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Scopula subpunctaria, an abundant pest in tea gardens, produce type-II sex pheromone components, which are critical for its communicative and reproductive abilities; however, genes encoding the proteins involved in the detection of type-II sex pheromone components have rarely been documented in moths. In the present study, we sequenced the transcriptomes of the male and female S. subpunctaria antennae. A total of 150 candidate olfaction genes, comprising 58 odorant receptors (SsubORs), 26 ionotropic receptors (SsubIRs), 24 chemosensory proteins (SsubCSPs), 40 odorant-binding proteins (SsubOBPs), and 2 sensory neuron membrane proteins (SsubSNMPs) were identified in S. subpunctaria. Phylogenetic analysis, qPCR, and mRNA abundance analysis results suggested that SsubOR46 may be the Orco (non-traditional odorant receptor, a subfamily of ORs) of S. subpunctaria. SsubOR9, SsubOR53, and SsubOR55 belonged to the pheromone receptor (PR) clades which have a higher expression in male antennae. Interestingly, SsubOR44 was uniquely expressed in the antennae, with a higher expression in males than in females. SsubOBP25, SsubOBP27, and SsubOBP28 were clustered into the moth pheromone-binding protein (PBP) sub-family, and they were uniquely expressed in the antennae, with a higher expression in males than in females. SsubOBP19, a member of the GOBP2 group, was the most abundant OBP in the antennae. These findings indicate that these olfactory genes, comprising five candidate PRs, three candidate PBPs, and one candidate GOBP2, may be involved in type II sex pheromone detection. As well as these genes, most of the remaining SsubORs, and all of the SsubIRs, showed a considerably higher expression in the female antennae than in the male antennae. Many of these, including SsubOR40, SsubOR42, SsubOR43, and SsubIR26, were more abundant in female antennae. These olfactory and ionotropic receptors may be related to the detection of host plant volatiles. The results of this present study provide a basis for exploring the olfaction mechanisms in S. subpunctaria, with a focus on the genes involved in type II sex pheromones. The evolutionary analyses in our study provide new insights into the differentiation and evolution of lepidopteran PRs.

For many herbivorous insects, vision is more important than olfaction in the prealighting stage of host habitat location. Tea leafhoppers, Empoasca onukii (Hemiptera, Cicadellidae), are serious pests that preferentially inhabit the tender leaves of tea plants across China. Here, we investigated whether tea leafhoppers could distinguish foliage colors associated with different leaf ages and use this visual cue to guide suitable habitat location from short distances. Similar to honeybees, the adult E. onukii has an apposition type of compound eye, and each ommatidium has eight retinular cells, in which three spectral types of photoreceptors are distributed, with peak sensitivities at 356 nm (ultraviolet), 435 nm (blue), and 542 nm (green). Both changes in spectral intensity and hue of reflectance light of the host foliage were correlated with varying leaf age, and the intensity linearly decreased with increasing leaf age. Behavioral responses also showed that adult E. onukii could discriminate between the simulated colors of host foliage at different leaf ages without olfactory stimuli and selected the bright colors that strongly corresponded to those of tender leaves. The results suggest that, compared with the spectral composition (hue), the intensity of light reflectance from leaves at different ages is more important for adult leafhoppers when discriminating host foliage and could guide them to tender leaves at the top of tea shoots.

Species-specific sex pheromone is biosynthesized and released in most female moths as a chemical cue in mating communication. However, information on genes involved in this pathway is limited. The beet armyworm, Spodoptera exigua, is a cosmopolitan agricultural pest that causes severe economic losses to many crops. In China, the female sex pheromones in sex pheromone glands (PGs) of S. exigua have been measured which comprises (Z,E)-9,12-tetradecadienyl acetate, (Z)-9-tetradecen-l-ol, (Z)-9-tetradecenyl acetate, and (Z,E)-9,12-tetradecadien-1-ol in a ratio of 47:18:18:17. Fifty-nine putative genes related to sex pheromone biosynthesis were identified in the present study by sequencing and analyzing the sex pheromone gland (PG) transcriptome of S. exigua. Expression profiles revealed that two desaturase (SexiDes5 and SexiDes11) and three fatty acyl reductase (SexiFAR2, 3, and 9) genes had PG-specific expression, and phylogenetic analysis demonstrated that they clustered with genes known to be involved in pheromone synthesis in other moth species. Our results provide crucial background information that could facilitate the elucidation of sex pheromone biosynthesis pathway of S. exigua as well as other Spodoptera species and help identify potential targets for disrupting sexual communication in S. exigua for developing novel environment-friendly pesticides.

Jasmonates (JAs), which can exogenously induce anti-herbivore defensive reaction in plants, are important in the field of plant physiology and plant protection. We summarized the ecological effects of exogenous JAs applied on 40 species of plants on various herbivores (such as Lepidoptera and Hemipteran) and their natural enemies in the past 20 years. We systematically genera-lized the research status about the direct and indirect antiherbivore defense induced by JAs, induced systemic defense, induction methods, induced performance in field, and the application status of JAs. Moreover, combining with the latest literatures, we reviewed the effects of JAs on plant growth-defense trade-offs from the cross-talk between phytohormone signaling pathways, and the regulation of nodes in the JA signaling pathway. Finally, we proposed the future directions and key aspects of the research on the plant anti-herbivore defense induced by exogenous JAs, which would promote the development of the related research and the application of JAs in field.茉莉酸类化合物(jasmonates, JAs)可外源诱导植物产生抗虫防御反应.因此,JAs在植物生理及植物保护学领域具有十分重要的研究价值.本文归纳了近20年来在约40种植物上外源施用JAs对鳞翅目、半翅目等植食者及其天敌所产生的生态影响,并从JAs诱导植物产生的直接与间接抗虫反应、诱导系统抗性、诱导方法、田间诱导抗虫表现、应用现状等方面对JAs诱导植物抗虫的研究现状进行了系统的总结.同时,本文结合最新研究结果,从植物体内信号交流、JA信号途径节点调控等方面,对植物体内与JA途径相关的生长-防御“权衡”进行了综述.最后,本文对今后外源JAs诱导植物抗虫研究的发展方向与重点方面进行了分析和展望,以期促进相关研究及JAs田间应用的创新发展.

Chrysoperla sinica is one of the most prominent natural enemies of many agricultural pests. Host seeking in insects is strongly mediated by olfaction. Understanding the sophisticated olfactory system of insect antennae is crucial for studying the physiological bases of olfaction and could also help enhance the effectiveness of C. sinica in biological control. Obtaining olfactory genes is a research priority for investigating the olfactory system in this species. However, no olfaction sequence information is available for C. sinica. Consequently, we sequenced female- and male-antennae transcriptome of C. sinica. Many candidate chemosensory genes were identified, including 12 odorant-binding proteins (OBPs), 19 chemosensory proteins (CSPs), 37 odorant receptors (ORs), and 64 ionotropic receptors from C. sinica. The expression patterns of 12 OBPs, 19 CSPs and 37 ORs were determined by RT-PCR, and demonstrated antennae-dominantly expression of most OBP and OR genes. Our finding provided large scale genes for further investigation on the olfactory system of C. sinica at the molecular level.

Moths can biosynthesize sex pheromones in the female sex pheromone glands (PGs) and can distinguish species-specific sex pheromones using their antennae. However, the biosynthesis and transportation mechanism for Type II sex pheromone components has rarely been documented in moths. In this study, we constructed a massive PG transcriptome database (14.72 Gb) from a moth species, Ectropis grisescens, which uses type II sex pheromones and is a major tea pest in China. We further identified putative sex pheromone biosynthesis and transportation-related unigenes: 111 cytochrome P450 monooxygenases (CYPs), 25 odorant-binding proteins (OBPs), and 20 chemosensory proteins (CSPs). Tissue expression and phylogenetic tree analyses showed that one CYP (EgriCYP341-fragment3), one OBP (EgriOBP4), and one CSP (EgriCSP10) gene displayed an enriched expression in the PGs, and that EgriOBP2, 3, and 25 are clustered in the moth pheromone-binding protein clade. We considered these our candidate genes. Our results yielded large-scale PG sequence information for further functional studies.

Odorant-degrading esterases (ODEs) act in the fast deactivation of ester pheromone components and plant volatiles in insects. However, only few ODEs have been characterised to date. In this study, six full-length putative ODE genes (designated SexiCXE4, 5, 17, 18, 20, and 31) were cloned from the male antennae of Spodoptera exigua. The deduced amino acid sequences possessed typical characteristics of a carboxylesterase (CXE) and shared high identities with reported insect CXEs. The tissue and temporal expression patterns were investigated by quantitative real time PCR. Although all six SexiCXEs are expressed in antennae of both sexes, SexiCXE4, 17 and 20 are antennae-enriched; while SexiCXE5 and SexiCXE18 are dominantly expressed in wings, and SexiCXE31 is mainly expressed in proboscises, heads and legs. With the highly biased expression in antennae and proboscises, SexiCXE4 was selected for further functional assay. The recombinant SexiCXE4 were expressed in High-five cells and purified by a Ni(2+) affinity column. SexiCXE4 has much higher enzyme activity against plant volatiles (Z)-3-hexenyl acetate and hexyl acetate than to the sex pheromone components, suggesting that it may function mostly in the degradation of the plant volatiles.

The normalization of quantitative real-time PCR (qPCR) is important to obtain accurate gene expression data, and the most common method for qPCR normalization is to use reference genes. However, reference genes can be regulated under different conditions. qPCR has recently been used for gene expression study in Laodelphax striatellus, but there is no study on validation of the reference genes. In this study, five new housekeeping genes (LstrTUB1, LstrTUB2, LstrTUB3, LstrARF and LstrRPL9) in L. striatellus were cloned and deposited in the GenBank with accession numbers of JF728809, JF728810, JF728811, JF728807 and JF728806, respectively. Furthermore, mRNA expressions of the five genes and β-actin were measured by qPCR with insect samples of different instar at nymph stage, and the expression stabilities were determined by the software geNorm and NormFinder. As a result, ARF and RPL9 were consistently more stable than β-actin, while three TUB genes were less stable than β-actin. To determine the optimal number of reference genes used in qPCR, a pairwise variations analysis by geNorm indicated that two references ARF and RPL9 were required to obtain the accurate quantification. These results were further confirmed by the validation qPCR experiment with chitinase gene as the target gene, in which the standard error of the mRNA quantification by using binary reference ARF-RPL9 was much lower than those by ARF, RPL9 or β-actin alone. Taken together, our study suggested that the combination of ARF-RPL9 could replace β-actin as the reference genes for qPCR in L. striatellus.

Odorant-binding proteins (OBPs), widely and abundantly expressed in insect olfactory organs, are important for insect olfaction. In this study, two novel Minus-C OBP genes (HarmOBP17 and HarmOBP18) were characterized from the cotton bollworm, Helicoverpa armigera, one of the most important pests in cotton and other crops. Quantitative RT-PCR results indicate that the expression levels of the two genes differed greatly in regard to developmental stages, tissues, and genders. Binding properties of the two OBPs with plant volatiles were investigated at different pH levels using a competitive binding assay. Both OBPs displayed the highest binding affinities (Ki13 μM) with Beta-ionone among the 85 tested compounds. Surprisingly, the two OBPs (HarmOBP17 in particular) displayed higher binding at pH 5.0 than at pH 7.4 and 10.0. This is the first reported Lepidopteran OBPs showing a contrary pH-dependent ligand binding behavior to other OBPs, implying a different mechanism of ligand binding and release. In addition, binding assays of the mutant OBPs (with the C-terminus eliminated) showed that the C-terminus was important for the ligand binding, but this importance varied with different ligands and OBPs, suggesting that their binding properties depend on the specific interactions between OBP and ligand.

Differences in sex pheromone component can lead to reproductive isolation. The sibling noctuid species, Helicoverpa armigera and Helicoverpa assulta, share the same two sex pheromone components, Z9-16:Ald and Z11-16:Ald, but in opposite ratios, providing an typical example of such reproductive isolation. To investigate how the ratios of the pheromone components are differently regulated in the two species, we sequenced cDNA libraries from the pheromone glands of H. armigera and H. assulta. After assembly and annotation, we identified 108 and 93 transcripts putatively involved in pheromone biosynthesis, transport and degradation in H. armigera and H. assulta, respectively. Semi-quantitative RT-PCR, qRT-PCR, phylogenetic and mRNA abundance analyses suggested that some of these transcripts involved in the sex pheromone biosynthesis pathways perform. Based on these results, we postulate that the regulation of desaturases, KPSE and LPAQ, might be key factor regulating the opposite component ratios in the two sibling moths. In addition, our study has yielded large-scale sequence information for further studies and can be used to identify potential targets for the bio-control of these species by disrupting their sexual communication.

Chrysopa pallens is an important natural predator of various pests in many different cropping systems. Understanding the sophisticated olfactory system of insect antennae is crucial for studying the physiological bases of olfaction and could also help enhance the effectiveness of C. pallens in biological control. However, functional studies of the olfactory genes in C. pallens are still lacking. In this study, we cloned five odorant-binding protein (OBP) genes from C. pallens (CpalOBPs). Quantitative RT-PCR results indicated that the five CpalOBPs had different tissue expression profiles. Ligand-binding assays showed that farnesol, farnesene, cis-3-hexenyl hexanoate, geranylacetone, beta-ionone, octyl aldehyde, decanal, nerolidol (Ki20 μM), and especially 2-pentadecanone (Ki=1.19 μM) and 2-hexyl-1-decanol (Ki=0.37 μM) strongly bound to CpalOBP2. CpalOBP15 exhibited high binding affinities for beta-ionone, 2-tridecanone, trans-nerolidol, and dodecyl aldehyde. Behavioral trials using the 14 compounds exhibiting high binding affinities for the CpalOBPs revealed that nine were able to elicit significant behavioral responses from C. pallens. Among them, farnesene and its corresponding alcohol, farnesol, elicited remarkable repellent behavioral responses from C. pallens. Our study provides several compounds that could be selected to develop slow-release agents that attract/repel C. pallens and to improve the search for strategies to eliminate insect pests.

The complete mitochondrial genome of the Chrysoperla sinica was sequenced in this study. It is 16,057 bp in length, and contains 22 transfer RNA genes, 13 protein-coding genes (PCGs), 2 rRNA genes and 1 non-coding AT-rich region. But, C. sinica lack long tandem repeats. The Chrysoperla sinica mitogenome displays a very high level of similarity (99.27%) with Chrysoperla nipponensis. The genome base composition is 39.24% A, 39.70% T, 11.98% C, and 9.08% G. Similar to other insects, all PCGs are initiated by a typical ATN codon, except COX1, which starts with TCG. Nine PCGs have the standard, complete stop codon (TAA or TAG), and the other four PCGs end with the incomplete stop codon (TA or T).

The complete mitochondrial genome of the tobacco budworm Helicoverpa assulta was sequenced. It is 15,351 bp in length, and contains 22 transfer RNA genes, 13 protein-coding genes (PCGs), 2 rRNA genes and a non-coding AT-rich region. The H. assulta mitogenome displays a very high level of identity (99.76%) with Helicoverpa armigera. The genome base composition is 40.50% A, 40.40% T, 11.32% C and 7.70% G. Similar to other Lepidoptera insects, all PCGs are initiated by a typical ATN codon, except CO1, which starts with CGA. Nine PCGs have the standard, complete stop codon (TAA), and the other four PCGs end with the incomplete stop codon TA-.

Chemosensory proteins (CSPs) have been proposed to capture and transport hydrophobic chemicals to receptors on sensory neurons. We identified and cloned 24 CSP genes to better understand the physiological function of CSPs in Helicoverpa armigera. Quantitative real-time polymerase chain reaction assays indicate that CSP genes are ubiquitously expressed in adult H. armigera tissues. Broad expression patterns in adult tissues suggest that CSPs are involved in a diverse range of cellular processes, including chemosensation as well as other functions not related to chemosensation. The H. armigera CSPs that were highly transcribed in sensory organs or pheromone glands (HarmCSPs 6, 9, 18, 19), were recombinantly expressed in bacteria to explore their function. Fluorescent competitive binding assays were used to measure the binding affinities of these CSPs against 85 plant volatiles and 4 pheromone components. HarmCSP6 displays high binding affinity for pheromone components, whereas the other three proteins do not show affinities for any of the compounds tested. HarmCSP6 is expressed in numerous cells located in or close to long sensilla trichodea on the antennae of both males and females. These results suggest that HarmCSP6 may be involved in transporting female sex pheromones in H. armigera.

Odorant-degrading esterases (ODEs) act in the fast deactivation of ester pheromone components and plant volatiles in insects. However, only few ODEs have been characterised to date. In this study, six full-length putative ODE genes (designated SexiCXE4, 5, 17, 18, 20, and 31) were cloned from the male antennae of Spodoptera exigua. The deduced amino acid sequences possessed typical characteristics of a carboxylesterase (CXE) and shared high identities with reported insect CXEs. The tissue and temporal expression patterns were investigated by quantitative real time PCR. Although all six SexiCXEs are expressed in antennae of both sexes, SexiCXE4, 17 and 20 are antennae-enriched; while SexiCXE5 and SexiCXE18 are dominantly expressed in wings, and SexiCXE31 is mainly expressed in proboscises, heads and legs. With the highly biased expression in antennae and proboscises, SexiCXE4 was selected for further functional assay. The recombinant SexiCXE4 were expressed in High-five cells and purified by a Ni(2+) affinity column. SexiCXE4 has much higher enzyme activity against plant volatiles (Z)-3-hexenyl acetate and hexyl acetate than to the sex pheromone components, suggesting that it may function mostly in the degradation of the plant volatiles.

Background The cotton aphid, Aphis gossypii Glover, is a destructive insect pest worldwide; it directly or indirectly damages (virus transmission) 300 species of host plants. Knowledge of their ecologically adaptive mechanisms at the molecular level may provide an essential and urgent method to effectively control this pest. However, no transcriptome information is available for the cotton aphid and sequence data are scarce. Therefore, we obtained transcriptome data. Results To facilitate such a study, two cotton aphid transcriptomes at different growth stages of cotton, seedling and summer, were sequenced. A total of 161,396 and 66,668 contigs were obtained and assembled into 83,671 and 42,438 transcripts, respectively. After combining the raw date for both transcriptomes, the sequences were reassembled into 66,695 transcripts, and 52,160 were annotated based on BLASTX analyses. Comparison of the transcriptomes revealed that summer presented less challenges for the cotton aphids than the seedling stage of cotton. In total, 58 putative heat shock protein genes and 66 candidate cytochrome p450 genes were identified with BLASTX. Conclusions Our results form a basis for exploring the molecular mechanisms of ecological adaption in the cotton aphid. Our study also provides a baseline for the exploration of abiotic stress responses. In addition, it provides large-scale sequence information for further studies on this species.

It is postulated that insect pheromone-binding proteins (PBPs) are involved in sex pheromone reception, while the general odorant-binding proteins (GOBPs) are involved in reception of the general odorants including plant volatiles. However, this functional specificity is not completely conclusive. In the present study, full-length sequences of two new OBP genes were molecularly identified as OachPBP1 and OachGOBP2 from Orthaga achatina, an important pest of the camphor tree Cinnamomum camphora. Quantification of transcript levels by qRT-PCR showed that the two genes highly expressed in antennae, with OachPBP1 male-biased and OachGOBP2 similar between sexes. These expression patterns are consistent with the generally proposed functions of PBPs and GOBPs. With the recombinant proteins obtained by a bacterial expression system, the binding specificity of these proteins was further investigated and compared using the competitive binding assay. OachPBP1 exhibited high binding affinities with all three putative sex pheromones and 10 pheromone analogs, supporting its role in pheromone reception. On the other hand, in addition to binding with some plant volatiles, OachGOBP2 surprisingly displayed similar or even higher binding affinities with the sex pheromones than OachPBP1. Therefore, we propose that OachGOBP2 might play roles in reception of sex pheromone. Additionally, plant volatiles farnesol and farnesene showed high binding with both OachGOBP2 and OachPBP1, suggesting that these volatile chemicals have regulatory functions in the behavior of O. achatina.

The complete mitochondrial genome of the tobacco budworm Helicoverpa assulta was sequenced. It is 15,351 bp in length, and contains 22 transfer RNA genes, 13 protein-coding genes (PCGs), 2 rRNA genes and a non-coding AT-rich region. The H. assulta mitogenome displays a very high level of identity (99.76%) with Helicoverpa armigera. The genome base composition is 40.50% A, 40.40% T, 11.32% C and 7.70% G. Similar to other Lepidoptera insects, all PCGs are initiated by a typical ATN codon, except CO1, which starts with CGA. Nine PCGs have the standard, complete stop codon (TAA), and the other four PCGs end with the incomplete stop codon TA–.

Background Chrysopa pallens (Rambur) are the most important natural enemies and predators of various agricultural pests. Understanding the sophisticated olfactory system in insect antennae is crucial for studying the physiological bases of olfaction and also could lead to effective applications of C. pallens in integrated pest management. However no transcriptome information is available for Neuroptera, and sequence data for C. pallens are scarce, so obtaining more sequence data is a priority for researchers on this species. Results To facilitate identifying sets of genes involved in olfaction, a normalized transcriptome of C. pallens was sequenced. A total of 104,603 contigs were obtained and assembled into 10,662 clusters and 39,734 singletons; 20,524 were annotated based on BLASTX analyses. A large number of candidate chemosensory genes were identified, including 14 odorant-binding proteins (OBPs), 22 chemosensory proteins (CSPs), 16 ionotropic receptors, 14 odorant receptors, and genes potentially involved in olfactory modulation. To better understand the OBPs, CSPs and cytochrome P450s, phylogenetic trees were constructed. In addition, 10 digital gene expression libraries of different tissues were constructed and gene expression profiles were compared among different tissues in males and females. Conclusions Our results provide a basis for exploring the mechanisms of chemoreception in C. pallens, as well as other insects. The evolutionary analyses in our study provide new insights into the differentiation and evolution of insect OBPs and CSPs. Our study provided large-scale sequence information for further studies in C. pallens.