Effects of an artificial blend of host‐infested plant volatiles on plant attractiveness to parasitic wasps

For the biological control of diamondback moth (DBM) larvae in commercial greenhouses, we have previously identified a blend of volatiles that attracted Cotesia vestalis, a parasitoid of DBM larvae. Here, we tested the effects of an artificial volatile blend on the attractiveness of komatsuna plants (Japanese mustard spinach; Brassica rapa var. perviridis) to C. vestalis under greenhouse conditions. First, we showed that female C. vestalis preferred infested komatsuna plants to uninfested plants in the greenhouse. Under the same conditions, placing the artificial attractants near both infested and uninfested plants did not affect the wasps’ preference. However, when comparing infested komatsuna plants coupled with the artificial attractants with infested plants without them, significantly more female C. vestalis were attracted to the former. The possible use of artificial C. vestalis attractants for the biological control of DBM is discussed.     

Combined use of herbivore‐induced plant volatiles and sex pheromones for mate location in braconid parasitoids

Herbivore‐induced plant volatiles (HIPVs) are important cues for female parasitic wasps to find hosts. Here, we investigated the possibility that HIPVs may also serve parasitoids as cues to locate mates. To test this, the odour preferences of four braconid wasps – the gregarious parasitoid Cotesia glomerata (L.) and the solitary parasitoids Cotesia marginiventris (Cresson), Microplitis rufiventris Kokujev and Microplitis mediator (Haliday) – were studied in olfactometers. Each species showed attraction to pheromones but in somewhat different ways. Males of the two Cotesia species were attracted to virgin females, whereas females of M. rufiventris were attracted to virgin males. Male and female M. mediator exhibited attraction to both sexes. Importantly, female and male wasps of all four species were strongly attracted by HIPVs, independent of mating status. In most cases, male wasps were also attracted to intact plants. The wasps preferred the combination of HIPVs and pheromones over plant odours alone, except M. mediator, which appears to mainly use HIPVs for mate location. We discuss the ecological contexts in which the combined use of pheromones and HIPVs by parasitoids can be expected. To our knowledge, this is the first study to show that braconid parasitoids use HIPVs and pheromones in combination to locate mates.     

Females of Cotesia vestalis,a parasitoid of diamondback moth larvae,learn to recognise cues from aphid‐infested plants to exploit honeydew

1. To maximise their reproductive success, the females of most parasitoids must not only forage for hosts but must also find suitable food sources. These may be nectar and pollen from plants, heamolymph from hosts and/or honeydew from homopterous insects such as aphids. 2. Under laboratory conditions, females of Cotesia vestalis, a larval parasitoid of the diamondback moth (Plutella xylostella) which does not feed on host blood, survived significantly longer when held with cruciferous plants infested with non‐host green peach aphids (Myzus persicae) than when held with only uninfested plants. 3. Naïve parasitoids exhibited no preference between aphid‐infested and uninfested plants in a dual‐choice test, but those that had been previously fed aphid honeydew significantly preferred aphid‐infested plants to uninfested ones. 4. These results suggest that parasitoids that do not use aphids as hosts have the potential ability to learn cues from aphid‐infested plants when foraging for food. This flexible foraging behaviour could allow them to increase their lifetime reproductive success.     

Plant volatiles induced by herbivore eggs prime defences and mediate shifts in the reproductive strategy of receiving plants

Plants can detect cues associated with the risk of future herbivory and modify defence phenotypes accordingly; however, our current understanding is limited both with respect to the range of early warning cues to which plants respond and the nature of the responses. Here we report that exposure to volatile emissions from plant tissues infested with herbivore eggs promotes stronger defence responses to subsequent herbivory in two Brassica species. Furthermore, exposure to these volatile cues elicited an apparent shift from growth to reproduction in Brassica nigra, with exposed plants exhibiting increased flower and seed production, but reduced leaf production, relative to unexposed controls. Our results thus document plant defence priming in response to a novel environmental cue, oviposition‐induced plant volatiles, while also showing that plant responses to early warning cues can include changes in both defence and life‐history traits.     

Insect attraction to synthetic herbivore‐induced plant volatile‐treated field crops

• 1 Plants produce natural enemy‐attracting semiochemicals known as herbivore‐induced plant volatiles (HIPV) in response to herbivore damage. Deployment of synthetic HIPV in crops could enhance the biological control of pests. To test this, six HIPV [methyl salicylate (MeSA), methyl anthranilate (MeA), methyl jasmonate (MeJA), benzaldehyde (Be), cis‐3‐hexenyl acetate (HA), cis‐hexen‐1‐ol (He)] in three concentrations (0.5%, 1.0% and 2.0% v/v) mixed with a vegetable oil adjuvant, Synertrol® (Organic Crop Protectants Pty Ltd, Australia), were sprayed onto winegrape, broccoli and sweet corn plants.
• 2 The relative abundance of insects within treated plots was assessed with non‐attracting, transparent sticky traps at varying time intervals up to 22 days after spraying.
• 3 In the vineyard experiment, Trichogrammatidae responded to Be and MeA (0.5%) and Be (1.0%); Encyrtidae and Bethylidae responded to MeA (1.0%); Scelionidae responded to all compounds at 1.0% and 2.0%; and predatory insects responded to MeA. In sweet corn, parasitoids as a group and Encyrtidae responded to MeA (0.5%); Braconidae responded to all compounds at 0.5% and Synertrol‐only; thrips responded to all compounds at 0.5% and 1.0%; while all parasitoids responded to all compounds at 0.5% and 1.0% and Synertrol‐only. In broccoli, parasitoids as a group and Scelionidae responded to Be, HA, He and Synertrol‐only; Trichogrammatidae responded to Be (0.5%), He (0.5% and 1.0%), MeJA (1.0%) and MeSA (0.5%); and thrips responded to all compounds at to 0.5% and 1.0%.
• 4 Significant attraction of insects occurred up to 6 days after the HIPV application, suggesting that plants may have been induced to produce endogenous volatiles that attracted insects over an extended period.
• 5 The results obtained are discussed in relation to the potential utility of synthetic HIPV to enhance the biological control of pests.

     

Infochemical use and dietary specialization in parasitoids: a meta‐analysis

Many parasitoid species use olfactory cues to locate their hosts. In tritrophic systems, parasitoids of herbivores can exploit the chemical blends emitted by plants in reaction to herbivore‐induced damage, known as herbivore‐induced plant volatiles (HIPVs). In this study, we explored the specificity and innateness of parasitoid responses to HIPVs using a meta‐analysis of data from the literature. Based on the concept of dietary specialization and infochemical use, we hypothesized that (i) specialist parasitoids (i.e., with narrow host ranges) should be attracted to specific HIPV signals, whereas generalist parasitoids (i.e., with broad host ranges) should be attracted to more generic HIPV signals and (ii) specialist parasitoids should innately respond to HIPVs, whereas generalist parasitoids should have to learn to associate HIPVs with host presence. We characterized the responses of 66 parasitoid species based on published studies of parasitoid behavior. Our meta‐analysis showed that (i) as predicted, specialist parasitoids were attracted to more specific signals than were generalist parasitoids but, (ii) contrary to expectations, response innateness depended on a parasitoid's target host life stage rather than on its degree of host specialization: parasitoids of larvae were more likely to show an innate response to HIPVs than were parasitoids of adults. This result changes our understanding of dietary specialization and highlights the need for further theoretical research that will help clarify infochemical use by parasitoids.     

Drought stress affects response of phytopathogen vectors and their parasitoids to infection‐ and damage‐induced plant volatile cues

1. The response of a phytopathogen vector to pathogen‐induced plant volatiles was investigated, as well as the response of the phytopathogen vector's parasitoid to herbivore‐induced plant volatiles released from plants with and without drought stress. 2. These experiments were performed with Asian citrus psyllid (Diaphorina citri), vector of the plant pathogen Candidatus Liberibacter asiaticus (CLas) and its parasitoid Tamarixia radiata as models. Candidatus Liberibacter asiaticus is the presumed causal pathogen of huanglongbing (HLB), also called citrus greening disease. 3. Diaphorina citri vectors were attracted to headspace volatiles of CLas‐infected citrus plants at 95% of their water‐holding capacity (WHC); such attraction to infected plants was much lower under drought stress. Attraction of the vector to infected and non‐stressed plants was correlated with greater release of methyl salicylate (MeSA) as compared with uninfected and non‐stressed control citrus plants. Drought stress decreased MeSA release from CLas‐infected plants as compared with non‐stressed and infected plants. 4. Similarly, T. radiata was attracted to headspace volatiles released from D. citri‐infested citrus plants at 95% of their WHC. However, wasps did not show preference between headspace volatiles of psyllid‐infested and uninfested plants when they were at 35% WHC, suggesting that herbivore‐induced defences did not activate to recruit this natural enemy under drought stress. 5. Our results demonstrate that herbivore‐ and pathogen‐induced responses are environmentally dependent and do not occur systematically following damage. Drought stress affected both pathogen‐ and herbivore‐induced plant volatile release, resulting in concomitant decreases in behavioural response of both the pathogen's vector and the vector's primary parasitoid.     

Integration of two herbivore‐induced plant volatiles results in synergistic effects on plant defence and resistance

Plants can use induced volatiles to detect herbivore‐ and pathogen‐attacked neighbors and prime their defenses. Several individual volatile priming cues have been identified, but whether plants are able to integrate multiple cues from stress‐related volatile blends remains poorly understood. Here, we investigated how maize plants respond to two herbivore‐induced volatile priming cues with complementary information content, the green leaf volatile (Z)‐3‐hexenyl acetate (HAC) and the aromatic volatile indole. In the absence of herbivory, HAC directly induced defence gene expression, whereas indole had no effect. Upon induction by simulated herbivory, both volatiles increased jasmonate signalling, defence gene expression, and defensive secondary metabolite production and increased plant resistance. Plant resistance to caterpillars was more strongly induced in dual volatile‐exposed plants than plants exposed to single volatiles.. Induced defence levels in dual volatile‐exposed plants were significantly higher than predicted from the added effects of the individual volatiles, with the exception of induced plant volatile production, which showed no increase upon dual‐exposure relative to single exposure. Thus, plants can integrate different volatile cues into strong and specific responses that promote herbivore defence induction and resistance. Integrating multiple volatiles may be beneficial, as volatile blends are more reliable indicators of future stress than single cues.     

Prospects of herbivore egg‐killing plant defenses for sustainable crop protection

Due to a growing demand of food production worldwide, new strategies are suggested to allow for sustainable production of food with minimal effects on natural resources. A promising alternative to the application of chemical pesticides is the implementation of crops resistant to insect pests. Plants produce compounds that are harmful to a wide range of attackers, including insect pests; thus, exploitation of their natural defense system can be the key for the development of pest‐resistant crops. Interestingly, some plants possess a unique first line of defense that eliminates the enemy before it becomes destructive: egg‐killing. Insect eggs can trigger (1) direct defenses, mostly including plant cell tissue growth or cell death that lead to eggs desiccating, being crushed or falling off the plant or (2) indirect defenses, plant chemical cues recruiting natural enemies that kill the egg or hatching larvae (parasitoids). The consequences of plant responses to eggs are that insect larvae do not hatch or that they are impeded in development, and damage to the plant is reduced. Here, we provide an overview on the ubiquity and evolutionary history of egg‐killing traits within the plant kingdom including crops. Up to now, little is known on the mechanisms and on the genetic basis of egg‐killing traits. Making use of egg‐killing defense traits in crops is a promising new way to sustainably reduce losses of crop yield. We provide suggestions for new breeding strategies to grow egg‐killing crops and improve biological control.     

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.     

The maize lipoxygenase,ZmLOX10, mediates green leaf volatile,jasmonate and herbivore‐induced plant volatile production for defense against insect attack

Fatty acid derivatives are of central importance for plant immunity against insect herbivores; however, major regulatory genes and the signals that modulate these defense metabolites are vastly understudied, especially in important agro‐economic monocot species. Here we show that products and signals derived from a single Zea mays (maize) lipoxygenase (LOX), ZmLOX10, are critical for both direct and indirect defenses to herbivory. We provide genetic evidence that two 13‐LOXs, ZmLOX10 and ZmLOX8, specialize in providing substrate for the green leaf volatile (GLV) and jasmonate (JA) biosynthesis pathways, respectively. Supporting the specialization of these LOX isoforms, LOX8 and LOX10 are localized to two distinct cellular compartments, indicating that the JA and GLV biosynthesis pathways are physically separated in maize. Reduced expression of JA biosynthesis genes and diminished levels of JA in lox10 mutants indicate that LOX10‐derived signaling is required for LOX8‐mediated JA. The possible role of GLVs in JA signaling is supported by their ability to partially restore wound‐induced JA levels in lox10 mutants. The impaired ability of lox10 mutants to produce GLVs and JA led to dramatic reductions in herbivore‐induced plant volatiles (HIPVs) and attractiveness to parasitoid wasps. Because LOX10 is under circadian rhythm regulation, this study provides a mechanistic link to the diurnal regulation of GLVs and HIPVs. GLV‐, JA‐ and HIPV‐deficient lox10 mutants display compromised resistance to insect feeding, both under laboratory and field conditions, which is strong evidence that LOX10‐dependent metabolites confer immunity against insect attack. Hence, this comprehensive gene to agro‐ecosystem study reveals the broad implications of a single LOX isoform in herbivore defense.     

Variability and stability of tea weevil‐induced volatile emissions from tea plants with different weevil densities,photoperiod and infestation duration

Abstract After herbivore attack, many plants emit herbivore‐induced plant volatiles (HIPVs). HIPVs can attract carnivores and/or repel herbivores, thereby mediating tritrophic plant–herbivore–carnivore interactions. HIPVs act as chemical information between organisms; hence, their variability and stability are vital. In the present study, variations in the volatile emissions, from the tea plant Camellia sinensis (O. Ktze) damaged by the tea weevil Myllocerinus aurolineatus (Voss) (Coleoptera: Curculionidae), with weevil densities, photoperiod and infestation duration, were investigated. The volatiles induced by high‐density weevils were more abundant in composition and amount than those induced by low‐density weevils, whether at noon, night or after weevil removal. The induced volatile emissions were similar on the second and third day after infestation, and the emissions of the major induced compounds displayed diurnal cycles. Linalool, (E,E)‐α‐farnesene, and benzyl nitrile were emitted mainly at noon, whereas 1,3,8‐p‐menthatriene and (E)‐β‐ocimene were maximally emitted at night. Given the different emission dynamics, significant differences were found between noon‐ and night‐induced volatiles. In summary, tea plants damaged by different weevil densities emitted a relatively stable signal at a particular time. This stability could be attributed to the similarities under the two densities of the main induced volatile compounds, their relative ratios and the emission dynamics of the induced volatiles.     

An ecogenomic analysis of herbivore‐induced plant volatiles in Brassica juncea

Upon herbivore feeding, plants emit complex bouquets of induced volatiles that may repel insect herbivores as well as attract parasitoids or predators. Due to differences in the temporal dynamics of individual components, the composition of the herbivore‐induced plant volatile (HIPV) blend changes with time. Consequently, the response of insects associated with plants is not constant either. Using Brassica juncea as the model plant and generalist Spodoptera spp. larvae as the inducing herbivore, we investigated herbivore and parasitoid preference as well as the molecular mechanisms behind the temporal dynamics in HIPV emissions at 24, 48 and 72 h after damage. In choice tests, Spodoptera litura moth preferred undamaged plants, whereas its parasitoid Cotesia marginiventris favoured plants induced for 48 h. In contrast, the specialist Plutella xylostella and its parasitoid C. vestalis preferred plants induced for 72 h. These preferences matched the dynamic changes in HIPV blends over time. Gene expression analysis suggested that the induced response after Spodoptera feeding is mainly controlled by the jasmonic acid pathway in both damaged and systemic leaves. Several genes involved in sulphide and green leaf volatile synthesis were clearly up‐regulated. This study thus shows that HIPV blends vary considerably over a short period of time, and these changes are actively regulated at the gene expression level. Moreover, temporal changes in HIPVs elicit differential preferences of herbivores and their natural enemies. We argue that the temporal dynamics of HIPVs may play a key role in shaping the response of insects associated with plants.     

Caterpillar‐induced plant volatiles attract conspecific herbivores and a generalist predator

Plants release volatiles in response to caterpillar feeding that attracts natural enemies of the herbivores, a tritrophic interaction which has been considered to be an indirect plant defence against herbivores. On the other hand, the caterpillar‐induced plant volatiles have been reported to either repel or attract conspecific adult herbivores. This work was undertaken to investigate the response of both herbivores and natural enemies to caterpillar‐induced plant volatiles in apple orchards. We sampled volatile compounds emitted from uninfested apple trees, and apple trees infested with generalist herbivore the pandemis leafroller moth, Pandemis pyrusana (Lepidoptera, Tortricidae) larvae using headspace collection and analysed by gas chromatography/mass spectrometry. Infested apple trees uniquely release six compounds (benzyl alcohol, phenylacetonitrile, phenylacetaldehyde, 2‐phenylethanol, indole and (E)‐nerolidol). These compounds were tested on two species of herbivores and one predator in apple orchards. Binary blends of phenylacetonitrile + acetic acid or 2‐phenylethanol + acetic acid attracted a large number of conspecific male and female adult herbivores. The response of pandemis leafroller to herbivore‐induced plant volatiles (HIPVs) was so pronounced that over one thousand and seven hundred conspecific male and female adult herbivores were caught in traps baited with HIPVs in three‐day trapping period. In addition, significantly higher number of male and female obliquebanded leafroller, Choristoneura rosaceana (Lepidoptera, Tortricidae), was caught in traps baited a binary blend of 2‐phenylethanol + acetic acid, or a ternary blend contains 2‐phenylethanol and phenylacetonitrile + acetic acid. This result challenges the current paradigm hypothesized that HIPVs repel herbivores and question the indirect defensive function proposed for these compounds. On the other hand, a ternary blend of phenylacetonitrile and 2‐phenylethanol + acetic acid attracted the largest numbers of the general predator, the common green lacewing, Chrysoperla plorabunda. To our knowledge, this is the first record of the direct attraction of conspecific adult herbivores as well as a predator to the caterpillar‐induced plant volatiles in the field.     

Host plant defense signaling in response to a coevolved herbivore combats introduced herbivore attack

Defense‐free space resulting from coevolutionarily naïve host plants recently has been implicated as a factor facilitating invasion success of some insect species. Host plants, however, may not be entirely defenseless against novel herbivore threats. Volatile chemical‐mediated defense signaling, which allows plants to mount specific, rapid, and intense responses, may play a role in systems experiencing novel threats. Here we investigate defense responses of host plants to a native and exotic herbivore and show that (1) host plants defend more effectively against the coevolved herbivore, (2) plants can be induced to defend against a newly‐associated herbivore when in proximity to plants actively defending against the coevolved species, and (3) these defenses affect larval performance. These findings highlight the importance of coevolved herbivore‐specific defenses and suggest that naïveté or defense limitations can be overcome via defense signaling. Determining how these findings apply across various host–herbivore systems is critical to understand mechanisms of successful herbivore invasion.     

The role of the plant in attracting parasitoids: response to progressive mechanical wounding

Based on the model system of Brussels sprouts [Brassica oleracea var. gemmifera (Brassicaceae)], the herbivore cabbage white caterpillar, Pieris brassicae (L.) (Lepidoptera: Pieridae), and the parasitoid wasp, Cotesia glomerata (L.) (Hymenoptera: Braconidae), the influence of plant damage type, and damage duration were assessed on plant volatile emission and subsequent recruitment of natural antagonists of the herbivore. Plants were damaged by three methods for a period of either 3 or 8 h: herbivore damage (HD), progressive mechanical damage, and final mechanical damage inflicted in a single event. Wind‐tunnel bioassays evaluated whether the mode of damage affected female parasitoid oriented flight. After both periods of damage, all treatments were highly significantly preferred by naïve C. glomerata to undamaged control plants. After 3 h, herbivore‐damaged plants were significantly preferred to plants with final damage (FD). Most remarkably, following 8‐h damage, the parasitoid preferred both herbivore‐damaged and progressively damaged plants to plants with FD and did not significantly discriminate between herbivore and progressively damaged plants, thus indicating a similarity in plant response to herbivore and progressive mechanical damage. In addition to wind‐tunnel bioassays, emitted plant volatiles were collected and analysed by thermal desorption gas chromatography/mass spectrometry, following 3 and 8 h of damage in order to correlate volatiles released from different damage types with the attraction of the parasitoid. Differences in volatile profiles from all damage types were similar following both 3 and 8 h of damage, with only (Z)‐3‐hexenyl acetate found to be emitted in significantly higher quantities by final mechanical damage compared with HD after 3 h. In conclusion, the plant's response to progressive mechanical damage was more similar to HD than final mechanical damage deployed at a single point in time, irrespective of damage duration, and C. glomerata did not significantly discriminate between progressive damage and HD.     

Host‐searching responses to herbivory‐associated chemical information and patch use depend on mating status of female solitary parasitoid wasps

1. In a tritrophic interaction system consisting of plants, herbivores, and their parasitoids, chemicals released from plants after herbivory are known to play important roles for many female parasitoids to find their hosts efficiently. On the plant side, chemical information associated with herbivory can act as an indirect defence by attracting the natural enemies of the host herbivores. 2. However, mated and virgin females of haplodiploid parasitoids might not necessarily respond to such chemical cues in the same way. Since virgin females can produce only sons, they might refrain from searching for hosts to invest eggs until copulation, in order to produce both sexes. 3. Here, we investigated differential host‐searching behaviours shown by mated and virgin females in the solitary parasitoid wasp, Cotesia vestalis, in response to herbivory‐associated chemical information from cruciferous plants infested by their host larvae, Plutella xylostella. 4. Mated females showed a significantly higher flight preference for host‐infested plants over intact plants, while no preference was observed with virgin females. Mated females also showed more intensive antennal searching and ovipositor probing behaviours to leaf squares with wounds caused by hosts than did virgin females. Furthermore, mated females stayed longer in host patches with higher parasitism rates than virgin females. 5. These results indicate that mating status of C. vestalis females clearly influences their host‐searching behaviour in response to herbivory‐associated chemical information and patch exploitation. Female parasitoids seem to forage for hosts depending on their own physiological condition in a tritrophic system.     

Effects of quantitative and qualitative differences in volatiles from host‐ and non‐host‐infested maize on the attraction of the larval parasitoid Cotesia kariyai

Cotesia kariyai Watanabe (Hymenoptera: Braconidae) is a specialist larval parasitoid of Mythimna separata Walker (Lepidoptera: Noctuidae). Cotesia kariyai wasps use herbivore‐induced plant volatiles (HIPVs) to locate hosts. However, complex natural habitats are full of volatiles released by both herbivorous host‐ and non‐host‐infested plants at various levels of intensity. Therefore, the presence of non‐hosts may affect parasitoid decisions while foraging. Here, the host‐finding efficiency of naive C. kariyai from HIPVs influenced by host‐ and non‐host‐infested maize [Zea mays L. (Poaceae)] plants was investigated with a four‐arm olfactometer. Ostrinia furnacalis Guenée (Lepidoptera: Crambidae) was selected as a non‐host species. One unit (1 U) of host‐ or non‐host‐infested plant was prepared by infesting a potted plant with five host or seven non‐host larvae. In two‐choice bioassays, host‐infested plants fed upon by different numbers of larvae, and various units of host‐ and non‐host‐infested plants (infestation units; 1 U, 2 U, and 3 U) were arranged to examine the effects of differences in volatile quantity and quality on the olfactory responses of C. kariyai with the assumption that volatile quantity and quality changes with differences in numbers of insects and plants. Cotesia kariyai was found to perceive quantitative differences in volatiles from host‐infested plants, preferring larger quantities of volatiles from larger numbers of larvae or plants. Also, the parasitoids discriminated between healthy plants, host‐infested plants, and non‐host‐infested plants by recognising volatiles released from those plants. Cotesia kariyai showed a reduced preference for host‐induced volatiles, when larger numbers of non‐host‐infested plants were present. Therefore, quantitative and qualitative differences in volatiles from host‐ and non‐host‐infested plants appear to affect the decision of C. kariyai during host‐habitat searching in multiple tritrophic systems.