Female adult puncture-induced plant volatiles promote mating success of the pea leafminer via enhancing vibrational signals

Abstract

Herbivore-induced plant volatiles (HIPVs) synergize with the sex pheromones of herbivorous insects to facilitate mate location. However, the synergism of HIPVs and acoustic signals for sexual communication remains unknown. Here, we investigated the synergy between HIPVs and vibrational duets for sexual communication and mating in the pea leafminer (Liriomyza huidobrensis). Our results indicated that adult leafminers do not produce species-specific pheromone, and female-puncture-induced plant volatiles facilitate the attraction of both sexes to host plant leaves and sexual encounters. Insect-derived cues do not participate in mate locations. Both sexes do not produce qualitatively different cuticular hydrocarbons (CHCs), and CHCs from females cannot elicit the antennal and behavioural responses of males. By contrast, induced green leaf volatiles, terpenoids and oximes elicit dramatic antennal responses in both sexes. Electrophysiological and behavioural tests consistently showed that the volatiles (Z)-3-hexenol and (Z)-3-hexenyl-acetate elicited the most intense gas chromatographic-electroantennographic responses, and attracted males and females. Remarkably, these volatiles significantly promoted the occurrence of vibrational duets between the sexes, thereby increasing the mating success of leafminers. Therefore, the synergism of HIPVs and vibrational signals largely promoted the mating success of leafminers, suggesting an alternative control strategy through precision trapping for non-pheromone-producing insects.

This article is part of the theme issue ‘Biotic signalling sheds light on smart pest management’.

1. Introduction

Plants respond to herbivore attacks by releasing specific blends of herbivore-induced plant volatiles (HIPVs), and some of them are biologically active compounds that promote plant survival by acting as the first lines of defence against herbivory [1–3]. The roles of HIPVs in tritrophic interactions among plants, herbivores and natural enemies have been extensively studied [4–8]. HIPVs can also trigger the priming defence of neighbouring unattacked plants [8–10]. Importantly, laboratory and fieldwork have shown that HIPVs are promising agents in push–pull strategies against several crop insect pests [11–13].

Given that plants provide nutrient-rich foods, habitats and mating sites, herbivores can also benefit from the use of HIPVs as important foraging cues. For example, leaf- and root-feeding beetles use conspecific species-induced HIPVs to aggregate on host plants for feeding and development [14,15]. HIPVs facilitate mate finding, which is mainly mediated by species-specific pheromones [16]. Green leaf volatiles (GLVs) enhance the attractiveness of sex pheromones in boll weevils, bark beetles, Mediterranean fruit flies and tobacco budworms [17–19]. Several terpenoids and aromatic compounds also intensify the trapping effects of synthetic female-produced pheromones in lepidopterous moths [20–22]. Some herbivorous insect species demonstrate increased mating activities and pheromone production upon perceiving volatiles released from host plants [23–25]. Therefore, HIPVs probably play additive roles in the sexual encounters and communications of herbivorous insects. Although pheromones are almost ubiquitous among arthropod species and effective over short to long spatial ranges [26,27], agromyzid flies use vibration for sexual communication on leaves [28] and the mating systems of specific insect species may even lack sex pheromones [29,30]. Under these circumstances, plant volatiles and conspecific species-induced HIPVs provide a reliable signal for the conundrum of habitat location and sex encounter [16]. However, reports illustrating that the synergistic effects of HIPVs and acoustic and vibrational signals on sexual communication and mating success remain insufficient.

The pea leafminer Liriomyza huidobrensis is a polyphagous pest of vegetables and ornamental plants worldwide [31]. The management of this pest requires novel control strategies given its resurgence and high resistance to chemical sprays [32]. Leafminers can employ plant volatiles for host location [33–35]. Both sexes of Liriomyza sativae respond positively to headspace volatile mixtures from host plants, especially those from damaged host plants [35]. Damage caused by female puncturing and larval feeding induce the emissions of a large amount of HIPVs, including GLVs, terpenoids and oximes, and the emitted volatiles provide signals that are useful for conspecific and heterospecific species [36]. Mutant tomato plants with reduced HIPV production are less attractive to adult leafminers than wild-type tomato plants [37]. These results indicate that HIPVs have crucial roles in host location by leafminers. However, to date, the behavioural responses of leafminers to specific HIPVs, volatile blends or specific chemical compounds derived from adult leafminers remain unknown. Despite several studies on the geographical phylogeny, cold tolerance and tritrophic interaction of agromyzid flies [32,38], sex communication and its underlying mechanism in agromyzid flies remain poorly understood. Recently, we reported that the leafminer individuals use vibrational duets for sexual communication on plant leaves. The playback of these sexual signals can elicit orientation behaviours in the opposite sex, implying that vibrational signals have potential applications in the trapping control of Liriomyza pests [28]. Given that adult females puncture leaf surfaces with ovipositors to facilitate feeding and oviposition and induce host plants to release a specific blend of volatiles, HIPVs may serve as important cues for sexual encounters. Thus, we hypothesized that HIPVs may coordinate with vibrational signals to facilitate the mating success of leafminers.

In this study, we aimed to determine whether (i) adult leafminers of both sexes use species-specific pheromones for mate finding; (ii) specific female-puncture-induced volatiles attract both sexes to host plants; and (iii) HIPVs and vibrational signals jointly facilitate sexual interaction and mating success. Our results demonstrated that the synergism of HIPVs and vibrational communication could significantly promote the mating success of leafminers and that the combined use of plant volatiles and acoustic signals was a potential strategy for agromyzid fly control through precision trapping by simulating specific HIPVs and vibrational waves.

2. Material and methods

(a). Plants and animals

Plants and animals were obtained as described previously [28]. Kidney bean plants (Phaseolus vulgaris L. cv Naibai) were cultivated in plastic pots (8 cm in diameter) filled with vermiculite and peat (1 : 2) in environmental chambers (25 ± 2°C, 14 L : 10 D photoperiod, 60% relative humidity and 30 000 lm m⁻²). Two-week-old bean plants with two fully developed true leaves were used in insect rearing and in our experiments. The population of the pea leafminer was maintained on kidney bean plants in the environmental chambers. Upon emerging from pupae, virgin adults were individually isolated in microtubes (1.5 ml cryogenic vial, Nalgene 5000–1020, USA) and then fed with 10% diluted honey. Two-day-old mature adults were used in all of the experiments.

(b). Experiment 1: olfactory responses to adult leafminer volatiles

(i). Behavioural bioassays

To determine whether adult leafminers use species-specific pheromones to facilitate mutual attraction between the sexes, we used a four-armed olfactometer to quantify the selection response of adults to the odours of the opposite sex. The behavioural set-up was designed as previously described [39] with some modifications (electronic supplementary material).

(ii). Gas chromatography–electroantennographic detection recording

A solid-phase microextraction (SPME) fibre (polydimethylsiloxane/divinylbenzene at 65 µm) was used to adsorb chemicals from a group of 30 two-day-old adult females for 30 min in a screw-top Supelco vial (2 ml). Thus, SPME extracts included volatiles and nonvolatiles. SPME fibres used for adsorption in empty vials served as the control. Antennae were prepared as previously reported [34]. For details, see the electronic supplementary material.

(iii). Collections and analyses of adult body extracts

Cuticular hydrocarbons (CHCs) were extracted by immersing 30–40 virgin 2-day-old adult flies of each sex in hexane (5 µl per fly) for 10 min. CHC extracts were transferred to 2 ml screw cap vials (Waters Co., USA) with Teflon/rubber septa and then stored at −20°C until further analysis. Chemical identification and analysis are described in detail in the electronic supplementary material.

(c). Experiment 2: olfactory responses to herbivore-induced plant volatiles

(i). Behavioural bioassays

We employed the same four-armed olfactometer to determine whether adult flies are attracted to HIPVs produced by adult female-punctured plants. Female-punctured leaves were prepared by placing one healthy bean plant in a mesh cage containing 20 female flies and then transferred to the environmental chamber for 16 h (from 17.00 to 9.00). For details, see the electronic supplementary material.

The preferences of females and males for individual compounds were assessed in a Y-tube (stem, 10 cm; arms, 23 cm at 60°; internal diameter, 2.3 cm) as previously reported [7]. For details, see the electronic supplementary material.

(ii). Gas chromatographic–electroantennographic detection recording

To identify the bioactive volatiles that are present in female-puncture-induced HIPVs, we measured the antennal responses of both sexes to common GLVs, terpenoids and oximes by using the same gas chromatographic–electroantennographic detection (GC–EAD) recording set-up and method described above with some modifications. For details, see the electronic supplementary material.

(iii). Collections and analyses of herbivore-induced plant volatiles

To test if females emit specific volatiles in the presence of host plants, we compared the profiles of volatiles from female-punctured leaves with those from females plus female-punctured leaves (electronic supplementary material, table S2). The female-punctured leaves were prepared as described above in Behavioural bioassays. For details, see the electronic supplementary material.

(d). Experiment 3: effect of herbivore-induced plant volatiles on vibrational communication

To identify the effect of HIPVs on vibrational signals and copulation, we used a cubic observation cage (15 cm × 15 cm × 20 cm) designed in accordance with a previous study [28]. Tests were performed between 9.00 and 17.00 at 25°C. Six substrates, namely, mesh, punctured leaves, a plant photo, mesh plus synthetic hexane, mesh plus synthetic HIPVs blend and mesh plus GLVs blend, were introduced to the bottom of the arena. The HIPV and GLV blends were prepared as described above at dosages of 1050 and 580 ng in 10 µl of hexane (electronic supplementary material, table S2). Blends and hexane were taken up into glass capillary tubes (10 µl; Sigma-Aldrich, Steinheim, Germany). A female was initially introduced into a cage 5 min before the test. A male was subsequently introduced. Two video cameras were used to record the behaviour of both sexes. When a male and a female meet in a cage, they generate vibrational duets; during this process, the female generates a conspicuous bobbing signal, and the male approaches the female [28]. The copulation occurrence was also recorded. Cases in which males and females met and performed duets were analysed and the location of flies was recorded with Observer 11 (Noduls Information Technology, Wageningen, The Netherlands). On each experimental day, 10 pairs were observed. A total of 60 pairs were monitored for each treatment.

(e). Statistical analysis

A χ²-test was performed to identify the significance of differences among the proportions of leafminers in the behavioural assays conducted in experiments 1 and 2 and to analyse the duet location identified in experiment 3. Electrophysiological data for experiments 1 and 2 were compared through one-way ANOVA followed by Tukey's honest significant difference (HSD) post hoc tests. The absolute amount of CHCs in experiment 1 was subjected to log(X + 1) transformation and then compared through a Student's t-test. The percentages of plant volatiles determined in experiment 2, the proportion of duetting and copulation and the time spent on the platform observed in experiment 3 were subjected to arcsin(X^1/2) transformation and compared through ANOVA and Tukey's HSD test. All of the tests were performed using SPSS v. 17.0 (SPSS Inc., Chicago, IL). Prior to statistical analysis, data were checked for normality through one-sample Kolmogorov–Smirnov tests.

3. Results

(a). Olfactory responses of adult leafminers to insect-derived chemicals

We quantified the responses of 2-day-old sexually mature virgin males and females in upwind flight to volatiles from opposite sexes in a four-armed olfactometer (figure 1a) to examine the effects of conspecific volatiles on both sexes (figure 1b). Neither volatiles from females nor from males could evoke remarkable upwind preferences in males or females (male to female volatiles: χ² = 0.40, p = 0.53, n = 21; male to male volatiles: χ² = 1.11, p = 0.30, n = 30; female to female volatiles: χ² = 0, p = 1, n = 20; female to male volatiles: χ² = 0.462, p = 0.50, n = 26; figure 1b). These results demonstrated that the volatiles emitted by sexually mature adults failed to induce attraction, aggregation or repulsion behaviours.

Figure 1.

Olfactory preferences and GC–EAD responses of conspecific adults to volatiles and CHCs. (a) Schematic of the four-armed olfactometer. The odour source was delivered to one arm, and clean air was delivered to the three other arms. Flies were released into the central glass chamber as indicated by the black arrow. The numbers of adults present in the insect-trapping bulbs were recorded. (b) Male and female preferences for conspecific odours in the four-armed olfactometer (χ² test, n.s., not significant). Dashed lines indicate the expected distribution (1 : 3) of choices between odour and air. n.c., no choice; ♂, male; ♀, female. (c) Representative gas chromatogram of CHCs derived from female (blue, lower peaks) and male (red, upper peaks) flies. The numbers above the peaks refer to the compounds listed in the electronic supplementary material, table S1. ♂, male; ♀, female. (d) Simultaneously recorded antennal responses of 2-day-old males (blue trace) to SPME-absorbed female body chemicals (black trace, upper) or to SPME-absorbed chemicals (black trace) from a blank vial as the control (lower). FID, flame ionization detection. Gas chromatogram peaks of CHCs with retention times (RTs) of 13 min to 16 min are listed in the electronic supplementary material, table S1. (Online version in colour.)

To test if sex-specific chemicals were present in the bodies of leafminer adults, we analysed the organic compounds washed from the whole bodies of sexually mature virgin adults. Gas chromatography–mass spectrometry (GC–MS) detected 22 CHCs (saturated C23-C29) in extracts from 2-day-old adults of both sexes (figure 1c and electronic supplementary material, table S1). These CHCs were not qualitatively different between the two sexes (electronic supplementary material, table S1). Females produced higher quantities of alkanes than males did (t-test, t₄ = 3.216, p = 0.032) because of the larger body size of the former [40]. These results suggested that leafminer adults did not produce sex-specific chemicals.

To further verify the behavioural responses of the flies to volatiles and to test the coupled GC–EAD responses to nonvolatile compounds derived from the adult bodies, we measured the antennal responses of males to the SPME extract of adult females. GC–EAD analysis showed that volatiles and nonvolatiles (corresponding to the compounds shown in figure 1d) could not elicit the specific and consistent antennal responses of males (volatiles: compounds with retention times of 4–10 min; nonvolatiles: compounds with retention times of 13–16 min; figure 1d). These results confirmed that the leafminer adults did not emit biologically active volatiles and that nonvolatile chemicals also could not elicit substantial electrophysiological activities. Thus, the pea leafminer adults did not produce the species-specific pheromone for sexual communication.

(b). Olfactory responses of adult leafminers to female adult puncture-induced herbivore-induced plant volatiles

To test if female-puncture-induced HIPVs could act as signals for conspecific individual communication, we used the four-armed olfactometer to quantify the behavioural responses of both sexes to volatile emissions from plants. In comparison with the air control, the puncture-induced blend evoked the significant upwind preferences of both sexes (male to female-punctured leaves: χ² = 37.56, p < 0.0001, n = 24; female to female-punctured leaves: χ² = 26.89, p < 0.0001, n = 24; male versus female: χ² = 0.44, p = 0.51; figure 2a). Thus, both sexes could use female-induced HIPVs as cues for the host location.

Figure 2.

Olfactory preferences and GC–EAD responses to volatiles from female-punctured leaves and the synthetic chemicals. (a) Preferences of males and females to volatiles from female-punctured leaves versus air control in a four-armed olfactometer. (b) Dual choices of males to volatiles from female-punctured leaves and from female-punctured leaves plus females in a four-armed olfactometer. Pie chart indicates the percentages of males (black, males choose odours; light grey, air control; white, n.c., no choice). (c) Amplitudes of spikes in male and female GC–EAD responses to six volatiles in a synthetic blend (mean ± s.e.). Bars labelled with different letters indicate statistically significant differences in average amplitude for different compounds (ANOVA, followed by Tukey's HSD test, p < 0.05). (d) Preferences of males and females in a Y-tube to synthesized single compounds. Compounds were prepared at dosages equivalent to the natural release rate in 1 h (electronic supplementary material, table S2). (a–d) Dashed lines represent the expected distribution in olfactometers. χ²-test for differences between the numbers of leafminers in each arm ; *p < 0.05, **p < 0.01, ***p < 0.001, n.s., not significant; n.c., no choice. Numbers in bars are the numbers of adults choosing the indicated odour source. ♂, male; ♀, female. (c,d) Bars labelled with different colours indicate GLVs (green), aldoximes (cyan), terpenoids (red) and air control (grey). (Online version in colour.)

To test if HIPVs could trigger the release of species-specific pheromones, we compared the volatile blends of female-punctured leaves and those of virgin females plus female-punctured leaves (female–plant complex). The volatile quality or quantity of the two blends did not differ significantly (electronic supplementary material, table S2). In addition, no insect-derived chemical was detected in the female–plant complex (electronic supplementary material, table S2). The two blends were equally attractive to males (χ² = 0, p = 1, n = 34; figure 2b). This result confirmed that leafminers lacked species-specific volatiles.

To identify the electrophysiologically active compounds in the tested volatiles, we examined the GC–EAD responses of both sexes to a synthetic volatile blend comprising major GLVs, terpenoids and oximes. These volatiles significantly elicited antennal responses from both sexes. Female antennae were more highly responsive to (Z)-3-hexenol, (Z)-3-hexenyl-acetate, (3E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), β-caryophyllene and 2-methylpropanal oxime than to 2-methylbutanal oxime (F = 5.69, p < 0.001; figure 2c). Male antennae were most responsive to (Z)-3-hexenol and highly tuned to DMNT and (Z)-3-hexenyl-acetate (F = 15.77, p < 0.0001; figure 2c).

To identify behaviourally active volatiles, we used the Y-tube to assess the insects' behavioural responses to single compounds. (Z)-3-hexenol and (Z)-3-hexenyl-acetate elicited the most intense GC–EAD responses and the most intense behavioural activities ((Z)-3-hexenol: males: χ² = 4, p = 0.046; females: χ² = 4, p = 0.029; (Z)-3- hexenyl-acetate: χ² = 6.38, p = 0.012; females: χ² = 9.64, p = 0.002; figure 2d). However, we did not observe the attractive effect of DMNT, β-caryophyllene, 2-methylbutanal oxime and 2-methylpropanal oxime on the preference of both sexes of adult leafminers (figure 2d).

(c). Promotion of the copulation behaviours by female-puncture-induced herbivore-induced plant volatiles and vibrational duetting

To investigate the synergism of HIPVs and vibrational signals for sexual communication and mating success, we recorded the courtship behaviour of the pairs in the presence of different stimuli (figure 3a). The proportions of the duetting pairs were higher in the female-punctured leaves in the mesh cages than in the empty control or a leaf photo in the mesh cages (F = 12.99, p < 0.01; figure 3b). Consistently, the proportions of duetting pairs were higher in the six-compound blend consisting of GLVs, terpenoids and oximes, or in the two-compound blend of (Z)-3-hexenol and (Z)-3-hexenyl-acetate than in the hexane solvent in the mesh cages (F = 14.95, p < 0.01; figure 3b). These results indicated that the HIPVs, particularly GLVs, increased the occurrence of duetting in the leafminers. When the duets were categorized on the basis of their location, the proportions of duetting on the platform where the stimulus was placed, were higher in the presence of punctures than in the absence of punctures (χ² = 31.00, p < 0.001; figure 3c). Consistently, the proportions were higher in the presence of the synthetic blends than in the presence of the hexane solvent control (HIPVs blend: χ² = 16.32, p < 0.001; GLVs blend: χ² = 10.38, p < 0.01; figure 3c). As a consequence, the proportions of the copulating pairs in the presence of the female-punctured leaves or the synthetic blends were more than two times higher than those in the absence of punctures or blends, respectively (empty control versus leaf photo versus punctured leaves: F = 8.77, p = 0.003; hexane versus HIPVs versus GLVs: F = 7.33, p = 0.006; figure 3d).

Figure 3.

Vibrational courtship and copulation behaviours in mesh cages in the presence of different stimuli. (a) Schematic of the arena employed to observe courtship behaviour. Adults were released into a mesh cage. Stimuli were introduced to the platform. The HIPV and GLV blends were prepared at dosages of 1050 and 580 ng in 10 µl of hexane (electronic supplementary material, table S2), respectively. (b) The proportion of duetting pairs in the presence of six stimuli (ANOVA, followed by Tukey's HSD test). (c) Percentages of duetting paired flies in the cage or the platform in the experimental arena (χ²-test). (d) The proportion of copulating pairs in the presence of six stimuli (ANOVA, followed by Tukey's HSD test). (e) Relative time spent by males and females on the platform (ANOVA, followed by Tukey's HSD test). ♂, male; ♀, female. (b–e) Data are shown as mean ± s.e. *p < 0.05, **p < 0.01. (Online version in colour.)

Through the analysis of male and female locations in the courtship assay, we found that both sexes spent significantly more time on the platform when they were provided with the female-punctured leaves (empty control versus leaf photo versus punctured leaves: male, F = 15.23, p < 0.001; female, F = 31.02, p < 0.001; figure 3e), HIPVs and GLVs than when they were provided with the controls (hexane versus HIPVs versus GLVs: male, F = 27.11, p < 0.001; female, F = 16.28, p < 0.001; figure 3e).

4. Discussion

In this study, we showed that HIPVs can enhance the vibrational communication of the leafminers, increase the attractiveness of the host plant, and promote the mating success. Our findings expanded the current understanding of the synergistic effect of HIPVs on other signalling systems involved in the sexual communication of herbivorous insects.

Although many studies have found that HIPVs exert synergistic or additive effects on insect sex pheromones, we did not discover a volatile sex pheromone that functioned in long-range attraction in pea leafminers. Moreover, we did not detect any gender-specific chemicals in adult cuticular extracts and headspace volatiles with potential functions in intersexual attraction. These results confirmed previous claims that this leafminer species may lack sex pheromones [32,40]. The functional reduction of sex pheromones has been reported in various insect groups. For example, some parasitic wasps do not use sex pheromones for long-range conspecific attraction [30], and male fruit flies do not release sex pheromones in the absence of plant odours [41]. Despite their various advantages, the production of sex pheromones may incur an energetic cost [30], increase the risks of eavesdropping by natural enemies [42] and induce the emission of alert signals by plants [43]. Given that HIPVs are highly attractive and have low molecular weights that enable their dispersion across long distances, the reliability and detectability of HIPVs may have driven leafminer adults to reduce the reliance on sex pheromones. This phenomenon implies the parsimonious evolution of sex signals under the selection pressure of complex environmental factors. However, this speculation needs further evidence from other species of agromyzid flies.

We found that volatiles from the leaves punctured by the females attracted the leafminer males and females. In Liriomyza species, pupae remain in the soil, and adults need to locate host plants after eclosion occurs [40]. Adult females puncture leaves to feed, lay eggs and assess plant quality [40]. Thus, female-puncture-induced HIPVs may convey signals about host quality and act as feeding and oviposition cues for conspecifics. For males, female-induced HIPVs provide clues for potential partners and food because of their feeding reliance on females. Thus, female-induced HIPVs serve as an indicator of food and mating sites. HIPVs play important roles in food and mate location [16]. HIPVs are ubiquitously released by various plant species and considered the most sensitive components to wounding [7]. We showed that (Z)-3-hexenol and (Z)-3-hexenyl-acetate were attractive to adult male and female leafminers. Given the diverse host range and sympatric distributions of some Liriomyza pest species, their acute responses to HIPVs may be an effective strategy for food and mate location. We proposed two explanations for the precise responses of different species to HIPVs during food and mate location. Firstly, herbivores and their developmental stages can affect HIPV composition. For example, individual components in the HIPV profiles of plant leaves quantitatively and qualitatively vary in response to herbivory by different species and leafminers in various developmental stages [36]. The ratio of volatiles can provide specific information to herbivores [44]. As such, different ratios of the main volatiles may be used for host assessment and spatial separation. Secondly, pairs of leafminers on plants use vibrational duets for efficient communication [28]. Given that vibrational communication modalities are species-specific across various taxa [30], HIPVs serve as common cues for host plant location among Liriomyza species, and vibrational signals are responsible for the accurate recognition of the sex and specific species of potential mates. Further studies should verify these hypotheses with other leafminer species.

In comparison with other signals, such as leaf photos, female-induced volatiles induced a high proportion of pairs to perform duetting and copulation. This result may be attributed to the synergistic effect of HIPVs and leaf-borne vibrations on facilitating mating success. Alternatively, the stimulatory effect of plant volatiles on the emission of sex signals may account for this result. Indeed, male and female insects can increase sex signalling in response to food odours. For example, the presence of food odours extends the duration of courtship songs produced by Drosophila males [45]. Several herbivores begin to release pheromones or increase mating activity after they sense plant volatiles [41,46]. Regardless of the mechanism underlying the effect of HIPVs on sex signals, the combined use of food odours and sex signals may have evolved as a common behavioural pattern for foraging and mating successes among phytophagous insects.

In the pea leafminer, the playback of female response signals is sufficient to elicit mate-searching behaviour in all males [28]. To increase the vibration-based trap effect, adult insects must be aggregated on the same substrates. In this study, we showed that HIPVs served as attractants over a large spatial range and promoted the possibility of vibrational communication. Thus, the use of HIPVs represents an opportunity for the application of chemical signals to enhance the effective space of acoustic traps. In addition, the combined use of plant volatiles and acoustic signals can increase control in the precision trapping of target insect pests and eliminate negative effects on natural enemies, given the sensitivity of beneficial insects to HIPVs.

In summary, our findings provided the synergistic mechanism of HIPVs and acoustic signal in the sexual interaction of leafminers. The benefit and cost analysis of HIPVs released by plants in the same or different systems is necessary to understand the evolution and ecology of HIPVs and to optimize their practical applications.

Ethics

The pea leafminer is reared in the laboratory and handled following local guidelines.

Data accessibility

The datasets supporting this article have been uploaded as part of the electronic supplementary material. Additional data available from authors upon request.

Authors' contributions

J.G., J.N.W. and L.K. generated conception and design. J.G., J.N.W., N.L. and Y.N.Y. acquired data. J.G. and J.N.W. analysed and interpreted data. J.N.W., J.G. and L.K. drafted the article. All authors approved the version to be published.

Competing interests

We have no competing interests.

Funding

This work was supported by the Strategic Priority Research Program of the Chinese Academy of Sciences (grant XDB11050600), National Key R&D Program of China (2017YFD0200400) and the National Nature Science Foundation of China (31170361).