chao-Xia, YAN Yi, ZHANG Fang-Mei, ZHU Xun, ZHANG Yun-Hui, LI Xiang-Rui
Acta Entomologica Sinica    2021, 64 (4): 439-448.   DOI: 10.16380/j.kcxb.2021.04.003
Abstract186)      PDF(pc) (1337KB)(70)    PDF(mobile) (1337KB)(29)    Save
【Aim】 This study aims to select the stably expressed reference genes for expression analysis of microRNAs (miRNAs) in the cereal aphid, Sitobion avenae, under specific conditions. 【Methods】 According to the Illumina sequencing results of S. avenae miRNAs, 10 candidate reference genes including miR-10-3p, miR-993, miR-276, miR-275, miR-252a, miR-1, miR-375, pc-15, pc-73 and one normal reference gene U6 were screened, and their relative expression levels in winged and wingless aphids of S. avenae at different developmental stages, in various tissues of winged and wingless aphids and in wingless aphids of S. avenae exposed to four insecticides (95.1% imidacloprid technical material, 97.8% thiacloprid technical material, 95% avermectins technical material and 40% omethoate emulsifiable concentrate) were detected by using qRT-PCR. The expression stability of the 10 candidate reference genes was evaluated by GeNorm, NormFinder, ΔCt method, BestKeeper, and RefFinder. 【Results】 The qRT-PCR results showed that the highly expressed reference genes in winged and wingless aphids of S. avenae at different developmental stages, in various tissues of winged and wingless aphids, and in wingless aphids of S. avenae exposed to four insecticides were miR-276, miR-276, miR-10-3p, and miR-10-3p, respectively. Integrating the analysis results of five approaches, more stably expressed reference genes under the above four conditions were miR-252a, miR-993, miR-275, and miR-993, respectively. The optimal number of reference genes was two under different conditions by GeNorm. Based on the comprehensive ranking of RefFinder, the optimal combinations of stably expressed reference genes in winged and wingless aphids of S. avenae at different developmental stages, in various tissues of winged and wingless aphids, and in wingless aphids of S. avenae exposed to four insecticides were miR-252a and miR-276, miR-993 and miR-276, miR-275 and miR-10-3p, miR-993 and miR-10-3p, respectively. 【Conclusion】 The optimal combinations of reference genes in winged and wingless aphids of S. avenae at different developmental stages, in various tissues of winged and wingless aphids, and in wingless aphids of S. avenae exposed to different insecticides are miR-252a and miR-276, miR-993 and miR-276, miR-275 and miR-10-3p, miR-993 and miR-10-3p, respectively. The results provide a basis of reference gene selection for quantitative expression analysis of miRNA genes in S. avenae.
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cDNA cloning, prokaryotic expression, and ligand binding characterization of the odorant binding proteins CpunOBP3 and CpunOBP4 of the yellow peach moth, Conogethes punctiferalis (Lepidoptera: Crambidae)
GUO Hong-Gang, WEI Chun-Hua, ZHANG Min-Zhao, QIN Xiao-Chun, DU Yan-Li
Acta Entomologica Sinica    2021, 64 (3): 327-336.   DOI: 10.16380/j.kcxb.2021.03.005
Abstract185)      PDF(pc) (1954KB)(65)    PDF(mobile) (1954KB)(11)    Save
【Aim】 This study aims to determine the physiological function of odorant binding proteins (OBPs) in the chemoreception of the yellow peach moth, Conogethes punctiferalis, so as to provide a theoretical basis for selecting OBPs that may be used as targets in C. punctiferalis biological control. 【Methods】 Two OBP genes (CpunOBP3 and CpunOBP4) were cloned from antennae of C. punctiferalis by PCR based on our previous antennal transcriptome data. The nucleotide and amino acid sequences of the two OBPs were analyzed with bioinformatics software. The recombinant  xpression vectors pET-30a/CpunOBP3 and pET-30a/CpunOBP4 were constructed. and the recombinant proteins CpunOBP3 and CpunOBP4 were obtained by prokaryotic expression and purification. The binding activity of the recombinant CpunOBP3 and CpunOBP4 to 24 ligands was analyzed by fluorescence competitive binding assay.【Results】 The open reading frame of CpunOBP3 gene (GenBank accession no.: GEDO010000010.1) is 387 bp in length, encoding a protein of 128 amino acids with the predicted molecular weight of 14.72 kD. The open reading frame of CpunOBP4 gene (GenBank accession no.: GEDO010000011.1) is 438 bp in length, encoding a protein of 145 amino acids. The predicted molecular weight of  CpunOBP4 without signal peptide is 12.82 kD. CpunOBP3 and CpunOBP4 share the typical structural features of OBPs, including six conservative cysteine residues. Both the recombinant CpunOBP3 and CpunOBP4 were mainly expressed in the inclusion. Fluorescence competitive binding assay indicated that the recombinant CpunOBP3 showed the binding ability to seven plant volatiles tested, with the strongest binding ability to 3-carene(Ki=10.33 μmol/L), but not to two sex pheromones tested. The recombinant CpunOBP4 exhibited the binding ability not only to the two sex pheromones tested (cis-10-hexadecenal with the Ki value of 14.65 μmol/L and hexadecanoyl with the Ki value of 7.83 μmol/L), but also to eight plant volatiles tested, with the strongest binding ability to ethyl butyrate (Ki=4.32 μmol/L). 【Conclusion】 Based on these results, we inferred that CpunOBP3 plays an important role in the host location and shift of C. punctiferalis, while CpunOBP4 possesses dual functions in recognizing sex pheromones and plant volatiles. The results provide a theoretic basis for controlling the occurrence and damage of C. punctiferalis via disturbing its olfactory reception.
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Influences of gall-inducing insects on the physiology and metabolism of host plants
YANG Meng-Ke, LIU Sai, QIAO Hai-Li, GUO Kun, XU Rong, XU Chang-Qing, CHEN Jun
Acta Entomologica Sinica    2021, 64 (4): 536-548.   DOI: 10.16380/j.kcxb.2021.04.012
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Plant galls, the abnormal outgrowths of plant tissues induced by gallinducing insects, are ideal materials to investigate the co-evolution between plants and insects. Gall-inducing insects are also important pests in agriculture and forestry. Investigations into the effects of gall-inducing insects on their host plants are useful to reveal the relationships between gall-inducing insects and plants, and can also help reveal the growth process of host plants. Besides, investigations into the responses of galled plants to gall-inducing insects are helpful to screen out the resistance indexes, resistance genes and sensitive genes of plants, providing a theoretical basis for resistance breeding. This review focuses mainly on the effects of gall-inducing insects on the photosynthesis, physiology and metabolism of their host plants. Gall-inducing insects generally reduce the photosynthetic pigments and photosynthetic rate of host plants, raise the contents of primary metabolites such as sugars and amino acids in inner tissues of galls, raise the contents of secondary metabolites such as non-volatile phenols and flavonoids and volatile terpenoids in outer tissues of galls, raise the activities of protective enzymes such as POD and SOD, and raise the contents of phytohormones such as IAA, SA and JA in host plants. The current research data indicate that investigations into the influences of gallinducing insects on the physiology and metabolism of host plants are even in their infancy, and the influencing mechanisms still need to be further explore.
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Expression patterns and pheromone-binding properties of the pheromone binding protein CpunPBP3 in Conogethes punctiferalis (Lepidoptera: Crambidae) (In English)
CHEN Qiu-Ying, YANG Xi, YOU Dong-Rui, YANG Mu, XU Zhi-Feng, XIAO Wei
Acta Entomologica Sinica    2021, 64 (3): 318-326.   DOI: 10.16380/j.kcxb.2021.03.004
Abstract183)      PDF(pc) (1706KB)(58)    PDF(mobile) (1706KB)(9)    Save

【Aim】 This study aims to better understand the sex pheromone perception mechanisms by identifying and characterizing a sex pheromone binding protein (PBP) in the yellow peach moth, Conogethes punctiferalis (CpunPBP3). 【Methods】 The cDNA sequence of CpunPBP3 of C. punctiferalis was amplified and analyzed, and the amino acid sequence was compared to those of the homologous proteins in other Crambidae species. The day-age-dependent changes and circadian fluctuations in the expression levels of CpunPBP3 in the male adult antenna of C. punctiferalis, and the changes in the expression level of CpunPBP3 in the antenna over 24 h-period following exposure of adult males to the sex pheromones E10-16∶Ald (150 ng) and Z10-16∶Ald (6 ng) were examined by qRT-PCR. The recombinant expression vector pET-30a(+)/CpunPBP3 was constructed, and the recombinant CpunPBP3 was expressed in Escherichia coli. The binding capacity of the purified recombinant protein CpunPBP3 with the above two sex pheromones was evaluated by fluorescence competitive binding assay. 【Results】 The phylogenetic analysis result revealed that CpunPBP3 and the previously identified C. punctiferalis PBP genes CpunPBP2 and CpunPBP5 clustered in different branches, but CpunPBP3 is similar to PBP genes in other insect species. The qRT-PCR results showed that the expression level of CpunPBP3 in the male adult antenna increased first and then decreased from day 0 to 8 after adult eclosion, with significantly higher expression level at 17∶00 than at 1∶00, but with no significant difference at other time points within 24-h photoperiod. However, the expression level of CpunPBP3 in the male adult antenna significantly decreased after induction by 150 ng E10-16∶Ald for 3 and 6 h, and significantly increased after induction by 6 ng Z10-16∶Ald for 6 and 24 h. Fluorescence competitive binding assay result showed that the recombinant CpunPBP3 had strong binding capacity with E10-16∶Ald and Z10-16∶Ald, with the Ki values of 9.267 and 8.656 μmol/L, respectively. 【Conclusion】 The study determined the nucleotide and amino acid sequences and the expression pattern of CpunPBP3, and CpunPBP3 was expressed in response to the sex pheromone induction. The recombinant CpunPBP3 has strong binding capacity with sex pheromone, indicating that CpunPBP3 is a sex pheromone binding protein in C. punctiferalis.

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Differences in the evolution of mitochondrial genome between pollinating and non-pollinating fig wasps (In English)
WANG Jian-Xia, ZHOU Yi, XIN Zhao-Zhe, ZHAO Dan, XIAO Jin-Hua, HUANG Da-Wei
Acta Entomologica Sinica    2021, 64 (4): 479-489.   DOI: 10.16380/j.kcxb.2021.04.007
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【Aim】 At present, there are few reports on the mitochondrial genomes of fig wasps. The purpose of this study is to explore whether there are some differences in the evolution of mitochondrial genome (mitogenome) between pollinating fig wasps (PFWs) and non-pollinating fig wasps (NPFWs). 【Methods】 Based on the mitogenomes from 15 fig wasp species, of which the mitogenomes of 11 species were newly determined, we used the comparative mitochondrial genomic method to analyze the sequence and evolutionary characteristics of the mitogenomes of fig wasps. 【Results】 The length of the mitogenomes of 11 fig wasps newly determined ranges from 12 768 to 17 060 bp, and the AT content in the 11 mitogenomes is more than 80%. The AT-skew is negative and the GC-skew is positive in most species except for the non-pollinating fig wasp Philotrypesis tridentata. Frequent mitochondrial gene rearrangement occurs in fig wasps, which may be valuable for phylogenetic analysis of the species. Further analysis of selection pressure indicates that the ω ratios of protein-coding genes (PCGs) in mitogenomes of fig wasps are far less than 1, suggesting that these genes have experienced purifying selection. However, most of the genes in PFWs may have accumulated more nonsynonymous mutations than those in NPFWs. Furthermore, compared with the NPFWs, the mitogenomes of PFWs have more gene rearrangements, and higher nucleotide diversity and amino acid substitution rate. 【Conclusion】 The mitogenome evolution of PFWs is faster than that of NPFWs, which may be related to the significantly different lifestyles or evolutionary histories of the two groups.

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Function of the cytoplasmic peptidoglycan recognition protein RfPGRP-L2 in maintaining the homeostasis of gut microbiota in Rhynchophorus ferrugineus (Coleoptera: Dryophthoridae) 
XIAO Rong, WANG Xing-Hong, LI Xiong-Wei, LIU Hui-Hui, LU Sheng-Ping, HOU You-Ming, SHI Zhang-Hong
Acta Entomologica Sinica    2021, 64 (3): 348-362.   DOI: 10.16380/j.kcxb.2021.03.007
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Abstract: 【Aim】 To determine the function of a cytoplasmic peptidoglycan recognition protein, RfPGRP-L2, in the maintenance and regulation of homeostasis of gut microbiota in the invasive insect pest, Rhynchophorus ferrugineus, so as to provide scientific basis and action targets for the development of new insect pest control strategies targeting to destroy the homeostasis of gut microbiota. 【Methods】 The sequence characteristics of RfPGRP-L2 were analyzed by bioinformatics methods. RT-qPCR was used to analyze the expression levels of RfPGRP-L2 in the different tissues (head, fat body, epidermis, foregut, mid-/hindgut and hemolymph) of the healthy 4th instar larvae of R. ferrugineus and in the gut and fat body of the 4th instar larvae of R. ferrugineus challenged with Escherichia coli DH5α and Staphylococcus aureus by injection with 1 μL bacterial suspension with the OD600 value of 1.6 and oral feeding with sugarcane slices smeared with 1 mL bacterial suspension with the OD600 value of 1.6, respectively. Prokaryotic expression of RfPGRP-L2 was carried out, and in vitro assays were applied to determine the agglutination and antibacterial activity of the recombinant RfPGRP-L2 to E. coli DH5α and S. aureus. After RNAi of RfPGRP-L2, the number of gut bacterial colonies of E. coli in the hemolymph and gut of the 4th instar larvae of R. ferrugineus was determined. The expression levels of antimicrobial peptide genes in the fat body and gut of the 4th instar larvae of R. ferrugineus after RNAi of RfPGRP-L2 were detected by RT-qPCR. By bacterial 16S rRNA-based high-throughput sequencing, the effect of RNAi of RfPGRP-L2 on the gut microbiota composition of the healthy 4th instar larvae of R. ferrugineus was analyzed.【Results】 The SMART analysis revealed that RfPGRP-L2 has no transmembrane domain and signal peptide, suggesting that RfPGRP-L2 is a cyoplasmic peptidoglycan recognition protein. The RT-qPCR results showed that RfPGRP-L2 was highly expressed in the immunity-related tissues, such as the hemolymph, gut and fat body of the healthy 4th instar larvae of R. ferrugineus. The expression level of RfPGRP-L2 in the fat body of the 4th instar larvae of R. ferrugineus was significantly up-regulated upon the challenge with injected E. coli and S. aureus for 6 h and 12 h, respectively. After oral feeding of E. coli for 6 h, the expression level of RfPGRP-L2 in the gut of the 4th instar larvae of R. ferrugineus was also increased significantly. Furthermore, in vitro assays revealed that the recombinant RfPGRP-L2 could result in the obvious agglutination of E. coli and S. aureus, suggesting that RfPGRP-L2 could recognize the two pathogenic bacteria. When RfPGRP-L2 was silenced in the 4th instar larvae of R. ferrugineus, the ability to clear the invaded EGFP-tagged E. coli in the gut and hemolymph was significantly impaired as compared to the control group, the expression level of antimicrobial peptide gene RfCecropin in the gut was significantly reduced, the number of gut bacterial colonies in the healthy 4th instar larvae was significantly increased compared to the control group, and the gut bacterial composition was also altered significantly. 【Conclusion】 The cyoplasmic peptidoglycan recognition protein RfPGRP-L2, acting as a pattern recognition receptor to discriminate pathogens and to activate the immune signaling pathways in intestinal epithelial cells, can promote the expression of antimicrobial peptide gene to modulate the homeostasis of gut microbiota in R. ferrugineus.

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Effects of the serine protease homologue SgSPH from the venom of Scleroderma guani (Hymenoptera: Bethylidae) on the phenoloxidase activity in the host hemolymph
LI Li-Fang, WU Chao-Yan, HAN Kai-Jian, WU Guo-Xing, ZHU Jia-Ying
Acta Entomologica Sinica    2021, 64 ( 2): 170-177.   DOI: 10.16380/j.kcxb.2021.02.004
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【Aim】 The aim of this study is to clone and express the venom serine protease homologue (SPH) gene of Sclerotium guani (SgSPH), and to investigate the effect of the venom protein encoded by this gene on the phenoloxidase activity in the host hemolymph. 【Methods】 The open reading frame (ORF) of venom SgSPH gene was cloned from S. guani by RT-PCR. Its sequence features were analyzed using bioinformatic software. The relative expression levels of the venom SgSPH gene at different developmental stages (egg, early instar larva, late instar larva, mature larva, spinning larva, pupa in yellow cocoon, pupa in black cocoon, and 1-5-day-old adults) and in different female adult tissues (head, thorax, abdomen without venom apparatus and venom apparatus) of S. guani were determined by qPCR. This venom gene was expressed with prokaryotic expression vector pSUMO-Mut. The expressed recombinant protein was purified using Ni-chelating affinity chromatography, and examined by SDS-PAGE and Western blot analysis. The inhibitory effect of the recombinant SgSPH on the phenoloxidase activity in the pupal haemolymph of Tenebrio molitor was measured by enzymatic activity assay. 【Results】 The ORF of the venom SgSPH gene (GenBank accession number: MT920663) of S. guani was cloned. It is 798 bp in length, encoding 265 amino acids, with the signal peptide consisting of amino acids 1-20, and the predicted protein molecular mass of 30.53 kD and pI of 9.59. Multiple sequence alignment results showed that SgSPH of S. guani shares low amino acid sequence identity (9%-17%) with the serine proteases and SPHs from venoms of other parasitoid wasps, and lacks a conservative catalytic triad. The qPCR results indicated that SgSPH gene was abundantly expressed at the adult stage and in the venom apparatus of S. guani. SDSPAGE and Western blot analyses showed that the recombinant SgSPH was successfully expressed and the highly purified recombinant SgSPH was obtained. Enzymatic activity assay results showed that the recombinant SgSPH was able to inhibit the phenoloxidase activity in the pupal hemolymph of the host T. molitor. 【Conclusion】 The results suggest that the venom SgSPH of S. guani can interfere with the host phenoloxidase cascade.

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Research progress of insect sodium channels#br#
WU Shao-Ying, DUAN Wen-Bo, LI Fen, YANG Lei, WANG Hao, WANG Li-Kui
Acta Entomologica Sinica    2021, 64 (7): 862-874.   DOI: 10.16380/j.kcxb.2021.07.010
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There are only one or two voltage-gated sodium channel α subunit genes in insects, but the two post-transcriptional modifications, alternative splicing and RNA editing, confer the functional diversity of insect sodium channels. The insect β accessory subunits, TipE and TEH1-4, also play important roles in the expression and regulation of sodium ion channels. Voltagegated sodium channel plays an important role in the generation and transmission of action potential and is the target site of many natural and synthetic neurotoxins and insecticides, including the pyrethroids, indoxacarb and metaflumizone. Pyrethroids can prolong the transmembrane sodium ion flow by controlling the inactivation and deactivation of sodium channels in insects, causing neuroexcitatory conduction disorders. Indoxacarb and metaflumizone block the neuronal action potential in the central and peripheral nervous system of insects. These neural agents can disturb the normal function of sodium channels in insects. Two pyrethroid binding sites have been commonly identified in sodium channels of insects, but sodium channels of different species have differences in binding sites for pyrethroids. Therefore, in this article we reviewed insect sodium channels and their interaction with insecticides, hoping to promote the research of insect nerve receptors and to provide important references for identification of mutations associated with resistance and development of effective insecticides.
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Insect metamorphosis: Nature, history, evolution and regulation-A review of the book Insect Metamorphosis: From Natural History to Regulation of Development and Evolution
LI Kang, SONG Jia-Sheng, ZHOU Shu-Tang, LI Sheng
Acta Entomologica Sinica    2021, 64 (6): 769-770.   DOI: 10.16380/j.kcxb.2021.06.012
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Identification and characterization of two aminopeptidases N from the midgut of the brown planthopper, Nilaparvata lugens (Hemiptera: Delphacidae)
LIN Li, YU Xiao-Qiang, GUAN Xiong, SHAO En-Si
Acta Entomologica Sinica    2021, 64 (7): 771-780.   DOI: 10.16380/j.kcxb.2021.07.001
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【Aim】 Aminopeptidases N (APNs) are a class of important proteases in the digestive system in insects. This study aims to verify the expression of two apn genes (nlapn1 and nlapn4) with high transcription level in the midgut epithelium of the brown planthopper, Nilaparvata lugens, and to identify and analyze the characteristics of their proteins. 【Methods】 Phylogenetic analysis of both NLAPN1 and NLAPN4 of N. lugens was conducted by maximum likelihood method. Western blotting and LC-ESI-MS/MS were respectively conducted to localize and identify NLAPN1 and NLAPN4 in the midgut brush border membrane vesicles (BBMVs) of N. lugens. NLAPN1 and NLAPN4 were respectively expressed in Drosophila S2 cells. Lacolization of both NLAPN1 and NLAPN4 in S2 cells was analyzed by Western blot and immunofluorescence. The enzymatic activities of NLAPN1 and NLAPN4 were determined through enzyme assays using leucine-p-NA, Ala-p-NA, Met-p-NA and Lys-p-NA as the substrate, respectively. 【Results】 Phylogenetic tree analysis showed that both NLAPN1 and NLAPN4 of N. lugens were clustered together with the APN proteins highly expressed in the midgut of other hemipteran insects. NLAPN1 and NLAPN4 with the molecular weight of ~160 kD were identified in the midgut BBMV of N. lugens by Western blot and LC-ESI-MS/MS. Western blot and immunofluorescence analysis showed that NLAPN1 and NLAPN4 were expressed on the cytomembrane of transfected S2 cells, while that of NLAPN4lackG without glycosylphosphatidylinositol (GPI) anchor site at the C-terminal end was distributed in the cytoplasm. Enzyme assay results revealed that both NLAPN1 and NLAPN4 showed certain enzymatic activity using Ala-p-NA and Lys-p-NA as the substrate, while using leucine-p-NA as the substrate, NLAPN1 showed extremely high enzymatic activity (>60 U/mg). 【Conclusion】 NLAPN1 and NLAPN4 are both highly expressed GPI-anchored membrane-bound aminopeptidases N located on the epithelial membrane of the midgut of N. lugens. Both NLAPN1 and NLAPN4 show similar structure and enzymatic characteristics to the previous identified membrane-bound APN proteins in lepidotperans, coleopterans and dipterans. Physiological and biochemical functions of membrane-bound APNs in the midgut of N. lugens and their interaction with exogenous pathogenic microorganisms need to be further studied.
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Oviposition preference of Serangium japonicum (Coleoptera: Coccinellidae) to eggplant varieties with different leaf trichome densities
MEI Wen-Juan, YAO Feng-Luan, LIN Shuo, DING Xue-Ling, ZHENG Yu, LU Xue-Song, HE Yu-Xian, WENG Qi-Yong
Acta Entomologica Sinica    2021, 64 (9): 1092-1103.   DOI: 10.16380/j.kcxb.2021.09.009
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 【Aim】 Oviposition is vital for the survival and development of insect populations. Our previous study demonstrated that Serangium japonicum prefer to lay eggs on eggplant (Solanum melongena) leaves. This study aims to explore the oviposition preference of S. japonicum to eggplant varieties with different leaf trichome densities, so as to further clarify the mechanisms regarding to leaf trichome-related oviposition preference in S. japonicum. 【Methods】 Leaf discs and plants of four eggplant varieties (Heidaodi, Heiniu, Riyouchangzhizhuang and Luoxing) were provided for S. japonicum to determine the oviposition preference. The offspring performance including the growth and development, and adult fecundity, predation capability and attachment force of S. japonicum on leaves of each of the four eggplant varieties was tested. Microstructure of the abaxial leaf surface of the four eggplant varieties, the preference of S. japonicum female adults to leaf odours, the length and width of Bemisia tabaci eggs on eggplant leaves, and the cannibalism risk of S. japonicum eggs by its female adults were examined. 【Results】 S. japonicum preferred to lay eggs on Heidaodi, where the shortest offspring developmental duration (15.41±0.21 d) and the highest fecundity (877.44±15.27 eggs laid per female) were found. However, the proportion of S. japonicum eggs had no significant correlation with offspring performance (developmental duration, survival rate and adult body weight) and adult performance (fecundity and predation capability), but was significantly positively correlated with the density and length of leaf trichomes on the abaxial leaf surface (ALS). Furthermore, the proportion of S. japonicum eggs laid on leaf discs was significantly positively correlated with the attachment force of its female adults. The attachment force of S. japonicum female adults was positively correlated with leaf trichome density but had no significant correlation with adult performance. The leaf odour, prey quality, and cannibalism risk on the four eggplant varieties had no significant effects on the oviposition preference of S. japonicum. 【Conclusion】 The density of ALS leaf trichome and the attachment force of S. japonicum mediated by leaf trichomes play an important role in the oviposition preference of S. japonicum.
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