Computational modeling and experimental validation of odor detection behaviors of classically conditioned parasitic wasp,Microplitis croceipes

A prototype chemical sensor named Wasp hound® that utilizes five classically conditioned parasitoid wasps, Microplitis croceipes (Cresson) (Hymenoptera: Braconidae), to detect volatile odors was successfully implemented in a previous study. To improve the odor‐detecting ability of Wasp Hound®, searching behaviors of an individual wasp in a confined area are studied and modeled through stochastic differential equations in this paper. The wasps are conditioned to 20 mg of coffee when associated with food and subsequently, tested to 5, 10, 20, and 40 mg of coffee. A stochastic model is developed and validated based on three positive behavioral responses (walking, rotation around odor source, and self‐rotation) from conditioned wasps at four different test dosages. The model is capable to reproducing the behaviors of conditioned wasps, and can be used to improve the ability of Wasp Hound® to assess changes in odor concentration. The model simulation results show the behaviors of conditioned wasps are significantly different when tested at different coffee dosages. We conjecture that the searching behaviors of conditioned wasps are based on the temporal and spatial neuron activity of olfactory receptor neurons and glomeruli, which are strongly correlated to the training dosages. The overall results demonstrate the utility of mathematical models for interpreting experimental observations, gaining novel insights into the dynamic behavior of classically conditioned wasps, as well as broadening the practical uses of Wasp Hound. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:596–606, 2015     

Influence of handling and conditioning protocol on learning and memory of Microplitis croceipes

Microplitis croceipes (Cresson) (Hymenoptera: Braconidae) learns odors in association with both hosts and food. The food‐associated ‘seeking’ behavior of M. croceipes was investigated under various training protocols utilizing the conditioning odor, 3‐octanone. We investigated the effects of odor training, or its lack, training duration, training frequency, time elapsed after training, wasp hunger state, and training reinforcement, on the food‐seeking responses of M. croceipes females. We found that odor‐trained females show strong food seeking responses, whereas non‐odor‐trained females do not respond to the odor, and that a single 10 s association with the odor whilst feeding on sugar water subsequently conditioned the wasps to exhibiting significant responses to it. Increases in training time to more than 10 s did not improve their responses. Repetition of the food–odor associations increased a wasp's recall, as well as its response over time, compared to a single exposure. Repeated exposure to the learned odor in the absence of a food reward decreased the responses of less hungry individuals. However, the level of response increased significantly following a single reinforcement with the food–odor association. Understanding the factors that influence learning in parasitoids can enhance our ability to predict their foraging behavior, and opens up avenues for the development of effective biological detectors.     

Learning and the role of host-specific volatiles during in-flight host-finding in the specialist parasitoid Microplitis croceipes

Abstract The effect of antennal contact with host-related stimuli on the subsequent flight response to host-specific volatiles was examined in the specialist parasitoid, Microplitis croceipes (Cresson) (Hymenoptera: Braconidae). In a choice test between two species of caterpillars feeding on cowpea, free-flying female M.croceipes lacking prior contact with host cues landed more often on sources of odour from its host, Heliothis tea (Boddie), than from non-host Spodoptera frugiperda (Smith). After female parasitoids contacted host cues with their antennae, more landings on sources containing hosts were observed only when caterpillars used in the pre-flight treatment and in the odour sources fed on the same plant species. The degree to which host-experienced wasps landed more often on hosts as compared with non-hosts varied with host-instar and plant species. The role of learning in host finding in specialist parasitoids is discussed.     

棉铃虫雌蛾对小麦花挥发性气味的触角电位反应

测定了棉铃虫Helicoverpa armigera处女雌蛾和交配雌蛾对小麦花中单一或多个挥发性物质的EAG反应。在单组分或多组分的测定中,棉铃虫处女蛾和交配蛾在P=0.05水平上并不存在显著性差异,但在混合的绿叶气味物质中加入含量为1×10^-5～5×10^-5（体积比）的正庚醛时,交配蛾对混合物的触角电位反应显著高于处女蛾的反应（P<0.05）。     

A learning-related variation in electroantennogram responses of a parasitic wasp

ABSTRACT Odour responses of parasitic insects, in search of their hosts, can change due to experience. Leptopilina heterotoma (Thomson) (Hymenoptera; Eucoilidae), a parasitic wasp of drosophilid larvae, is known to alter its preference for odours emanating from host food substrates through learning. These kinds of behavioural modifications in insects are assumed to be the result of complex processes in the brain. The results presented in this report, however, suggest that this learning-related behavioural variation is not restricted to brain processes but that it involves changes in sensitivity of olfactory receptor neurones.     

两种寄生蜂的卵巢结构与卵成熟特点初步观察

对斜纹夜蛾的内寄生蜂双斑侧沟茧蜂MicroplitisbicoloratusChen和南美斑潜蝇的外寄生蜂潜蝇姬小蜂Diglyhusisaea(Walker)的卵巢结构进行了研究。双斑侧沟茧蜂的2个卵巢管底部膨大有大量的卵,通过凹陷的颈部与萼区相连接,在萼区里大量成熟待产的卵浸泡在萼液中。产卵量随产卵时间延长而降低,在日产卵实验中,日产卵量202(±114SE),从1000至1800每2h的产卵量分别占日产卵量的38,32,17和13%;在一生产卵量实验中,9d产卵期,每头雌峰一生平均产卵量为502(±78SE)粒,第1d产卵量占一生总产卵量的42%。潜蝇姬小蜂的每个卵巢由3条卵巢管组成,每条输卵管中仅有1粒成熟卵,整个卵巢中只有6粒成熟卵,每次产卵数不超过6粒。结果表明,M.bicoloratus是早熟卵(proovigenic)类昆虫;D.isaea是同步卵(synovigenic)类昆虫。     

Comparison of detection ability of learned odors between males and females in the larval parasitoid Microplitis croceipes

Although female parasitic wasps are known to learn to associate odors with hosts and food, the ability of males to learn and detect odors has been neglected. We conducted laboratory experiments to compare the detection ability of learned odors between males and females in the larval parasitoid Microplitis croceipes (Cresson) (Hymenoptera: Braconidae). We first conditioned males and females to associate sucrose water with methyl benzoate, 3‐octanone, or cyclohexanone, and then observed their behavior toward various concentrations (40 ng l^?1?4 mg l^?1) of the trained odors. Conditioned male wasps responded as well as female wasps to various concentrations of the three odors. Response times by wasps to these three odors were not significantly different between sexes. For the three odors, response times of both sexes were longer at the intermediate concentrations (40–400 µg l^?1) than the higher or lower concentrations. The present study suggests that M. croceipes males can learn and respond to the three chemicals tested as well as the females, and conditioned males are as sensitive to learned odors as conditioned females. By using their sensitive learning and odor‐detection capabilities, M. croceipes males could search for food sources as efficiently as females under natural conditions.     

中红侧沟茧蜂对烟草挥发物的触角电生理及行为反应

中红侧沟茧峰 Microplitis mediator是棉铃虫 H elicoverpa armigera的主要内寄生蜂 ,是优良的蜂种 ,在生物防治上具有广阔的应用前景。但迄今 ,有关中红侧沟茧蜂的化学生态的研究甚少。利用触角电位技术 ,测定了中红侧沟茧蜂对烟草叶片粗提物和烟草中 10种挥发物的触角电生理反应 ,并测定了中红侧沟茧蜂的触角电位反应随日龄的变化。以嗅觉仪测定了中红侧沟茧蜂对烟草粗提物、烟草中 4种挥发物的行为反应。结果表明中红侧沟茧蜂利用烟草释放的气味向烟草定向 ,进而找到其寄主——棉铃虫 ,也佐证了烟草同样是棉铃虫的重要寄主植物的论点。雌蜂对脂肪族化合物反应最强 ,对芳香族化合物和糠醛 (杂环化合物 )的反应居中 ,对萜类化合物反应较弱或者不反应。表明烟草挥发性物质在中红侧沟茧蜂寻找寄主的过程中起着重要的作用 ,但挥发物各组分的作用存在着差异。雌性中红侧沟茧蜂对顺 - 3-己烯醇的触角电位反应随日龄的增加而变化。 1日龄的雌蜂的反应较弱 ,2日龄、3日龄的雌蜂反应较强 ,4日龄、5日龄的雌蜂的反应又减弱。这与该蜂产卵习性相符 :羽化当天的雌蜂产卵较少 ,羽化 2～ 3d之后的雌蜂产卵量较大     

Electroantennogram responses of the Mediterranean fruit fly, Ceratitis capitata, to the volatile constituents of nectarines

Electroantennograms (EAGs) were recorded from laboratory-reared, male and female Ceratitis capitata (medfly) in response to individual volatiles identified from ripe nectarines. Uniformity in EAG response between the sexes was observed for most test compounds. Only 10 volatiles, of the 44 nectarine volatiles tested, elicited significantly different EAG responses between the sexes. No correlation was observed between the magnitude of antennal responsiveness and the concentration of a particular volatile in the headspace odor of ripe nectarines. The most abundant (major) nectarine volatiles were among the least EAG stimulatory compounds tested. And certain minor and trace volatiles were the most potent compounds in eliciting EAGs. Moreover, the magnitude of antennal response to a nectarine volatile was related to the functional-group, chain-length, and unsaturation of the compound. The degree of potency of the compounds was as follows: six-carbon unsaturated aldehydes and alcoholsmethyl and ethyl hexanoates and octanoateshexenyl acetates and monoterpenes>shorter chain-length acetates and alcohols>lactones. Unsaturated aldehydes, alcohols, and acetates generally elicited larger EAGs than their saturated analogs, with the (E)-2-isomers being the most potent isomeric configurations. In addition, medfly antennae exhibited long recovery periods (i.e., >10 sec.) for the EAG tracings to return to baseline potential after stimulations with certain classes of compounds, e.g., C₆ to C₈ acid esters, monoterpenes, and hexen-1-ols. The potential adaptiveness to medflies for selective sensitivity to these volatiles is discussed.     

Electroantennogram and behavioral responses of Cotesia plutellae to plant volatiles

Plant volatiles have been demonstrated to play an important role in regulating the behavior of Cotesia plutellae, a major larval parasitoid of the diamondback moth (DBM), Plutella xylostella, but little is currently known about the function of each volatile and their mixtures. We selected 13 volatiles of the DBM host plant, a cruciferous vegetable, to study the electroantennogram (EAG) and behavioral responses of C. plutellae. EAG responses to each of the compounds generally increased with concentration. Strong EAG responses were to 100 μL/mL of trans‐2‐hexenal, benzaldehyde, nonanal and cis‐3‐hexenol, and 10 μL/mL of trans‐2‐hexenal and benzaldehyde with the strongest response provoked by trans‐2‐hexenal at 100 μL/mL. In the Y‐tube olfactometer, C. plutellae, was significantly attracted by 1 μL/mL of trans‐2‐hexenal and benzaldehyde. β‐caryophyllene, cis‐3‐hexenol or trans‐2‐hexenal significantly attracted C. plutellae at 10 μL/mL, while nonanal, benzyl alcohol, cis‐3‐hexenol or benzyl cyanide at 100 μL/mL significantly attracted C. plutellae. Trans‐2‐hexenal significantly repelled C. plutellae at 100 μL/mL. EAG of C. plutellae showed strong responses to all mixtures made of five various compounds with mixtures 3 (trans‐2‐hexenal, benzaldehyde, nonanal, cis‐3‐hexenol, benzyl cyanide, farnesene, eucalyptol) and 4 (trans‐2‐hexenal, benzaldehyde, benzyl alcohol, (R)‐(+)‐limonene, β‐ionone, farnesene, eucalyptol) significantly attracting C. plutellae. These findings demonstrate that the behavior of C. plutellae can be affected either by individual compounds or mixtures of plant volatiles, suggesting a potential of using plant volatiles to improve the efficiency of this parasitoid for biocontrol of P. xylostella.     

中红侧沟茧蜂对棉花挥发性物质的触角电位反应

中红侧沟茧蜂Microplitis mediator (Haliday) 是棉铃虫 Helicoverpa amigera (Hübner)的主要寄生蜂之一。在寻找害虫的过程中,害虫的寄主植物释放出的气味起着至关重要的作用。本文测定了中红侧沟茧蜂雌蜂对棉花中17种气味物质的触角电位反应,比较了雌雄茧蜂对6种气味反应的差别,并且测定了雌蜂对4种气味物质的剂量 反应。结果表明,在所测的气味物质中,中红侧沟茧蜂对脂肪族化合物的反应最强,对芳香族化合物的反应居中,而对萜类化合物的反应较弱。对所测的气味物质的反应,雌雄茧蜂之间的差异不显著。雌蜂对正己醛、苯乙酮、正壬醇的反应阈值为25 μL, 对苯甲醛的反应阈值为250 μL,在所测的浓度范围内,对4种化合物的反应均未达到饱和。     

Functional characterization of a Niemann–Pick type C2 protein in the parasitoid wasp Microplitis mediator

Niemann–Pick type C2 (NPC2) is a type of small soluble protein involved in lipid metabolism and triglyceride accumulation in vertebrates and arthropods. Recent studies have determined that NPC2 also participates in chemical communication of arthropods. In this work, two novel NPC2 proteins (MmedNPC2a and MmedNPC2b) in Microplitis mediator were identified. Real‐time quantitative PCR (qPCR) analysis revealed that MmedNPC2a was expressed higher in the antennae than in other tissues of adult wasps compared with MmedNPC2b. Subsequent immunolocalization results demonstrated that NPC2a was located in the lymph cavities of sensilla placodea. To further explore the binding characterization of recombinant MmedNPC2a to 54 candidate odor molecules, a fluorescence binding assay was performed. It was found MmedNPC2a could not bind with selected fatty acids, such as linoleic acid, palmitic acid, stearic acid and octadecenoic acid. However, seven cotton volatiles, 4‐ethylbenzaldehyde, 3,4‐dimethylbenzaldehyde, β‐ionone, linalool, m‐xylene, benzaldehyde and trans‐2‐hexen‐1‐al showed certain binding abilities with MmedNPC2a. Moreover, the predicted 3D model of MmedNPC2a was composed of seven β‐sheets and three pairs of disulfide bridges. In this model, the key binding residues for oleic acid in CjapNPC2 of Camponotus japonicus, Lue68, Lys69, Lys70, Phe97, Thr103 and Phe127, are replaced with Phe85, Ser86, His87, Leu113, Tyr119 and Ile143 in MmedNPC2a, respectively. We proposed that MmedNPC2a in M. mediator may play roles in perception of plant volatiles.     

麦长管蚜和禾谷缢管蚜对小麦挥发物的触角电位反应

采用活体蚜虫测定法,利用EAG技术比较分析了麦长管蚜Sitobion avenae和禾谷缢管蚜Rhopalosiphum padi有翅及无翅成蚜对小麦挥发物及麦蚜取食诱导挥发物组分的嗅觉反应,揭示了两种麦蚜的嗅觉变异特点。结果表明：麦长管蚜对水杨酸甲酯、反-2-己烯醛、反-2-己烯醇、-6-甲基-5-庚烯-2-酮和-6-甲基-5-庚烯-2醇的反应较强,禾谷缢管蚜对水杨酸甲酯、反-3-己烯乙酸酯、-6-甲基-5-庚烯-2-酮和-6-甲基-5-庚烯-2-醇的反应较强,并得到了剂量反应曲线。麦长管蚜的有翅和无翅成蚜对6-甲基-5-庚烯-2-酮、反-2-己烯醇和水杨酸甲酯的反应差异显著;禾谷缢管蚜的有翅和无翅成蚜对反-2-己烯醇、辛醛、里那醇、水杨酸甲酯和反-3-己烯乙酸酯的EAG反应差异显著,其原因与禾谷缢管蚜迁移及转主为害的生物学特性有关。     

两近缘种夜蛾对低浓度烟草挥发物的触角电位反应

寡食性的烟夜蛾Helicoverpa assulta (Guenée)和广食性的棉铃虫Helicoverpa armigera (Hübner)是实夜蛾属的2近缘种昆虫,在它们的寄主植物中,只有烟草为其共同寄主,为明确它们对烟草挥发物嗅觉感受能力的差异,测定了这2种夜蛾的处女雌蛾、交配雌蛾及雄蛾对20种低浓度(1.0×10-5 mol · L-1)烟草挥发物的触角电位反应,并对其中6种化合物进行了剂量反应测定,结果表明,(1) 2种夜蛾对测试化合物均可产生EAG反应且反应趋势相似:对植物中普遍存在的绿叶气味、脂肪族和芳香族化合物的反应显著高于具有烟草"种"特异性的单萜类、倍半萜类和杂环化合物,表明2种夜蛾远距离定向时,对植物"种"的辨识能力均较差.(2) 2种夜蛾对脂肪族C6_化合物的EAG反应最强,碳链延长或缩短都会导致反应强度的降低.(3) 烟夜蛾对大多数化合物的EAG反应大于棉铃虫,表明在远距离寄主定向时其嗅觉感受器对寄主挥发物的感受能力强于棉铃虫.(4) 烟夜蛾交配雌蛾对β-水芹烯和桧烯的EAG反应高于其处女雌蛾和雄蛾,除此之外2种夜蛾的EAG反应未见显著的性别差异,推测β-水芹烯和桧烯可能是烟夜蛾在烟草上产卵的"信号"物质.(5) 剂量反应曲线表明,在测试的浓度范围内,2种夜蛾对6种类型化合物的EAG反应强度与其浓度呈正相关,且均未达到饱和.由此可知,与棉铃虫相比,烟夜蛾嗅觉神经系统在其寄主定向行为中所起的作用更重要、也更专化.     

油松毛虫雌蛾对油松松针两种手性化合物的触角电位反应

采用触角电位技术测定了油松毛虫雌蛾触角对油松挥发物的两对手性单萜的剂量反应,并在饱和剂量下测定了α-蒎烯、β-蒎烯手性化合物及其消旋体的触角电位值。剂量反应测试表明,油松毛虫雌蛾对(-)-α-蒎烯的饱和剂量为1 600 μL,对(+)-α-蒎烯、(+)-β-蒎烯、(-)-β-蒎烯的饱和剂量为800 μL。α-蒎烯右旋异构体的反应值高于(-)-α-蒎烯的反应值,说明(+)-α-蒎烯能够更有效的与感受器中的分子受体结合;β-蒎烯则相反,(-)-β-蒎烯的EAG反应值高于(+)-β-蒎烯,说明β-蒎烯的右旋异构体不能有效的与多数感受器中的分子受体结合。在饱和剂量下的测定结果表明油松毛虫雌蛾对α-蒎烯和β-蒎烯消旋体的反应均与对其手性异构体反应相当,说明油松毛虫雌蛾用同一个感受细胞来接受α-蒎烯的两个对映异构体,同样β-蒎烯的两个对映异构体也是被同一个受体细胞来接受。结果提示,手性化合物的比例可能在油松毛虫寄主识别中有重要作用。     

小菜蛾对合成植物挥发物的活性反应

在室内检验了小菜蛾已交配和未交配雌、雄成虫对3种合成植物挥发物乙酸顺-3-己烯酯（Z-3-hexenyl acetate,ACTE）、异硫氰酸丙烯酯（allyl isothiocyanate,NCS）和顺-3-己烯醇（Z-3-hexen-1-ol,OH）的触角电位(EAG)反应。不同浓度（0.008 μg/μL、0.08 μg/μL、0.2 μg/μL、0.8 μg/μL、8 μg/μL、20 μg/μL和40 μg/μL）实验表明,随着刺激化合物浓度的提高,小菜蛾反应活性增强。但小菜蛾的性别及其交配状态可能影响其对3种化合物的反应强度：不同性别及交配状态的小菜蛾对ACTE的反应差异不大;未交配雌、雄虫对OH的反应强于已交配雌、雄虫;未交配雌虫对低浓度的NCS反应较强,NCS超过一定浓度时已交配雌虫的反应强于未交配雌虫,雄虫对NCS反应较小且与交配状态无关。以小菜蛾性信息素作为对照,在湖北长阳和越南河内试验了这3种植物挥发物诱芯（6 μL/诱芯）对小菜蛾的引诱作用。结果表明,在湖北,第1天时 ACTE、NCS对雄虫具有很强的引诱作用,引诱量显著大于性诱剂的引诱量,但随着诱芯放置时间延长,NCS引诱作用迅速下降,第2天时引诱作用已经很小;ACTE的引诱作用下降缓慢,第4天的诱蛾量才显著小于性信息素的诱蛾量,OH的引诱力较弱。在河内,ACTE、NCS、OH第1天对小菜蛾引诱作用很强,引诱量大于性信息素,但差异不显著;随着诱芯放置时间延长,3种挥发物对小菜蛾的引诱量均下降很快,第4天的引诱力就很弱。3种植物挥发物的混合物(体积比1∶1∶1)对小菜蛾的引诱作用与性信息素相似,且诱蛾活性持效期相对较长。无论在越南或湖北,植物挥发物或其混合物均很少引诱到雌虫。     

Responses of Microplitis croceipes to host and nonhost plants of Heliothis virescens in a wind tunnel

The relative attractiveness of velvet leaf (Abutilon theophrasti Medicus), cotton (Gossypium hirsutum L.) (host plants) and groundcherry (Physalis angulata L.) (nonhost plant), and cotton plants with or without nectaries and with or without glands to Microplitis croceipes Cresson (Hymenoptera: Braconidae) was determined in a wind tunnel. Female parasitoids flew significantly more to glandless than to glanded cotton; response to nectaried and nectariless cotton was similar. Velvet leaf and cotton were favored significantly over groundcherry; parasitoids being equally responsive to both host plant species. Addition of larval frass alone or in combination with host larvae significantly improved the attractivity of the nonhost plant (groundcherry) to the parasitoids. There was no difference in attractiveness of groundcherry terminals with or without host larvae. Parasitoid search time was significantly increased with addition of larval frass. In the presence of cotton, however, kairomone-treated groundcherry remained unattractive.

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Résumé L'influence des nectaires et des glandes du coton, et l'attractivité des feuilles veloutées du coton (plante hôte) et de Physalis angulata sur Microplitis croceipes, ont été déterminées dans des expériences avec tunnel à vent. Les résultats ont montré que les parasitoïdes femelles sont significativement plus attirés par les cotons sans glandes que par les cotons glanduleux, tandis que la présence ou l'absence de nectaires ne modifie pas l'attractivité du coton. Elles réagissent de la même façon aux feuilles veloutées. P. angulata traité avec des crottes de chenilles, présenté seul ou en combinaison avec du coton, attire plus de femelles que des pousses de P. angulata non traitées. L'addition de chenilles hôtes seules n'améliore pas significativement l'attractivité de la plante. P. angulata traité avec des kairomones est demeuré inattractif quand il était proposé en même temps que du coton.