Electroantennogram Responses Research Articles

Odor-Pollution From Fungicides Disrupts Learning and Recognition of a Common Floral Scent in Bumblebees (Bombus impatiens)

Background and AimsBumblebees provide vital pollination services to both natural and agricultural ecosystems. Consequently their declines in species-diversity and population size over the last five decades is alarming. Direct contributors to these declines include pesticides, habitat loss, and disease. However, given that colony fitness is linked to foraging success, successful conservation requires mitigation of any anthropogenic practices that negatively impact foraging. Previous work has shown that agrochemical odor-pollution, including that of fungicides, can modulate bumblebee foraging behavior. This study investigates how odor pollution from three common fungicides (Safer® Brand Garden Fungicide II, Scotts® Lawn Fungus Control, and Reliant® Systemic Fungicide) affects Bombus impatiens’ floral-odor learning and recognition using an associative learning paradigm.MethodsThe effects of fungicide-odor pollution were tested in three ways: (1) background pollution during floral-odor learning; (2) background pollution during floral-odor recognition; and (3) point (localized) pollution during floral-odor recognition. Electroantennogram (EAG) recordings from B. impatiens confirmed the salience of all odor-stimuli and examined impacts of background fungicide-odor on antennal responses to floral-odor. To better understand how fungicide-odor structure related to behavioral data, scents were sampled (Solid Phase Microextraction) and analyzed using gas chromatography–mass spectrometry. Odors were then characterized using the Compounds Without Borders (CWB) vectorization method.ConclusionAll fungicides tested disrupted floral-odor learning and recognition for at least one concentration tested, and Scotts® was universally disruptive at all tested concentrations. All fungicides induced EAG responses, indicating they provide perceivable odor stimuli. Interestingly, two of three tested fungicides (Scotts® and Reliant®) inhibit antennal responses to Monarda fistulosa odor. Odor characterization supports previous findings that sulfurous scents could be disruptive to odor-driven foraging behaviors. Inability for foraging bumblebees to associate to rewarding floral odors in the presence of fungicidal odor pollution could have negative large-scale implications for colony health and reproductive fitness.

Identification and functional analysis of a chemosensory protein from Bactrocera minax (Diptera: Tephritidae).

Olfaction has an indispensable role in insect behavior, enabling location of suitable host plants and oviposition sites, finding mates and evasion of natural enemies. Chemosensory proteins (CSPs) function to screen external odorants and transport them to olfactory receptor neurons, thereby increasing the sensitivity of the olfactory system. At present, CSP genes have been identified in many insect species, but there are relatively few studies on the function of CSP, especially in Tephritidae. In this study, we sequenced and analyzed 12 transcriptomes of Bactrocera minax and identified five CSP genes. The results of polymerase chain reactions with reverse transcription showed that BminCSP3 was highly expressed only in antennae. Results from competitive binding experiments showed that BminCSP3 has good binding ability to citral compared with 23 other volatile organic compounds. The docking model with citral showed hydrogen bond formation with residues (ARG97); however, no hydrogen bonds were formed in the docking of five other ligands (furfuryl alcohol, linalool, cis-3-hexenyl acetate, (R)-(+)-limonene and (+)-carvone). Electroantennogram (EAG) analyses revealed that citral was active in B. minax at the antennal level, and the EAG response value of female adults was significantly higher than that of male adults. Furthermore, the results of behavioral bioassays showed that females were significantly attracted to citral. Our results suggest that BminCSP3 plays an important role in the recognition of citral by B. minax adults. © 2022 Society of Chemical Industry.

Identification of a unique three-component sex pheromone produced by the tea black tussock moth, Dasychira baibarana (Lepidoptera: Erebidae: Lymantriinae).

The tea black tussock moth Dasychira baibarana Matsumura is a devastating pest in tea plantations that causes substantial economic losses. Presently, there is no effective method to control this pest other than pesticide application. The identified sex pheromone of D. baibarana could be used for detecting and monitoring this pest. Gas chromatography-electroantennogram detection showed that D. baibarana male moth antennae responded strongly to three components in the female sex pheromone gland, which were identified as: (3Z,6Z)-cis-9,10-epoxyhenicosa-3,6-diene (Z3,Z6,epo9-21:H) (I), (3Z,6Z,11E)-cis-9,10-epoxyhenicosa-3,6,11-triene (Z3,Z6,epo9,E11-21:H) (II) and (3Z,6Z)-henicosa-3,6-dien-11-one (Z3,Z6-21:11-one) (III). Dasychira baibarana uses a unique composition of an epoxydiene, epoxytriene, and dienone with the same 3,6-dienyl motif as its sex pheromone. The epoxytriene and dienone were not previously characterized as insect pheromone components. Electroantennogram analysis showed that each synthetic compound strongly stimulated male antennae, and compounds II and III elicited stronger responses than compound I. A wind tunnel bioassay and field trapping experiments proved that, ternary blends of compounds I-III attracted D. baibarana. Efficient attraction was achieved with a rubber septum baited with 500 μg of a mixture of compounds I-III at the ratio 25:20:55. The three identified compounds elicited an electroantennogram response in D. baibarana male moth antennae, and a mixture of the three components at the ratio 25:20:55 attracted D. baibarana male moths in a wind tunnel assay and field trapping experiments. This blend could be used for integrated management of D. baibarana in tea plantations. © 2022 Society of Chemical Industry.

Encounter With a Selfish Virus Sabotages Its Vector to Orient Toward Requisite Host Plant: A Case Study With Chili Leaf Curl Virus-Whitefly

Interactions of a virus with its vector and host plant have challenged entomologists, pathologists and biologists alike. Phytophagous insects depend on specific host volatile cues to locate suitable host plants for feeding and oviposition. Several studies have revealed that plant viruses modify their insect vector’s orientation toward specific host plants to facilitate their spread and survival. The ecological and molecular basis of this vector behavior modification remains largely unknown and was therefore explored in this study. Interestingly, host volatile preference for non-viruliferous female whiteflies [Bemisia tabaci (Genn.)] was found to be preferentially oriented toward infected chili plant [with chili leaf curl (ChLCV)] volatiles, while viruliferous whiteflies preferred healthy chili plant (Capsicum annum L.) volatiles in olfactometer. The electrophysiological studies involving electroantennogram (EAG) assays exhibited similar trend in EAG response amplitudes. Gas Chromatography linked electroantennodetection (GC EAD) revealed specific plant volatile cues responsible for altered host orientation behavior of the vector. Transcriptome profiling of the viruliferous and non-viruliferous whiteflies and Realtime qPCR validation showed differential expression of certain odorant binding proteins (OBPs) in viruliferous whiteflies. Our results suggest that there is a plant virus mediated altered chemoecological behavior in the vector with respect to orientation toward its host plant. Based on the findings we speculate that the virus mediates such change in the vector for a continued transmission success to the host.

Molecular characterization and its binding properties of general odorant binding protein 2 in Dioryctria abietella

AbstractDioryctria abietella is a major pest of conifer cones that can initiate specific behaviours (locating hosts, finding mates and selecting oviposition sites) through sensitive olfactory systems. General odorant‐binding proteins are involved in the first step in odorant perception in insects, but their specific functional roles in D. abietella chemoreception have not been fully elucidated. In this study, quantitative real‐time polymerase chain reaction (PCR) examinations showed that DabiGOBP2 is predominantly expressed in the antennae of female adults. Western blot experiments further confirmed that these proteins show biased expression in the tissues. Binding property analysis demonstrated that DabiGOBP2 exhibited ligand specificity to terpenes, especially myrcene (Ki = 8.19 ± 0.05). Electroantennogram (EAG) and Y‐tube assays showed that three plant volatiles elicited high EAG responses, but only dodecanal showed a strong attraction to females. However, myrcene and isobutyl benzoate were significantly repellent to males at high doses in the behaviour test. In conclusion, exploring the functions of DabiGOBP2 can help in better understanding the mechanism of the olfactory system of D. abietella and developing novel strategies to control this pest.

Effect of Soybean Volatiles on the Behavior of the Bean Bug, Riptortus pedestris.

The bean bug, Riptortus pedestris, is a polyphagous insect that feeds primarily on leguminous plants, especially soybean (Glycine max). Although the bean bug is an economically important pest of soybean, little is known about how the insect locates soybean fields. In this study, we examined the electroantennogram responses of R. pedestris to soybean volatiles and examined the behavioral responses of the adult bean bugs. R. pedestris adults were attracted more to their host-plant soybean, even when physical contact was absent, than to air or a non-host plant. Accordingly, we hypothesized that R. pedestris can recognize soybean through a plant's volatile organic compounds (VOCs). Five VOCs were identified from intact soybean plants at the vegetative stage: (Z)-3-hexen-1-ol, (Z)-3-hexenyl acetate, 4-ethylbenzaldehyde, α-farnesene, and methyl salicylate. Response spectra of the antennae to these volatiles clearly showed that both male and female R. pedestris can detect soybean volatiles. The adult bean bugs did not show behavioral orientation to any individual compounds but showed significant orientation to a particular blend of synthetic soybean volatiles when tested under laboratory conditions. In the field, this soybean volatile blend did not significantly attract the bean bugs, but it did interact synergistically with the aggregation pheromone to attract the bean bugs. These results highlight the role of host plant volatiles in the sensory ecology of R. pedestris and help explain colonization pattern of the bean bugs in soybean fields.

Electroantennogram responses of Batocera horsfieldi (Hope) to the selected volatile components of host plants, Rosa cymosa Tratt. and Rosa multiflora Thunb.

To screen for host volatile components for effectively monitoring and trapping Batocera horsfieldi Hope (Coleoptera: Cerambycidae), Headspace Solid Phase Microextraction (HS-SPME) and Gas Chromatograph-Mass Spectrometry (GC-MS) were combined to analyze volatiles emitted from Rosa cymosa Tratt. and Rosa multiflora Thunb. The results showed that alcohols (55.519%) and aldehydes (81.387%) were the primary chemicals emitted by R. cymosa and R. multiflora , respectively. An electroantennogram response (EAG) test was used to compare and analyze the responses of male and female beetles to each of the selected fifteen components, and the results showed that only nine of these components triggered significant EAG responses. Sex and volatile concentration-specific responses were observed in B. horsfieldi . Male adults showed the strongest EAG response to 1-octen-3-ol, while female adults expressed the most powerful EAG responses to salicylaldehyde and linalool. These results indicated that the sensitivity of B. horsfieldi to host volatile components was different, and the EAG response was positively correlated with the concentration of the volatile components in a certain range. Male and female adults showed different preferences for the volatile components. Furthermore, B. horsfieldi showed different preferences for volatile components and exhibited diverse recognition reactions to specific functional groups and homologous isomers. This work provided favorable data support for the development of insect attractant, and laid a theoretical foundation for the local forestry ecological protection. • R. cymosa and R. multiflora mainly contain aldehydes and alcohols. • EAG response had positive correlation with concentration. • B. horsfieldi could identify different functional groups and homologous isomers. • Male and female showed different preferences for volatile components.

The Inhibitory Effect of Powdery Mildew-Induced Volatiles from Rose on Host Selection Behavior of Beet Armyworm Moths (Lepidoptera: Noctuidae)

Abstract Plant-mediated indirect interactions (PMIIs) between phytopathogenic fungi and herbivorous insects on shared host plants occur in nature. Knowledge of PMIIs is critical in plant molecular breeding and integrated pest management. We studied the response and chemical mechanism of beet armyworm, Spodoptera exigua (Hübner), adults to rose plants, Rosa chinensis Jacquin, infected with rose powdery mildew, Podosphaera pannosa (Wallr.: Fr.) de Bary. Using gas chromatography-electroantennographic detection (GC-EAD) coupled with electroantennogram (EAG), we found that beet armyworm antennae responded to 8, 11, and 3 volatile organic compounds (VOCs) from noninfected roses, mildew-infected roses, and mildew alone, respectively. The EAG analyses showed 11 chemicals (e.g., limonene [1], 2-ethyl-1-hexanol [2], linalool [3], nonanal [4], (E)-β-caryophyllene [5], 1-dodecanol [7], nhexadecane [9], 1-hexadecanol [11], methyl palmitate [12], 1-octadecanol [14], and n-butyl hexadecanoate [15]) elicited electrophysiological responses of beet armyworm antennae with significant dose-response relationships (P < 0.05). The EAG responses to the three chemicals (3, 11, and 15) were greater than that to the reference chemical [i.e., (E)-2-hexenal] at 0.5, 5.0, and 50.0 mg/ml. Olfactory and ovipositional behavior assays indicated that three chemicals (2, 3, and 5) significantly attracted beet armyworm females and four chemicals (7, 11, 14, and 15) strongly repelled females. Chemicals 2, 3, and 5 from healthy roses appear to be responsible for the attraction of beet armyworm moths to healthy roses, whereas chemicals 7, 11, 14, and 15 from mildew-infected roses play key roles in inhibiting attraction of moths. VOCs from mildew alone did not attract or repel beet armyworm moths.

Optimization and field evaluation of sex-pheromone of potato tuber moth, Phthorimaea operculella Zeller (Lepidoptera: Gelechiidae).

The potato tuber moth, Phthorimaea operculella is a key pest of potato and tobacco in China. We compared the sex pheromone titers released by P. operculella female, and the electroantennogram (EAG) responses of male antennae to pheromone compounds under laboratory conditions. Then, the optimal sex pheromone ratio was screened in the field. The P. operculella sex pheromone feeding on potato or tobacco was extracted by solvent-extraction method. Main sex pheromone compounds including E4, Z7-13: AC (PTM1) and E4, Z7, Z10-13: AC (PTM2) were found in gland. The titer and relative ratio of P. operculella sex pheromone compounds secreted by female adult moths differ between host populations, while the relative EAG responses trend of P. operculella males to the pheromone components were the same when directly stimulated. In field trials, PTM1:PTM2=8:1-4:1 and PTM1:PTM2=1:4-1:7 was the most attractive ratio to P. operculella males in tobacco and potato fields, respectively. During 2019-2020, after 8 weeks of mass trapping for each year, the sex pheromone could reduce the population of P. operculella in the potato and tobacco fields. In this study, fixed property and quantity analyzing methods were adopted to compare sex pheromones from feeding on potato versus tobacco. EAG tests then were carried out on P. operculella males. Finally, we optimized the ratio of two sex pheromone compounds in potato and examined how this influenced field trapping. By further improving the parameters of sex pheromone application in the field, we demonstrate that deployment of sex pheromones can provide effective control of P. operculella. © 2021 Society of Chemical Industry.

Identification of Sex Pheromone Components of Korean Dioryctria abietella (Lepidoptera: Pyralidae) Population and Synergism of Pheromone and Pine Cone Volatile Blends.

(9Z,11E)-tetradecadienyl acetate (9Z,11E-14:OAc) and (3Z,6Z,9Z,12Z,15Z)-pentacosapentaene (C25 pentaene), known as the sex pheromone of the European Dioryctria abietella population, were identified in the Korean D. abietella population. The ratio of 9Z,11E-14:OAc to C25 pentaene extracted from female pheromone glands was 1:23. The electroantennogram (EAG) response of male antennae to 9Z,11E-14:OAc was stronger than to C25 pentaene, whereas an opposite EAG response was observed in female antennae. Major volatile compounds in mature Korean pine cone (Pinus koraiensis), such as limonene, myrcene, and (-)-α-pinene, elicited dose-dependent EAG responses of male and female antennae. In field attraction testing, traps baited with 100:1,000, 100:2,000, and 100:3,000 ug of 9Z,11E-14:OAc to C25 pentaene were the most attractive to male D. abietella. More males were attracted to traps baited with 9Z,11E-14:OAc+C25 pentaene+limonene than traps baited with 9Z,11E-14:OAc+C25 pentaene. This showed that limonene acted as a synergist to sex pheromone. Delta traps, wing traps, and diamond traps were superior to bucket traps for capturing D. abietella males. This study showed that pheromone traps baited with 100:1,000 ~ 100:3,000 ug of 9Z,11E-14:OAc to C25 pentaene and limonene will be useful for the exact monitoring of D. abietella flight phenology in Pinus koraiensis Sieb. & Zucc. (Pinales: Pinaceae) forests.

Olfactory dysfunction and potential mechanisms caused by volatile organophosphate dichlorvos in the silkworm as a model animal

Volatile pesticides impair olfactory function in workers/farmers and insects, but data on molecular responses and mechanisms are poorly understood. This study aims to reveal the mechanisms of olfactory dysfunction in the silkworm after exposure to volatile dichlorvos. Our results demonstrated that acute exposure for 12 h significantly reduced electroantennogram responses, and over 62.50% of the treated male moths cannot locate the pheromone source. Transcriptional and proteomic responses of the antennae and heads were investigated. A total of 101 differentially expressed genes (DEGs) in the antennae, 138 DEGs in the heads, and 43 differentially expressed proteins (DEPs) in the heads including antennae were revealed. We discovered that upregulations of Arrestin1 and nitric oxide synthase1 (NOS1) may inhibit cyclic nucleotide-gated channels and hinder calcium influx in the antennae. In the central nervous systems (CNS), downregulations of tyrosine hydroxylase (TH) and tyrosine decarboxylase (TDC) may inhibit olfactory signal transduction by reducing the second messenger biosynthesis. Meanwhile, an abnormal increase of brain cell apoptosis was revealed by Annexin V-mCherry staining, often leading to persistent neurologic impairment. Taken together, this study highlighted olfactory dysfunction caused by dichlorvos, which may provide a novel perspective for understanding the toxicity mechanism of volatile pesticides in other organisms.

Identification of Components of the Aggregation Pheromone of the Guam Strain of Coconut Rhinoceros Beetle, Oryctes rhinoceros, and Determination of Stereochemistry

The coconut rhinoceros beetle, Oryctes rhinoceros (Linnaeus 1758) (Coleoptera: Scarabaeidae: Dynastinae) (CRB), is endemic to tropical Asia where it damages both coconut and oil palm. A new invasion by CRB occurred on Guam in 2007 and eradication attempts failed using commonly applied Oryctes rhinoceros nudivirus (OrNV) isolates. This and subsequent invasive outbreaks were found to have been caused by a previously unrecognized haplotype, CRB-G, which appeared to be tolerant to OrNV. The male-produced aggregation pheromone of the endemic, susceptible strain of O. rhinoceros (CRB-S) was previously identified as ethyl 4-methyloctanoate. Following reports from growers that commercial lures containing this compound were not attractive to CRB-G, the aim of this work was to identify the pheromone of CRB-G. Initial collections of volatiles from virgin male and female CRB-G adults from the Solomon Islands failed to show any male- or female-specific compounds as candidate pheromone components. Only after five months were significant quantities of ethyl 4-methyloctanoate and 4-methyloctanoic acid produced by males but not by females. No other male-specific compounds could be detected, in particular methyl 4-methyloctanoate, 4-methyl-1-octanol, or 4-methyl-1-octyl acetate, compounds identified in volatiles from some other species of Oryctes. Ethyl 4-methyloctanoate elicited a strong electroantennogram response from both male and female CRB-G, but these other compounds, including 4-methyloctanoic acid, did not. The enantiomers of ethyl 4-methyloctanoate and 4-methyloctanoic acid were conveniently prepared by enzymatic resolution of the commercially-available acid, and the enantiomers of the acid, but not the ester, could be separated by gas chromatography on an enantioselective cyclodextrin phase. Using this approach, both ethyl 4-methyloctanoate and 4-methyloctanoic acid produced by male CRB-G were shown to be exclusively the (R)-enantiomers whereas previous reports had suggested male O. rhinoceros produced the (S)-enantiomers. However, re-examination of the ester and acid produced by male CRB-S from Papua New Guinea showed that these were also the (R)-enantiomers. In field trapping experiments carried out in the Solomon Islands, both racemic and ethyl (R)-4-methyloctanoate were highly attractive to both male and female CRB-G beetles. The (S)-enantiomer and the corresponding acids were only weakly attractive. The addition of racemic 4-methyloctanoic acid to ethyl 4-methyloctanoate did significantly increase attractiveness, but the addition of (R)- or (S)-4-methyloctanoic acid to the corresponding ethyl esters did not. Possible reasons for the difference in assignment of configuration of the components of the CRB pheromone are discussed along with the practical implications of these results.

Volatiles Induced from Hypolepis punctata (Dennstaedtiaceae) by Herbivores Attract Sclomina erinacea (Hemiptera: Reduviidae): Clear Evidence of Indirect Defense in Fern.

Simple SummaryPlants have developed many kinds of indirect defense mechanisms against herbivorous insects, through which the natural enemies are attracted by herbivore-induced plant volatiles to prey on these phytophagous species. There have been many reports regarding this mechanism in spermatophytes, but little is known in ferns. In this study, the relationship between the fern species Hypolepis punctata and the predatory insect Sclomina erinacea was studied. Employing field observations, plant volatile analysis, electrophysiological experiments, and behavioral experiments, we found that not only the plant can attract the assassin bug in the field, but compared with the healthy individuals, the fern being damaged by herbivorous insects also emitted several novel volatile organic compounds, which were strongly attractive to the assassin bug. The results indicate that ferns may also have indirect defense mechanisms using volatile organic compounds, and it is the first report of an indirect defense mechanism in fern.Plants have evolved various self-defense mechanisms against insect feeding. There are many reports regarding both direct and indirect defense mechanisms in seed-plant. However, only direct defenses on ferns were considered and the indirect defense mechanism has never been reported. In this study, it was observed that the fern Hypolepis punctata can attract the assassin bug Sclomina erinacea in the field. We collected and analyzed volatiles from H. punctata healthy individuals and the ones wounded by Bertula hadenalis, using dynamic headspace and GC-MS. We recorded the electroantennogram responses of antennae of S. erinacea to different standards of volatile compounds identified from the GC-MS analysis. We also analyzed the behavior of male and female S. erinacea adults in response to volatiles collected from H. punctata using a Y-tube olfactometer. The results showed that a number of volatile compounds were produced when the fern was damaged by B. hadenalis. Electroantennography and Y-tube olfactometer results showed that some herbivore-induced volatiles and volatiles from undamaged leaves could attract S. erinacea. Our research suggests that H. punctata can attract insect predators by releasing herbivory-induced volatile organic compounds, and for the first time we found ferns may also have indirect defense mechanisms using volatile organic compounds.

Detection of Volatile Organic Compounds by Antennal Lamellae of a Scarab Beetle

Chemoreceptive sensilla are abundantly distributed on antennal lamellae of scarab beetles. Olfactory reception by these sensory lamellae plays a major role in feeding behaviors and sexual communication of these beetles. A new electroantennogram (EAG) recording technique is here described for evaluation of electrophysiological responses of antennal lamellae of Pseudosymmachia flavescens to sex pheromones and host plant-related compounds. EAG responses were recorded simultaneously from each lamella and the closed antennal club. All test stimuli elicited similar EAG depolarization profiles in all the three lamellae and the closed club although EAG amplitudes from the same lamella or the club varied widely among different chemical stimuli. The mid lamella tended to produce significantly greater EAG responses. EAG responses evoked by a sex pheromone component, anisole, showed a significant correlation with the density of sensilla placodea subtype 1 (SP1). However, no general patterns were obtained for correlations between the density of any test sensilla type and EAG amplitudes evoked by all the six plant volatiles. Single sensillum recordings are needed to elucidate the specific roles of these sensilla in intraspecific sexual communication and perception of host plant volatiles.

CRISPR/Cas9 mutagenesis abolishes odorant-binding protein BdorOBP56f-2 and impairs the perception of methyl eugenol in Bactrocera dorsalis (Hendel)

Olfaction underpins many insect behaviors, such as foraging, host location, mating, and predator avoidance. In the first step of insect olfaction, odorant-binding proteins (OBPs) bind hydrophobic odorants and transport them to odorant receptors. Methyl eugenol (ME) is a powerful attractant for mature males of the oriental fruit fly Bactrocera dorsalis (Hendel), one of the most destructive fruit pests. The underlying molecular mechanism is unclear, but there is in vitro evidence that BdorOBP56f-2 is involved in ME perception. We used microscale thermophoresis to confirm that BdorOBP56f-2 directly binds ME with strong affinity in vitro. We then used CRISPR/Cas9 to knock out the BdorOBP56f-2 gene, allowing us to establish a homozygous mutant B. dorsalis line. The electroantennogram response and behavioral attraction to ME were significantly reduced in the mutant, providing in vivo evidence that BdorOBP56f-2 is necessary for efficient ME perception. Our results offer insight into the molecular mechanism of ME perception in B. dorsalis and provide a theoretical basis for the functional analysis of other OBPs.

Development of a semiochemical-based trapping technique for efficient control ofOdoiporus longicollisOlivier (Coleoptera: Curculionidae)

AbstractThe banana stem weevil,Odoiporus longicollisOlivier (Coleoptera: Curculionidae), is an important economic insect pest of bananas throughout the world. In this study, several important volatile substances were analysed to develop a semiochemical-based trapping technique for the control ofO. longicollis. Electroantennogram responses of both sexes were associated with electroantennogram-active compounds (1-hexanol, 1-heptanol,cis-3-Nonen-1-ol, methyl jasmonate, and terpenoids such as (–)-linalool, (+)-(1R)-α-pinene, (–)-(1S)-α-pinene, and 1,8-cineole) and then were analysed using a Y-tube olfactometer. The results revealed that only methyl jasmonate and 1-hexanol elicited strong behavioural responses inO. longicollis. These two semiochemicals and the host plant extract were evaluated individually and in a mixture for their efficiencies in attractingO. longicollisin order to develop a semiochemical-based trapping technique. The methyl jasmonate + host plant extract and 1-hexanol + host plant extract mixtures captured more weevils (> 3.5 times) than the host plant extract alone did in both high- and low-altitude regions. The captured weevils were female-dominated, with a mean sex ratio of 1:1.29 (males:females). This is the first report on the use of methyl jasmonate or 1-hexanol and the host plant extract as a potential attractant in mass trapping and managingO. longicollis.

Sodium channel activation underlies transfluthrin repellency in Aedes aegypti

BackgroundVolatile pyrethroid insecticides, such as transfluthrin, have received increasing attention for their potent repellent activities in recent years for controlling human disease vectors. It has been long understood that pyrethroids kill insects by promoting activation and inhibiting inactivation of voltage-gated sodium channels. However, the mechanism of pyrethroid repellency remains poorly understood and controversial.Methodology/Principal findingsHere, we show that transfluthrin repels Aedes aegypti in a hand-in-cage assay at nonlethal concentrations as low as 1 ppm. Contrary to a previous report, transfluthrin does not elicit any electroantennogram (EAG) responses, indicating that it does not activate olfactory receptor neurons (ORNs). The 1S-cis isomer of transfluthrin, which does not activate sodium channels, does not elicit repellency. Mutations in the sodium channel gene that reduce the potency of transfluthrin on sodium channels decrease transfluthrin repellency but do not affect repellency by DEET. Furthermore, transfluthrin enhances DEET repellency.Conclusions/SignificanceThese results provide a surprising example that sodium channel activation alone is sufficient to potently repel mosquitoes. Our findings of sodium channel activation as the principal mechanism of transfluthrin repellency and potentiation of DEET repellency have broad implications in future development of a new generation of dual-target repellent formulations to more effectively repel a variety of human disease vectors.

Functional Characterization of a Candidate Sex Pheromone Receptor AlinOR33 Involved in the Chemoreception of Adelphocoris lineolatus.

Sex pheromones are deemed to play a significant role in sexual communication of most insects. Although many sex pheromone components in mirid bugs have been identified, the roles of odorant receptors in sex pheromone perception in Adelphocoris spp. (Hemiptera: Miridae) remain unknown so far. Here, AlinOR33, a candidate sex pheromone receptor in Adelphocoris lineolatus was functionally characterized. Phylogenetic analysis showed that AlinOR33 clustered with the sex pheromone receptor AlucOR4 fromApolygus lucorum. Quantitative real-time PCR measurement revealed that the expression of AlinOR33 increased gradually from nymph to adult stage and reached its peak in the antennae of 3-day-old mated male bugs. The subsequent in situ hybridization demonstrated that AlinOR33 was mainly expressed in sensilla trichoid on the antennae of A. lineolatus. In the two-electrode voltage clamp recordings, AlinOR33/AlinOrco was specifically tuned to four sex pheromone components including butyl butyrate, hexyl hexanoate, trans-2-hexenyl butyrate and hexyl butyrate, and especially most sensitive to the major component trans-2-hexenyl butyrate. After dsAlinOR33 injection, the electroantennogram responses of males to four sex pheromone components were reduced significantly (∼50%). Compared to control bugs, dsAlinOR33-injected male bugs almost lost behavioral preference for trans-2-hexenyl butyrate. Furthermore, the wingbeat frequency of dsAlinOR33-injected male bugs notably declined. Therefore, we conclude that as a candidate sex pheromone receptor, AlinOR33 plays essential roles in the sexual behavior of A. lineolatus.

Functional Characterization of Olfactory Proteins Involved in Chemoreception of Galeruca daurica.

Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) play a fundamental role in insect olfaction. Galeruca daurica (Joannis) is a new pest with outbreak status in the Inner Mongolia grasslands, northern China. In this study, six olfactory protein genes (GdauOBP1, GdauOBP6, GdauOBP10, GdauOBP15, GdauCSP4, and GdauCSP5) were cloned by RACE and expressed by constructing a prokaryotic expression system. Their binding affinities to 13 compounds from host volatiles (Allium mongolicum) were determined by fluorescence-binding assay. In order to further explore the olfactory functions of GdauOBP15 and GdauCSP5, RNA interference (RNAi) and electroantennogram (EAG) experiments were conducted. Ligand-binding assays showed that the binding properties of the six recombinant proteins to the tested volatiles were different. GdauOBP6, GdauOBP15, GdauCSP4, and GdauCSP5 could bind several tested ligands of host plants. It was suspected that GdauOBP6, GdauOBP15, GdauCSP4, and GdauCSP5 were related to the host location in G. daurica. We also found that there were different EAG responses between males and females when the GdauOBP15 and GdauCSP5 genes were silenced by RNAi. The EAG response of G. daurica females to 2-hexenal was significantly decreased in dsRNA-OBP15-injected treatment compared to the control, and the dsRNA-CSP5-treated females significantly reduced EAG response to eight tested host volatiles (1,3-dithiane, 2-hexenal, methyl benzoate, dimethyl trisulfide, myrcene, hexanal, 1,3,5-cycloheptatriene, and p-xylene). However, the EAG response had no significant difference in males. Both GdauOBP15 and GdauCSP5 may have different functions between males and females in G. daurica and may play more important roles in females searching for host plants.

Pyrethrins elicit olfactory response and spatial repellency in Aedes albopictus.

Pyrethrum from dry flowers of Chrysanthemum is a well-known botanical insecticide and repellent. Its insecticidal activity attributes to its six insecticidal esters, collectively known as pyrethrins. Pyrethrins and its synthetic analogs pyrethroids exert their toxic action by modifying the function of voltage-gated sodium channels. Aside from insecticidal activity, pyrethrum has also been used to repel mosquitoes for centuries. Today, pyrethrum continues to be used as an active ingredient in mosquito coils and other mosquito-repellent devices globally. However, the mechanism of pyrethrum repellency remains largely unknown. Here we report that pyrethrum vapor induced spatial (non-contact) repellency in Aedes albopictus, a major vector of dengue and West Nile viruses. Using electroantennogram (EAG) recordings from adult antennae, we found that pyrethrum elicited EAG response in a dose-dependent manner. We then isolated the six insecticidal esters, pyrethrins I and II, cinerins I and II, jasmolins I and II from pyrethrum extract and discovered that five of the six esters, except jasmolin I, all elicited EAG responses. Furthermore, pyrethrins I and II, cinerin II and jasmolin II induced repellency, whereas cinerin I and jasmolin I did not. Of the six pyrethrins, four of them, pyrethrins I and II, cinerin II and jasmolin II, activate olfactory-receptor neurons and elicit spatial repellency in Ae. albopictus. Our study provided a foundation for future structure-function studies of pyrethrins, their cognate olfactory receptors and efficacies of repellency and for the development of new and more effective mosquito repellents for controlling vector-borne human diseases. © 2021 Society of Chemical Industry.