Functional characterization of sensory neuron membrane protein 1a involved in sex pheromone detection of Apolygus lucorum(Hemiptera:Miridae)

The mirid bug Apolygus lucorum(Hemiptera:Miridae)is a polyphagous pest that affects a wide range of host plants.Its control remains challenging mainly due to its rapid reproduction,necessitating an understanding of sex pheromone communication.The recognition of sex pheromones is vital for courtship and mating behaviors,and is mediated by various chemosensory-associated proteins.Among these,sensory neuron membrane protein(SNMP),a CD36-related protein,is suggested to play crucial roles in detecting sex pheromones.In this study,we employed transcriptomic and genomic data from A.lucorum and phylogenetic approaches,and identified four putative SNMP genes(AlucSNMP1a,AlucSNMP1b,AlucSNMP2a,and AlucSNMP2b)with full open reading frames.Expression analysis revealed the ubiquitous presence of AlucSNMP transcripts in multiple tissues,with only AlucSNMP1a exhibiting male-biased expression in the antennae,suggesting its potential role in male chemosensation.Functional analysis using the Xenopus oocyte expression system,coupled with two-electrode voltage clamp recording,demonstrated that the co-expression of AlucSNMP1a with specific pheromone receptors(PRs)and the Odorant receptor co-receptor(Orco)significantly enhanced electrophysiological responses to sex pheromones compared to the co-expression of PRs and Orco alone.Moreover,the results indicated that the presence of AlucSNMP1a not only affected the responsiveness to sex pheromones but also influenced the kinetics(activation and inactivation)of the induced signals.In contrast,the co-expression of AlucSNMP1b with AlucPR/Orco complexes had no impact on the inward currents induced by two pheromone compounds.An examination of the selective pressures on SNMP1 genes across 20 species indicated strong purifying selection,implying potential functional conservation in various insects.These findings highlight the crucial role of AlucSNMP1a in the response to sex pheromones.

n-octyl acrylate is a candidate sex pheromone component involved in courtship in parasitoid wasp Microplitis mediator

Sex pheromones are considered to play critical roles in partner communication of most parasitic Hymenoptera.However,the identification of sex pheromone components remains limited to a few families of parasitoid wasps.In this study,we functionally characterized a candidate sex pheromone component in Microplitis mediator(Hymenoptera:Braconidae),a solitary parasitoid of Noctuidae insects.We found that the body surface extract from female wasps could significantly stimulate courtship behavior of males.Gas chromatography-electroantennographic detection(GC-EAD)analysis revealed that a candidate semiochemical from extract triggered significant electrophysiological response of antennae of males.By performing gas chromatography-mass spectrometer(GC-MS)measurement,GC-EAD active compound was identified as n-octyl acrylate,a candidate sex pheromone component in female M.mediator.In electroantennogram(EAG)tests,antennae of male wasps showed significantly higher electrophysiological responses to n-octyl acrylate than those of females.Y-tube olfactometer assays indicated that male wasps significantly chose n-octyl acrylate compared with the control.Furthermore,male wasps showed a remarkable preference for n-octyl acrylate in a simulated field condition behavioral trial;simultaneously,n-octyl acrylate standard could also trigger significant courtship behavior in males.We propose that n-octyl acrylate,as a candidate vital sex pheromone component,could be utilized to design behavioral regulators of M.mediator to implement the protection and utilization of natural enemies.

Molecular characterization and expression of sensory neuron membrane proteins in the parasitoid Microplitis mediator (Hymenoptera: Braconidae)

Sensory neuron membrane proteins(SNMPs),homologs of the human fatty acid transport protein CD36 family,are observed to play a significant role in chemoreception,especially in detecting sex pheromone in Drosophila and some lepidopteran species.In the current study,two full‐length SNMP transcripts,MmedSNMP1 and MmedSNMP2,were identified in the parasitoid Microplitis mediator(Hymenoptera:Braconidae).Quantitative real‐time polymerase chain reaction analysis showed that the expression of MmedSNMP1 was significantly higher in antennae than in other tissues of both sexes.In addition,the MmedSNMP1 transcript was increased dramatically in newly emerged adults and there were no significant differences between adults with or without mating and parasitic experiences.However,compared with MmedSNMP1,the expression of MmedSNMP2 was widely found in various tissues,significantly increased at half‐pigmented pupae stage and remained at a relatively constant level during the following developmental stages.It was found that MmedSNMP1 contained eight exons and seven introns,which was highly conserved compared with other insect species.In situ hybridization assay demonstrated that MmedSNMP1 transcript was distributed widely in antennal flagella.Among selected chemosensory genes(odorant binding protein,odorant receptor,and ionotropic receptor genes),MmedSNMP1 only partially overlapped with MmedORco in olfactory sensory neurons of antennae.Subsequent immunolocalization results further indicated that MmedSNMP1 was mainly expressed in sensilla placodea of antennae and possibly involved in perceiving plant volatiles and sex pheromones.These findings lay a foundation for further investigating the roles of SNMPs in the chemosensation of parasitoids.

Characterization and target gene analysis of microRNAs in the antennae of the parasitoid wasp Microplitis mediator

MicroRNAs(miRNAs),a class of non-coding single-strand RNA molecules encoded by endogenous genes,are about 21–24 nucleotides long and are involved in the post-transcriptional regulation of gene expression in plants and animals.Generally,the types and quantities of miRNAs in the different tissues of an organism are diverse,and these divergences may be related to their specific functions.Here we have identified 296 known miRNAs and 46 novel miRNAs in the antennae of the parasitoid wasp Microplitis mediator by high-throughput sequencing.Thirty-three miRNAs were predicted to target olfactory-associated genes,including odorant binding proteins(OBPs),chemosensory proteins,odorant receptors(ORs),ionotropic receptors(IRs)and gustatory receptors.Among these,17 miRNAs were significantly highly expressed in the antennae,four miRNAs were highly expressed both in the antennae and head or wings,while the remaining 12 miRNAs were mainly expressed in the head,thorax,abdomen,legs and wings.Notably,miR-9a-5p and miR-2525-3p were highly expressed in male antennae,whereas miR-1000-5p and novel-miR-13 were enriched in female antennae.The 17 miRNAs highly expressed in antennae are likely to be associated with olfaction,and were predicted to target one OBP(targeted by miR-3751-3p),one IR(targeted by miR-7-5p)and 14 ORs(targeted by 15 miRNAs including miR-6-3p,miR-9a-5p,miR-9b-5p,miR-29-5p,miR-71-5p,miR-275-3p,miR-1000-5p,miR-1000-3p,miR-2525-3p,miR-6012-3p,miR-9719-3p,novel-miR-10,novel-miR-13,novel-miR-14 and novel-miR-28).These candidate olfactory-associated miRNAs are all likely to be involved in chemoreception through the regulation of chemosensory gene expression in the antennae of M.mediator.

Effect of five diatomaceous earth formulations against Tribofium castaneum （Coleoptera： Tenebrionidae）, Oryzaephflus surinamensis （Coleoptera： Silvanidae） and Rhyzopertha dominica （Coleoptera： Bostrychidae）

Laboratory bioassays were conducted to determine the effect of food source on the survival of Tribolium castaneum Herbst, Oryzaephilus surinamensis L. and Rhyzopertha dominica F., after exposure to five diatomaceous earth （DE） formulations： Protect-It, Insecto, Perma-GuardTM, Dryacide and SilicoSec. Adults of these species were exposed to DEs at the rate of 0.5 mg/cm^2 for 1 day on filter paper inside plastic Petri dishes. After exposure, the initial mortality was counted and live individuals of the three species were held for a week in glass vials containing 50 mg wheat flour, rice and whole wheat, respectively. In the second experiment, after 1 day exposure to DEs, beetles were transferred to Petri dishes without food and held for a week to determine if the presence of food source would decrease the mortality of beetles. Experiments were carried out at 27℃ and 55% RH in the dark. The initial mortality in both of the experiments reached 100% for the three species exposed to Protect-It and in the case ofR. dominica and O. surinamensis exposed to Dryacide. In contrast, low level of mortality （〈 10%） was observed for T. castaneum exposed to Perma-GuardTM and Insecto. The mortality after the post-treatment period on food was decreased for the three species exposed to Perma-GuardTM and in the case of T. castaneum and R. dominica exposed to Insecto and SilicoSec. Adults of O. surinamensis were the most susceptible followed by R. dominica and 100% adult mortality was obtained, whereas T. castaneum were the least susceptible beetles to DEs. Protect-It and Dryacide were the most efficient DE formulations and can be used effectively in a stored grain integrated pest management program.