Odorant-binding Protein Genes Research Articles

The Neurotranscriptome of Monochamus alternatus.

The Japanese pine sawyer Monochamus alternatus serves as the primary vector for pine wilt disease, a devastating pine disease that poses a significant threat to the sustainable development of forestry in the Eurasian region. Currently, trap devices based on informational compounds have played a crucial role in monitoring and controlling the M. alternatus population. However, the specific proteins within M. alternatus involved in recognizing the aforementioned informational compounds remain largely unclear. To elucidate the spatiotemporal distribution of M. alternatus chemosensory-related genes, this study conducted neural transcriptome analyses to investigate gene expression patterns in different body parts during the feeding and mating stages of both male and female beetles. The results revealed that 15 genes in the gustatory receptor (GR) gene family exhibited high expression in the mouthparts, most genes in the odorant binding protein (OBP) gene family exhibited high expression across all body parts, 22 genes in the odorant receptor (OR) gene family exhibited high expression in the antennae, a significant number of genes in the chemosensory protein (CSP) and sensory neuron membrane protein (SNMP) gene families exhibited high expression in both the mouthparts and antennae, and 30 genes in the ionotropic receptors (IR) gene family were expressed in the antennae. Through co-expression analyses, it was observed that 34 genes in the IR gene family were co-expressed across the four developmental stages. The Antenna IR subfamily and IR8a/Ir25a subfamily exhibited relatively high expression levels in the antennae, while the Kainate subfamily, NMDA subfamily, and Divergent subfamily exhibited predominantly high expression in the facial region. MalIR33 is expressed only during the feeding stage of M. alternatus, the MalIR37 gene exhibits specific expression in male beetles, the MalIR34 gene exhibits specific expression during the feeding stage in male beetles, the MalIR8 and MalIR39 genes exhibit specific expression during the feeding stage in female beetles, and MalIR8 is expressed only during two developmental stages in male beetles and during the mating stage in female beetles. The IR gene family exhibits gene-specific expression in different spatiotemporal contexts, laying the foundation for the subsequent selection of functional genes and facilitating the full utilization of host plant volatiles and insect sex pheromones, thereby enabling the development of more efficient attractants.

The Female-Biased General Odorant Binding Protein 2 of Semiothisa cinerearia Displays Binding Affinity for Biologically Active Host Plant Volatiles.

Herbivorous insects rely on volatile chemical cues from host plants to locate food sources and oviposition sites. General odorant binding proteins (GOBPs) are believed to be involved in the detection of host plant volatiles. In the present study, one GOBP gene, ScinGOBP2, was cloned from the antennae of adult Semiothisa cinerearia. Reverse-transcription PCR and real-time quantitative PCR analysis revealed that the expression of ScinGOBP2 was strongly biased towards the female antennae. Fluorescence-based competitive binding assays revealed that 8 of the 27 host plant volatiles, including geranyl acetone, decanal, cis-3-hexenyl n-valerate, cis-3-hexenyl butyrate, 1-nonene, dipentene, α-pinene and β-pinene, bound to ScinGOBP2 (KD = 2.21-14.94 μM). The electrical activities of all eight ScinGOBP2 ligands were confirmed using electroantennography. Furthermore, oviposition preference experiments showed that eight host volatiles, such as decanal, cis-3-hexenyl n-valerate, cis-3-hexenyl butyrate, and α-pinene, had an attractive effect on female S. cinerearia, whereas geranyl acetone, 1-nonene, β-pinene, and dipentene inhibited oviposition in females. Consequently, it can be postulated that ScinGOBP2 may be implicated in the perception of host plant volatiles and that ScinGOBP2 ligands represent significant semiochemicals mediating the interactions between plants and S. cinerearia. This insight could facilitate the development of a chemical ecology-based approach for the management of S. cinerearia.

Molecular and cellular organization of odorant binding protein genes in Drosophila

Chemosensation is important for the survival and reproduction of animals. The odorant binding proteins (OBPs) are thought to be involved in chemosensation together with chemosensory receptors. While OBPs were initially considered to deliver hydrophobic odorants to olfactory receptors in the aqueous lymph solution, recent studies suggest more complex roles in various organs. Here, we use GAL4 transgenes to systematically analyze the expression patterns of all 52 members of the Obp gene family and 3 related chemosensory protein genes in adult Drosophila, focusing on chemosensory organs such as the antenna, maxillary palp, pharynx, and labellum, and other organs such as the brain, ventral nerve cord, leg, wing, and intestine. The OBPs were observed to express in diverse organs and in multiple cell types, suggesting that these proteins can indeed carry out diverse functional roles. Also, we constructed 10 labellar-expressing Obp mutants, and obtained behavioral evidence that these OBPs may be involved in bitter sensing. The resources we constructed should be useful for future Drosophila OBP gene family research.

Identifying the Key Role of Plutella xylostella General Odorant Binding Protein 2 in Perceiving a Larval Attractant, (E,E)-2,6-Farnesol.

There is currently a lack of effective olfaction-based techniques to control diamondback moth (DBM) larvae. Identifying behaviorally active odorants for DBM larvae and exploring their recognition mechanisms can provide insights into olfaction-based larval control strategies. Through the two-choice assay, (E,E)-2,6-farnesol (farnesol) was identified as a compound exhibiting significant attractant activity toward DBM larvae, achieving an attraction index of 0.48 ± 0.13. PxylGOBP1 and PxylGOBP2, highly expressed in the antennae of DBM larvae, both showed high affinity toward farnesol. RNAi technology was used to knock down PxylGOBP1 and PxylGOBP2, revealing that the attraction of DBM larvae to farnesol nearly vanished following the knockdown of PxylGOBP2, indicating its critical role in recognizing farnesol. Further investigation into the PxylGOBP2-farnesol interaction revealed the importance of residues like Thr9, Trp37, and Phe118 in PxylGOBP2's binding to farnesol. This research is significant for unveiling the olfactory mechanisms of DBM larvae and developing larval behavior regulation techniques.

Genetic and behavioral differences between above and below ground Culex pipiens bioforms.

Efficiency of mosquito-borne disease transmission is dependent upon both the preference and fidelity of mosquitoes as they seek the blood of vertebrate hosts. While mosquitoes select their blood hosts through multi-modal integration of sensory cues, host-seeking is primarily an odor-guided behavior. Differences in mosquito responses to hosts and their odors have been demonstrated to have a genetic component, but the underlying genomic architecture of these responses has yet to be fully resolved. Here, we provide the first characterization of the genomic architecture of host preference in the polymorphic mosquito species, Culex pipiens. The species exists as two morphologically identical bioforms, each with distinct avian and mammalian host preferences. Cx. pipiens females with empirically measured host responses were prepared into reduced representation DNA libraries and sequenced to identify genomic regions associated with host preference. Multiple genomic regions associated with host preference were identified on all 3 Culex chromosomes, and these genomic regions contained clusters of chemosensory genes, as expected based on work in Anopheles gambiae complex mosquitoes and in Aedes aegypti. One odorant receptor and one odorant binding protein gene showed one-to-one orthologous relationships to differentially expressed genes in A. gambiae complex members with divergent host preferences. Overall, our work identifies a distinct set of odorant receptors and odorant binding proteins that may enable Cx. pipiens females to distinguish between their vertebrate blood host species, and opens avenues for future functional studies that could measure the unique contributions of each gene to host preference phenotypes.

Odorant Binding Protein Expressed in Legs Enhances Malathion Tolerance in Bactrocera dorsalis (Hendel).

Bactrocera dorsalis is a highly invasive species and is one of the most destructive agricultural pests worldwide. Organophosphorus insecticides have been widely and chronically used to control it, leading to the escalating development of resistance. Recently, odorant binding proteins (OBPs) have been found to play a role in reducing insecticide susceptibility. In this study, we used RT-qPCR to measure the expression levels of four highly expressed OBP genes in the legs of B. dorsalis at different developmental stages and observed the effect of malathion exposure on their expression patterns. The results showed that OBP28a-2 had a high expression level in 5 day old adults of B. dorsalis, and its expression increased after exposure to malathion. By CRISPR/Cas9 mutagenesis, we generated OBP28a-2-/- null mutants and found that they were more susceptible to malathion than wild-type adults. Furthermore, in vitro direct affinity assays confirmed that OBP28a-2 has a strong affinity for malathion, suggesting that it plays a role in reducing the susceptibility of B. dorsalis to malathion. Our findings enriched our understanding of the function of OBPs. The results highlighted the potential role of OBPs as buffering proteins that help insects survive exposure to insecticides.

Molecular and Functional Characterization of Three General Odorant-Binding Protein 2 Genes in Cydia pomonella (Lepidoptera: Tortricidae).

General odorant-binding proteins (GOBPs) play a crucial role in the detection of host plant volatiles and pheromones by lepidopterans. Previous studies identified two duplications in the GOBP2 gene in Cydia pomonella. In this study, we employed qRT-PCR, protein purification, and fluorescence competitive binding assays to investigate the functions of three GOBP2 genes in C. pomonella. Our findings reveal that CpomGOBP2a and CpomGOBP2b are specifically highly expressed in antennae, while CpomGOBP2c exhibits high specific expression in wings, suggesting a potential divergence in their functions. Recombinant proteins of CpomGOBP2a, CpomGOBP2b, and CpomGOBP2c were successfully expressed and purified, enabling an in-depth exploration of their functions. Competitive binding assays with 20 host plant volatiles and the sex pheromone (codlemone) demonstrated that CpomGOBP2a exhibits strong binding to four compounds, namely butyl octanoate, ethyl (2E,4Z)-deca-2,4-dienoate (pear ester), codlemone, and geranylacetone, with corresponding dissolution constants (Ki) of 8.59993 μM, 9.14704 μM, 22.66298 μM, and 22.86923 μM, respectively. CpomGOBP2b showed specific binding to pear ester (Ki = 17.37481 μM), while CpomGOBP2c did not exhibit binding to any tested compounds. In conclusion, our results indicate a functional divergence among CpomGOBP2a, CpomGOBP2b, and CpomGOBP2c. These findings contribute valuable insights for the development of novel prevention and control technologies and enhance our understanding of the evolutionary mechanisms of olfactory genes in C. pomonella.

Odorant-Binding and Chemosensory Proteins in Fig Wasps: Evolutionary Insights From Comparative Studies.

Fig wasps (Agaonidae; Hymenoptera) are the only pollinating insects of fig trees (Ficus; Moraceae), forming the most closely and highly specific mutualism with the host. We used transcriptome sequences of 25 fig wasps from six genera to explore the evolution of key molecular components of fig wasp chemosensory genes: odorant-binding proteins (OBPs) and chemosensory proteins (CSPs). We identified a total 321 OBPs and 240 CSPs, with each species recording from 6 to 27 OBP genes and 6-19 CSP genes. 318 OBP genes are clustered into 17 orthologous groups and can be divided into two groups: PBP sensitive to pheromone and GOBP sensitive to general odor molecules, such as alcohols, esters, acids, ketones, and terpenoids. 240 CSP genes are clustered into 12 orthologous groups, which can be divided into three major groups and have functions, such as olfactory, tissue formation and/or regeneration, developmental, and some specific and unknown function. The gene sequences of most orthologous groups vary greatly among species and are consistent with the phylogenetic relationships between fig wasps. Strong purifying selection of both OBP and CSP genes was detected, as shown by low ω values. A positive selection was detected in one locus in CSP1. In conclusion, the evolution of chemosensory proteins OBPs and CSPs in fig wasps is relatively conservative, and they play an indispensable role in the life activities of fig wasps. Our results provide a starting point for understanding the molecular basis of the chemosensory systems of fig wasps.

Genome-Wide Identification of the Genes of the Odorant-Binding Protein Family Reveal Their Role in the Olfactory Response of the Tomato Leaf Miner (Tuta absoluta) to a Repellent Plant

The remarkable biological and evolutionary adaptations of insects to plants are largely attributed to the powerful chemosensory systems of insects. The tomato leaf miner (Tuta absoluta) is a destructive invasive pest with a global distribution that poses a serious threat to the production of nightshade crops, especially tomatoes. Functional plants can attract or repel insect pests by releasing volatiles that interact with the olfactory system of insects, thereby reducing the damage of insect pests to target crops. However, there is limited research on the interaction between T. absoluta olfactory genes and functional plants. In this study, 97 members of the putative odorant-binding protein (OBP) family have been identified in the whole genome of T. absoluta. Phylogenetic analysis involving various Lepidopteran and Dipteran species, including D. melanogaster, revealed that OBP gene families present conserved clustering patterns. Furthermore, the Plus-C subfamily of OBP showed extremely significant expansion. Moreover, the expression levels of the OBP genes varied significantly between different developmental stages; that is, the highest number of OBP genes were expressed in the adult stage, followed by the larval stage, and fewer genes were expressed in high abundance in the egg stage. On the other hand, through a Y-tube olfactometer, we identified a functional plant—Plectranthus tomentosa—that significantly repels adult and larval T. absoluta. Finally, we screened the OBP genes in response to tomato and P. tomentosa volatiles at the genomic level of T. absoluta using RT-qPCR. These results laid a good foundation for controlling T. absoluta with functional plants and further studying olfactory genes.

An OBP gene highly expressed in non-chemosensory tissues affects the phototaxis and reproduction of Spodoptera frugiperda.

Insect odorant binding proteins (OBPs) were initially regarded as carriers of the odorants involved in chemosensation. However, it had been observed that a growing number of OBP genes exhibited broad expression patterns beyond chemosensory tissues. Here, an OBP gene (OBP31) was found to be highly expressed in the larval ventral nerve cord, adult brain and male reproductive organ of Spodoptera frugiperda. An OBP31 knockout strain (OBP31-/- ) was generated by CRISPR/Cas9 mutagenesis. For OBP31-/- , the larvae needed longer time to pupate, but there was no difference in the pupal weight between OBP31-/- and wild type (WT). OBP31-/- larvae showed stronger phototaxis than the WT larvae, indicating the importance of OBP31 in light perception. For mating rhythm of adults, OBP31-/- moths displayed an earlier second mating peak. In the cross-pairing of OBP31-/- and WT moths, the mating duration was longer, and hatchability was lower in OBP31-/- group and OBP31+/- ♂ group than that in the WT group. These results suggested that OBP31 played a vital role in larval light perception and male reproductive process and could provide valuable insights into understanding the biological functions of OBPs that were not specific in chemosensory tissues.

Minus-C odorant binding protein TcasOBP7G contributes to reproduction and defense against phytochemical in the red flour beetle, Tribolium castaneum

Insect odorant binding proteins (OBPs) paly an important role in the recognition and transmission signals. One odorant binding protein gene, TcasOBP7G, was significantly up-regulated after exposure to eucalyptol. Nevertheless, the causal role of TcasOBP7G in Tribolium castaneum is still unclear. Here, TcasOBP7G was cloned and characterized from T. castaneum, aligning with the typical characteristic of Minus-C OBPs. The analysis of gene expression profile showed that TcasOBP7G is highly expressed at late larval and adult stage, with particularly high expression levels in ovary and testis of adults, suggesting that TcasOBP7G might be associated with reproduction of T. castaneum. Furthermore, RNAi of TcasOBP7G led to a decrease in spawning, while the development of gonads was normal, suggesting that TcasOBP7G may medicate the reproduction of T. castaneum. Additionally, the mRNA level of TcasOBP7G was significantly increased after exposure to eucalyptol, and knockdown of TcasOBP7G resulted in an increased susceptibility of T. castaneum to eucalyptol, indicating that TcasOBP7G is associated with the tolerance of T. castaneum to eucalyptol. Altogether, these results indicate that TcasOBP7G is required for both reproduction and defense against phytochemical in T. castaneum. This study will provide a theoretical basis for the development of novel control strategies based on the olfaction of insects.

The binding affinity of two general odorant binding proteins in Spodoptera frugiperda to general volatiles and insecticides

Spodoptera frugiperda is a kind of polyphagous pest, and can damage a large number different host plants around the worldwide. The molecular mechanisms of two general odorant binding proteins (GOBPs) binding with general volatiles and insecticides are still blank. In this study, we investigated the function of two GOBPs in S. frugiperda, by expressing two SfruGOBPs and tested the binding affinities by the fluorescence competition binding assays. The results exhibited that SfruGOBP1 has binding affinities to 4 of 38 general volatiles and 3 of 7 insecticides. In contrast, SfruGOBP2 showed a broader ligand-binding spectrum to 21 volatiles and 4 insecticides, suggesting SfruGOBP2 may plays a more important role in perceiving host volatiles than SfruGOBP1. Furthermore, we used molecular docking and site-directed mutagenesis assay to explored the key amino acid residues of two SfruGOBP to insecticides ligand. This study provides some valuable information to exploring the olfactory mechanism of two GOBPs bound the host plant volatiles and insecticides in S. frugiperda.

Selection Behavior and OBP-Transcription Response of Western Flower Thrips, Frankliniella occidentalis, to Six Plant VOCs from Kidney Beans.

Plant volatile organic compounds (VOCs) are an important link that mediates chemical communication between plants and plants, plants and insects, and plants and natural enemies of insect pests. In this study, we tested the response in the selective behavior of western flower thrips, Frankliniella occidentalis, to the VOCs of kidney bean, Phaseolus vulgaris L., to explore their "attraction" or "repellent" effects regarding their application in integrated pest management (i.e., IPM). The results indicated that 12.7 μL/mL (E, E, E, E)-squalene, 3.2 μL/mL dioctyl phthalate, and 82.2 μL/mL ethyl benzene had a significantly attractive effect on the selective behavior of F. occidentalis, while 10.7 μL/mL and 21.4 μL/mL 2,6-ditert-butyl-4-methyl phenol had a significantly repulsive effect on the selective behavior of F. occidentalis, showing that F. occidentalis responds differently to specific concentrations of VOCs from P. vulgaris plant emissions. Interestingly, the three compounds with the specific above concentrations, after being mixed in pairs, significantly attracted F. occidentalis compared to the control treatment; however, the mixture with the three above compounds had no significant different effect on F. occidentalis compared to the control treatment. It can be seen that the effect with the mixtures of three kinds of VOCs had the same function and may not get better. Simultaneously, the reasons for this result from the transcription levels of odorant-binding protein genes (OBPs) were determined. There were differences in the types and transcription levels of OBPs, which played a major role in the host selection behavior of F. occidentalis under the mixed treatment of different VOCs. It is presumed that there are specific VOCs from P. vulgaris plants that have a good repellent or attracting effect on the selective behavior of F. occidentalis, which can be used for the development of plant-derived insect attractants and repellents to serve as IPM in fields. But attention should be paid to the antagonism between plant-derived preparations and VOCs produced by plants themselves after application.

Antennal transcriptome analysis of odorant-binding proteins and characterization of GOBP2 in the variegated cutworm Peridroma saucia.

Odorant-binding proteins (OBPs) are expressed at extremely high concentrations in the chemo-sensilla lymph of insects and have long been thought to be crucial for delivering the semiochemicals to the odorant receptors. They are represented by multiple classes: general odorant-binding proteins (GOBP1 and GOBP2) and pheromone-binding proteins. In the current study, we identified a total of 35 OBPs in the antennal transcriptome of Peridroma saucia, a worldwide pest that causes serious damage to various crops. A gene expression value (TPM, transcripts per million) analysis revealed that seven OBPs (PsauPBP1/2/3, PsauGOBP1/2, PsauOBP6, and PsauOBP8) were highly abundant in the antennae. Next, we focused on the expression and functional characterization of PsauGOBP2. Real-time quantitative-PCR analysis demonstrated that PsauGOBP2 was predominantly expressed in the antennae of both sexes. Fluorescence binding assays showed that the recombinant PsauGOBP2 strongly binds to the female sex pheromone components Z11-16: Ac (Ki = 4.2μM) and Z9-14: Ac (Ki = 4.9μM) and binds moderately (6µM ≤ Ki ≤ 13µM) to the host plant volatiles phenylethyl acetate, β-myrcene, and dodecanol. Further 3D structural modeling and molecular docking revealed that several crucial amino acid residues are involved in ligand binding. The results not only increase our understanding of the olfactory system of P. saucia but also provide insights into the function of PsauGOBP2 that has implications for developing sustainable approaches for P. saucia management.

Identification and characterization of odorant receptors in Plutella xylostella antenna response to 2,3-dimethyl-6-(1-hydroxy)-pyrazine

Diamondback moth (Plutella xylostella), a worldwide migratory pest that is developing strong resistance to various chemical insecticides. It has been determined that four natural pyrazines isolated from Allium tuberosum showed significant repellent activity to P. xylostella, but the molecular target still unknown. In the present study, a novel synthetic route for 2,3-dimethyl-6-(1-hydroxy)-pyrazine which has the most significant repellent activity with a purity of 90.60% was established. Simultaneously, the bioassay result declared that the repellent grade was IV at a dosage of 0.01 mg which was the same as to the published data. Transcriptomics analysis detected 1643 upregulated and 3837 downregulated genes in P. xylostella antennae following this pyrazine exposure. Then, 2142 differentially expressed genes were annotated using Gene Ontology and 2757 genes were annotated by Kyoto Encyclopedia of Genes and Genomes. Moreover, this procedure identified 84 odour perception-related genes, 58 odorant receptor (OR) genes including 57 conventional ORs and the odorant receptor co-receptor (Orco, atypical odorant receptor) gene, and 26 odorant-binding protein (OBP) genes. Based on quantitative real time PCR (RT-qPCR) and differential expression results, 9 OR genes including the Orco were cloned and characterised. In summary, this study provides important basis for the utilization of pyrazines as the main active ingredients or lead compounds to developing new botanical pesticides, which will reduce application of chemical pesticides and postpone the development of resistance.

Roles of GOBP1 in perception of host plant volatiles revealed by CRISPR/Cas9‐mediated mutagenesis in Spodoptera litura

AbstractThe general odorant‐binding proteins (GOBPs) in Lepidoptera form a distinct OBP subclass, which exclusively has two members (GOBP1 and GOBP2) in a single species. GOBPs are long thought to function in perception of plant odorants and recently in sex pheromones, but the in vivo evidence is rarely provided. Here, by using CRISPR/Cas9 technique, we demonstrated that GOBP1 in a noctuid moth Spodoptera litura is involved in perception of host plant volatiles but not of the female sex pheromone, and GOBP1 in females is less important than GOBP2 in host plant perception. First, the GOBP1 knockout strain (GOBP1−/−) of S. litura was obtained by CRISPR/Cas9 technique. Second, in the electroantennography (EAG) tests, GOBP1−/− females displayed significant reduction in responses to plant volatile farnesene, and leaf discs of the host plant cabbage (males only to high leaf doses), but GOBP1−/− adults did not show significant EAG reduction to the major sex pheromone component and two pheromone analogues. Third, compared with GOBP2−/− females, GOBP1−/− females were significantly lower in reduction (in relation to WT moths) of EAG response to host plant leaves. Our results provide in vivo evidence that GOBP1 is involved in perception of host plant volatiles but is less important than GOBP2 in S. litura.

Expression profile and functional characterization of odorant binding proteins in a forest pest, Dioryctria abietella (Lepidoptera: Pyralidae).

In the forest ecosystem dominated by the Pinaceae plants, this boring pest Dioryctria abietella is subject to a variety of odorants derived from host and nonhost plants, in which olfactory-related proteins enriched in antennae are key behavioral modulators for the orientation of feeding and ovipositing hosts. Here, we addressed the odorant binding protein (OBP) gene family in D. abietella. Expression profiles revealed that the majority of OBPs were abundantly expressed in the antennae at a female-biased level. A male-antenna-biased DabiPBP1 was a strong candidate for detecting type I and type II pheromones of D. abitella female moths. Using a prokaryotic expression system combined with affinity chromatography, we harvested two antenna-dominant DabiOBPs. In the ligand-binding assays, the two DabiOBPs exhibited different odorant response spectra, as DabiOBP17 was tuned to most odorants with higher affinities compared to DabiOBP4. Of these, DabiOBP4 could strongly bind syringaldehyde and citral (dissociation constants (Ki)<14μM). A floral volatile, benzyl benzoate (Ki=4.72±0.20μM), was the best ligand for DabiOBP17. Remarkably, several green leaf volatiles were found to strongly interact with DabiOBP17 (Ki<8.5μM), including Z3-hexenyl acetate, E2-hexenol, Z2-hexenal and E2-hexenal that may mediate a repellent response to D. abietella. Structural analyses of ligands revealed that the binding of the two DabiOBPs to odorants was associated with carbon-chain lengths and functional groups. Molecular simulations identified several key residues involved in the interactions of DabiOBPs and ligands, suggesting specific binding mechanisms. This study highlights olfactory roles of two antennal DabiOBPs in D. abietella, helping the identification of potentially behavioral compounds for the population control of this pest.

Functional differentiation of two general-odorant binding proteins in Hyphantria cunea (Drury) (Lepidoptera: Erebidae).

General odor-binding proteins (GOBPs) play critical roles in insect olfactory recognition of sex pheromones and plant volatiles. Therefore, the identification of GOBPs in Hyphantria cunea (Drury) based on their characterization to pheromone components and plant volatiles is remain unknown. In this study, two H. cunea (HcunGOBPs) genes were cloned, and their expression profiles and odorant binding characteristics were systematically analyzed. Firstly, the tissue expression study showed that both HcunGOBP1 and HcunGOBP2 were highly expressed in the antennae of both sexes, indicating their potential involvement in the perception of sex pheromones. Secondly, these two HcunGOBPs genes were expressed in Escherichia coli and ligand binding assays were used to assess the binding affinities to its sex pheromone components including two aldehydes and two epoxides, and some plant volatiles. HcunGOBP2 showed high binding affinities to two aldehyde components (Z9, Z12, Z15-18Ald and Z9, Z12-18Ald), and showed low binding affinities to two epoxide components (1, Z3, Z6-9S, 10R-epoxy-21Hy and Z3, Z6-9S, 10R-epoxy-21Hy), whereas HcunGOBP1 showed weak but significant binding to all four sex pheromone components. Furthermore, both HcunGOBPs demonstrated variable binding affinities to the plant volatiles tested. Thirdly, in silico studies of HcunGOBPs utilized homology, structure modeling, and molecular docking revealed critical hydrophobic residues might be involved in the binding of HcunGOBPs to their sex pheromone components and plant volatiles. Our study suggests that these two HcunGOBPs may serve as potential targets for future studies of HcunGOBPs ligand binding, providing insight in the mechanism of olfaction in H. cunea. © 2023 Society of Chemical Industry.

Identification and characterization of odorant‐binding protein genes in the brown‐winged green bug, Plautia stali

AbstractThe brown‐winged green bug, Plautia stali Scott (Hemiptera: Pentatomidae), is a major pest in orchards in Japan and Korea. Large numbers of adults are attracted by aggregation pheromone [methyl (E,E,Z)‐2,4,6‐decatrienoate] and cause extensive damage to the fruit. To identify olfactory‐related genes in P. stali, we performed RNA sequencing (RNA‐seq) of adult antennae. Among the 27299 contigs, 16 putative odorant‐binding protein (OBP) genes were identified and designated as PstaOBP1–16, including 10 classic (i.e., containing six conserved cysteines) and six Plus‐C (i.e., containing more than six cysteines) OBP genes. Quantitative reverse‐transcription polymerase chain reaction (qRT‐PCR) revealed that four genes (PstaOBP1, 3, 6, and 7) were strongly expressed in the antennae. PstaOBP2 was detected throughout the insect body, with higher expression in the antennae, stylet, and tarsus. The remaining OBP genes were expressed in very small amounts. PstaOBP1, which showed the highest expression in adult antennae, was expressed in Escherichia coli (Migula) Castellani &amp; Chalmers cells and the recombinant protein was subjected to a ligand‐binding competitive assay. PstaOBP1 bound to methyl (E,E,Z)‐2,4,6‐decatrienoate with high affinity. Of the other 26 compounds tested, hinokitiol and tropolone bound strongly to PstaOBP1. Hinokitiol is an odor component of hinoki cypress, a host tree of P. stali. Electrophysiological responses to methyl (E,E,Z)‐2,4,6‐decatrienoate and hinokitiol were detected using adult P. stali antennae. Combined, these results suggest that PstaOBP1 is involved in P. stali responses to pheromone and host plant volatiles.

Breeding honey bees (Apis mellifera L.) for low and high Varroa destructor population growth: Gene expression of bees performing grooming behavior.

Social organisms, including honey bees (Apis mellifera L.), have defense mechanisms to control the multiplication and transmission of parasites and pathogens within their colonies. Self-grooming, a mechanism of behavioral immunity, seems to contribute to restrain the population growth of the ectoparasitic mite Varroa destructor in honey bee colonies. Because V. destructor is the most damaging parasite of honey bees, breeding them for resistance against the mite is a high priority of the beekeeping industry. A bidirectional breeding program to select honey bee colonies with low and high V. destructor population growth (LVG and HVG, respectively) was conducted. Having high and low lines of bees allowed the study of genetic mechanisms underlying self-grooming behavior between the extreme genotypes. Worker bees were classified into two categories: 'light groomers' and 'intense groomers'. The brains of bees from the different categories (LVG-intense, LVG-light, HVG-intense, and HVG-light) were used for gene expression and viral quantification analyses. Differentially expressed genes (DEGs) associated with the LVG and HVG lines were identified. Four odorant-binding proteins and a gustatory receptor were identified as differentially expressed genes. A functional enrichment analysis showed 19 enriched pathways from a list of 219 down-regulated DEGs in HVG bees, including the Kyoto Encyclopedia of Genes and Genomes (KEGG) term of oxidative phosphorylation. Additionally, bees from the LVG line showed lower levels of Apis rhabdovirus 1 and 2, Varroa destructor virus -1 (VDV-1/DWV-B), and Deformed wing virus-A (DWV-A) compared to bees of the HVG line. The difference in expression of odorant-binding protein genes and a gustatory receptor between bee lines suggests a possible link between them and the perception of irritants to trigger rapid self-grooming instances that require the activation of energy metabolic pathways. These results provide new insights on the molecular mechanisms involved in honey bee grooming behavior. Differences in viral levels in the brains of LVG and HVG bees showed the importance of investigating the pathogenicity and potential impacts of neurotropic viruses on behavioral immunity. The results of this study advance the understanding of a trait used for selective breeding, self-grooming, and the potential of using genomic assisted selection to improve breeding programs.