Plant volatile organic compounds (VOCs) are major vehicles of information transfer between organisms and mediate many ecological interactions []. Altering VOC emission in response to herbivore damage has been hypothesized to be adaptive, as it can deter subsequent herbivores [], attract natural enemies of herbivores [], or transmit information about attacks between distant parts of the same plant []. Neighboring plants may also respond to these VOC cues by priming their own defenses against oncoming herbivory, thereby reducing future damage []. However, under which conditions such information sharing provides fitness benefits to emitter plants, and, therefore, whether selection by herbivores affects the evolution of such VOC signaling, is still unclear []. Here, we test the predictions of two alternative hypotheses, the kin selection and mutual benefits hypotheses [], to uncover the selective environment that may favor information sharing in plants. Measuring the response to natural selection in Solidago altissima, we found strong effects of herbivory on the way plants communicated with neighbors. Plants from populations that experienced selection by insect herbivory induced resistance in all neighboring conspecifics by airborne cues, whereas those from populations experiencing herbivore exclusion induced resistance only in neighbors of the same genotype. Furthermore, the information-sharing plants converged on a common, airborne VOC signal upon damage. We demonstrate that herbivory can drive the evolution of plant-plant communication via induction of airborne cues and suggest plants as a model system for understanding information sharing and communication among organisms in general.

Results and Discussion

12 Karban R.

Yang L.H.

Edwards K.F. Volatile communication between plants that affects herbivory: a meta-analysis. 15 Karban R. Plant behaviour and communication. 13 Heil M.

Karban R. Explaining evolution of plant communication by airborne signals. 16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. 17 Hamilton W.D. The genetical evolution of social behaviour. II. 16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. Plant-plant communication occurs when undamaged receivers induce resistance in response to a cue from a damaged neighbor, the emitter []. One popular explanation for the seemingly common phenomenon of volatile organic compound (VOC) perception by plants is that neighboring plants tune in on cues that evolved for within-plant signaling or trophic interactions []. These eavesdropping neighbors can achieve a competitive advantage over the emitter by gaining information about oncoming herbivores and readying their defenses preemptively. However, there are two hypotheses that could explain the emergence of VOC-mediated communication between different plant individuals in a way that would ultimately benefit the emitter of the information. First, the emitter may indirectly benefit by transmitting information about herbivory exclusively to their own kin (i.e., individuals especially likely to share genes) or among clones of the same plant, thereby increasing its inclusive fitness []. Such a scenario should select for the emission of genotype-specific signals that only kin can perceive, decipher, and respond to. This should simultaneously favor signals that minimize eavesdropping by neighboring non-related plants. Hence, under this kin selection hypothesis, selection by herbivores is expected to favor private communication channels among kin and lead to divergence of VOC blends between non-related genotypes [].

13 Heil M.

Karban R. Explaining evolution of plant communication by airborne signals. 18 Heil M. Herbivore-induced plant volatiles: targets, perception and unanswered questions. 19 Bruin J.

Dicke M. Chemical information transfer between wounded and unwounded plants: Backing up the future. 20 Rubin I.N.

Ellner S.P.

Kessler A.

Morrell K.A. Informed herbivore movement and interplant communication determine the effects of induced resistance in an individual-based model. 21 Morrell K.

Kessler A. Plant communication in a widespread goldenrod: keeping herbivores on the move. 14 Kessler A.

Kalske A. Plant secondary metabolite diversity and species interactions. Figure 1 Communicating Plants Show full caption (A and B) Tall goldenrod, Solidago altissima (A), transmits chemical information to neighbors about attack by herbivores, such as the larvae of the goldenrod leaf beetle, Trirhabda virgata (B). (C) Adult T. virgata. Alternatively, under the mutual benefits hypothesis, communicating the risk of herbivory benefits the individual emitters regardless of their relatedness to the receivers [] if information exchange between neighbors promotes herbivore movement away from the emitter plant or a patch of neighbors []. Inducing resistance in neighboring plants may facilitate dispersion of herbivory and reduce the probability of herbivores (re-) entering the ever-increasing patch of resistant plants, yielding a benefit to the emitter. Here, emitter plants would benefit from signaling all neighbors, and thus, herbivory is predicted to select for information sharing regardless of kinship, open-communication channels, and the convergence of VOC blends among genotypes []. By measuring the response to natural selection by herbivores, we test whether herbivory drives the evolution of VOC-mediated communication in tall goldenrod Solidago altissima ( Figure 1 ) and address the predictions of the two plant-plant communication hypotheses.

2 = 22.16, df = 1, p < 0.001; 21 Morrell K.

Kessler A. Plant communication in a widespread goldenrod: keeping herbivores on the move. 2 = 23.03, df = 1, p < 0.001). Similar findings have been reported from sagebrush plants, Artemisia tridentata, that strongly respond only to VOCs emitted from damaged kin, but not non-kin genotypes [ 16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. 22 Karban R.

Wetzel W.C.

Shiojiri K.

Ishizaki S.

Ramirez S.R.

Blande J.D. Deciphering the language of plant communication: volatile chemotypes of sagebrush. Figure 2 Induction of Resistance in Response to Self and Non-Self Neighbors Show full caption damage / R control ), where R damage and R control are resistance in receiver plants when exposed to damaged and control emitter plants, respectively. We calculated resistance as R = (1 − [damaged leaves/total leaf number]). The graph shows means ± 95% confidence intervals. Asterisks indicate significant induction of resistance at 0.05 level. See also Number of unique genotypes was 32 and 29 from H+ and H− plots, respectively. Number of replicates/genotypes in each treatment was as follows: H+ self C = 14/13, D = 32/30; H+ non-self C = 24/23, D = 26/25; H− self C = 18/17, D = 26/25; and H− self C = 25/24, D = 21/21. We calculated induction of resistance using response ratios as ln(R/ R), where Rand Rare resistance in receiver plants when exposed to damaged and control emitter plants, respectively. We calculated resistance as R = (1 − [damaged leaves/total leaf number]). The graph shows means ± 95% confidence intervals. Asterisks indicate significant induction of resistance at 0.05 level. See also Figure S1 and Tables S1 and S2 We conducted a field experiment in the native habitat, whereby dedicated potted emitter plants (damaged by the chrysomelid specialist Trirhabda virgata [ Figure 1 B] or undamaged controls) were placed in the center of a group of potted receiver plants that were of the same genotype (self) and different genotypes (non-self) as emitter ( Figure 2 ). In this experiment, undamaged receiver plants, exposed to VOCs from herbivore-attacked (emitter) plants, experienced lower herbivory (mean ± SE of proportion of damage leaves; 0.228 ± 0.019) than those receivers exposed to undamaged control emitters (0.378 ± 0.02; generalized linear mixed model, damage treatment: Χ= 22.16, df = 1, p < 0.001; Table S1 ). This result confirmed earlier findings [] that VOCs from a damaged neighbor can elicit resistance in S. altissima under realistic field conditions. In general, receiver plants responded more strongly to self than non-self emitters, resulting in lower damage when the neighbor was a self emitter (0.181 ± 0.023) compared to a non-self emitter (0.285 ± 0.0311; damage treatment × relatedness: Χ= 23.03, df = 1, p < 0.001). Similar findings have been reported from sagebrush plants, Artemisia tridentata, that strongly respond only to VOCs emitted from damaged kin, but not non-kin genotypes [], and non-kin genotypes and kin express similar VOC chemotypes [].

2 = 4.05; df = 1; p = 0.044). In our field experiment, approximately half of the plant genotypes originated from populations that had been exposed to ambient herbivory for 12 years (H+), and the rest were from populations where herbivory was excluded by insecticide spraying for that time period (H−). Thus, we were able to explore the response to selection directly by comparing plants from the two origins that had diverged primarily due to differential survival and clonal reproduction. Receiver plants from populations that had experienced ambient herbivory (H+) generally induced resistance in response to VOC cues from both damaged plants of the same genotypes (self) and different genotypes (non-self) equally. In contrast, receivers from populations where insect herbivory had been excluded (H−) only responded to VOC cues from emitters of the same genotype (self) by inducing resistance to subsequent herbivory ( 16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. 23 Lynch M. The similarity index and DNA fingerprinting. 1,34 = 0.88; p = 0.354), and there was no genetic structure between the populations (PCoA; However, in S. altissima, the effect of relatedness on VOC-mediated plant-plant information transfer depended on the plant population’s history of exposure to herbivory (origin), as evidenced by a three-way interaction between damage treatment, relatedness, and plant origin (Χ= 4.05; df = 1; p = 0.044). In our field experiment, approximately half of the plant genotypes originated from populations that had been exposed to ambient herbivory for 12 years (H+), and the rest were from populations where herbivory was excluded by insecticide spraying for that time period (H−). Thus, we were able to explore the response to selection directly by comparing plants from the two origins that had diverged primarily due to differential survival and clonal reproduction. Receiver plants from populations that had experienced ambient herbivory (H+) generally induced resistance in response to VOC cues from both damaged plants of the same genotypes (self) and different genotypes (non-self) equally. In contrast, receivers from populations where insect herbivory had been excluded (H−) only responded to VOC cues from emitters of the same genotype (self) by inducing resistance to subsequent herbivory ( Figure 2 ). Such differences in among-genotype communication could arise from differences in relatedness within a population []. However, we found no evidence for that, as the mean genetic distance (Lynch) [] between non-self pairs of emitters and receivers did not differ between H− and H+ populations (F= 0.88; p = 0.354), and there was no genetic structure between the populations (PCoA; Figure S1 ). Thus, our results are more likely explained by differential selection on information transfer between plants from H+ and H− populations. Importantly, these results identify insect herbivory as an important driver of the evolution of plant-plant communication, because sharing information about oncoming herbivores (open channel of communication) seems to be favored when S. altissima is exposed to an environment with high risk of herbivory.

21 Morrell K.

Kessler A. Plant communication in a widespread goldenrod: keeping herbivores on the move. 22 Karban R.

Wetzel W.C.

Shiojiri K.

Ishizaki S.

Ramirez S.R.

Blande J.D. Deciphering the language of plant communication: volatile chemotypes of sagebrush. The design of the field experiment, which excluded root and tactile information transfer (all plants were in pots and emitters covered with a mesh bag), suggested a major role of VOC cues in mediating the differential responses to self versus non-self neighbors by plants from H+ and H− populations. Moreover, previous work had established that the VOC cues from damaged plants alone are sufficient to induce metabolic changes in the neighbor and so mediate increased resistance to herbivores []. This, however, raises the question of what attributes of the VOC blends may mediate such specific differences between H+ and H− populations? A relatively open communication channel observed in H+ populations may result if genotypes within the population emit and respond to similar VOC blends such that VOCs from self and non-self are indistinguishable. In contrast, closed (private) communication channels should then be characterized by genotype-specific and distinct VOC induction patterns []. Furthermore, the receivers in H+ populations can be predicted to be under increased selection to more efficiently eavesdrop on their neighbors’ chemical cues. Accordingly, the open communication in H+ populations may simply be due to selection on improved perception of VOC emission in the receivers, in which case we would not necessarily see signal convergence of the emitters despite the strong plant-plant effect in non-self receivers ( Figure 2 ).

1,33 = 3.33; p = 0.022), but plant origin did not affect induction of sesquiterpene emissions in response to herbivory per se (damage treatment × origin F 1,33 = 0.51, p = 0.822; Figure 3 Induction of Sesquiterpene Blends Show full caption (A) NMDS of ordination of 41 sesquiterpene compounds identified from the VOC blend of S. altissima. Number of replicates/genotypes in each treatment was as follows: H+C = 17/10, H+D = 17/10 and H–C = 12/8, H–D = 13/9. We used means for genotypes with two replicates for the analyses to avoid pseudoreplication. Damage by T. virgata caused sesquiterpene emission in the damaged H+ plants (H+D) to converge (ordiareatest with 999 permutations; p = 0.010), but not in the damaged H− plants (H–D; p = 0.483) or control plants from either origin (H+C, p = 0.815; H–C, p = 0.677). Points are genotype means, and ellipses outline the SEM for the four groups. Dashed line, control; solid line, damage treatment. (B) Homogeneity of multidimensional group dispersion of the sesquiterpene composition among genotypes between damage and control plants tended to be lower in plants from H+ populations (betadisper; F1,18 = 4.083; p = 0.058), but not in plants from H− population (F1,15 = 0.009; p = 0.926). In other words, damaged plants from H+ tended to be more similar to one another than plants in other treatment × origin combinations. The plot represents the median distance to the cetroid with the upper and lower quartiles (box) and the minimum and maximum values within 1.5 times of the box (whiskers). See also Table S2 To test for variation in induced VOC emission by plant origin, we collected VOCs under damage and control treatments in a common garden from H+ and H− plants ( Figure 3 ). Sesquiterpene composition of the VOC blend changed upon damage (PERMANOVA; damage treatment F= 3.33; p = 0.022), but plant origin did not affect induction of sesquiterpene emissions in response to herbivory per se (damage treatment × origin F= 0.51, p = 0.822; Figure 2 Table S2 ). Although neither damage treatment nor plant origin affected the VOC blend overall, composition and abundance of the sesquiterpene bouquet (the most dominant compound class in S. altissima) varied with both ( Table S2 ). This indicates that, although subtle, the composition of herbivore-induced VOC emission (specifically terpenoids) changes in different ways in plants from H− and H+ populations. Most interestingly, damage-induced sesquiterpene blends were less variable among H+ genotypes than expected when assuming randomness (ordiareatest: p = 0.010; Figure 3 ), whereas this was not the case among damaged H− genotypes (p = 0.483).

24 Babushok V.I.

Linstrom P.J.

Zenkevich I.G. Retention indices for frequently reported compounds of plant essential oils. 14 Kessler A.

Kalske A. Plant secondary metabolite diversity and species interactions. 25 Zebelo S.A.

Matsui K.

Ozawa R.

Maffei M.E. Plasma membrane potential depolarization and cytosolic calcium flux are early events involved in tomato (Solanum lycopersicon) plant-to-plant communication. 26 Nagashima A.

Higaki T.

Koeduka T.

Ishigami K.

Hosokawa S.

Watanabe H.

Matsui K.

Hasezawa S.

Touhara K. Transcriptional regulators involved in responses to volatile organic compounds in plants. 16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. 22 Karban R.

Wetzel W.C.

Shiojiri K.

Ishizaki S.

Ramirez S.R.

Blande J.D. Deciphering the language of plant communication: volatile chemotypes of sagebrush. Figure 4 Response Patterns of Five Sesquiterpene Compounds Commonly Induced in H+ Plants Show full caption Compounds were classified characteristic of induction by random forest. We tentatively identified compounds by comparing their mass spectra to the NIST library and by comparison of retention times to retention indices as (A) β-Bourbonene, (B) β-elemene, (C) epi-bicyclosesquiphellandrene, (D) alloaromadendrene, and (E) germacrene D. Bars represent means, and error bars are SEM. We next investigated which compounds drove the similarity in the induced sesquiterpene blend of genotypes of H+ origin and compared the results to those of the genotypes of H− origin. The induction in H+ genotypes was mostly characterized by increased emissions of five compounds (Boruta and random forest; tentatively identified by comparison of mass spectra with NIST library and retention indices [] as β-Bourbonene, β-elemene, epi-bicyclosesquiphellandrene, alloaromadendrene, and germacrene D; Figure 4 ). In contrast, H− genotypes varied in their induction pattern, and the analysis did not identify any compounds similarly induced across all genotypes. Both the clustering of genotypes in the induced state in H+, but not in H−, population in the non-metric multidimensional scaling (NMDS) ( Figure 3 ) and the identification of compounds indicating induction only in the H+ population suggest that an environment with a high risk of herbivory favored genotypes that shared a similar VOC induction pattern, whereas the environment without herbivory did not. Taken together, the results are indicative of an open information channel [] and support the two major predictions of the mutual-benefit hypothesis: (1) a convergence on a shared signal and (2) communication among non-self individuals in the H+ populations. Whether or not receivers are also under selection to more efficiently eavesdrop on their neighbors’ chemical information remains unresolved, mainly because of our current lack of measurable traits that can serve as a proxy for the plants’ VOC perception mechanism []. In contrast to these findings in H+ populations, the exclusive response to self and the genotype-specific VOC induction patterns in plants from H− populations are indicative of private channel communication, similar to that observed in sagebrush (Artemisia tridentata) [], and thus support the kin selection hypothesis.

27 Junker R.R.

Kuppler J.

Amo L.

Blande J.D.

Borges R.M.

van Dam N.M.

Dicke M.

Dötterl S.

Ehlers B.K.

Etl F.

et al. Covariation and phenotypic integration in chemical communication displays: biosynthetic constraints and eco-evolutionary implications. 28 Dutour M.

Léna J.-P.

Lengagne T. Mobbing calls: a signal transcending species boundaries. 28 Dutour M.

Léna J.-P.

Lengagne T. Mobbing calls: a signal transcending species boundaries. 29 Wheatcroft D.

Price T.D. Learning and signal copying facilitate communication among bird species. 30 Fallow P.M.

Gardner J.L.

Magrath R.D. Sound familiar? Acoustic similarity provokes responses to unfamiliar heterospecific alarm calls. 21 Morrell K.

Kessler A. Plant communication in a widespread goldenrod: keeping herbivores on the move. 31 Bode R.F.

Kessler A. Herbivore pressure on goldenrod (Solidago altissima L., Asteraceae): its effects on herbivore resistance and vegetative reproduction. Our results suggest that insect herbivory can drive the evolution of open-communication channels, where plants broadcast the presence of herbivores by a convergent, inducible VOC emission that any conspecific neighbor can respond to by inducing resistance. Hence, in S. altissima populations under continued insect herbivore pressure, we observe chemical communication displays in which the proportional compositions of the herbivory-induced VOC bouquets are conserved across individuals (e.g., high phenotypic integration). High phenotypic integration, in turn, has been linked to higher ecological functionality []. Such convergence on common anti-predator signaling has previously been found in animal acoustic communication for warning and mobbing calls for predators []. However, it is still hotly debated whether signal convergence in animals is innate or based on learning []. Our results suggest that, in S. altissima, evolutionary change in patterns of VOC induction likely underlie the observed differences in communication strategy. VOC induction patterns are mechanistically and functionally linked to overall plant metabolism []. Thus, convergence in induced VOC emission blends in H+ populations may be indirectly favored through their metabolic and functional links to other traits, such as direct defense chemicals [] and endogenous plant signaling in general.

25 Zebelo S.A.

Matsui K.

Ozawa R.

Maffei M.E. Plasma membrane potential depolarization and cytosolic calcium flux are early events involved in tomato (Solanum lycopersicon) plant-to-plant communication. 26 Nagashima A.

Higaki T.

Koeduka T.

Ishigami K.

Hosokawa S.

Watanabe H.

Matsui K.

Hasezawa S.

Touhara K. Transcriptional regulators involved in responses to volatile organic compounds in plants. Although our data demonstrate VOC emission convergence on the emitter side, receivers in environments selecting for enhanced plant communication should also be expected to be fine-tuned on their ability to respond to those specific cues. Unfortunately, the lack of knowledge of how plants perceive VOCs currently limits our ability to measure associated receiver traits. New findings suggest that VOCs may directly interact with cell membranes, which, through changes in cell membrane potentials, could induce endogenous signal transduction cascades [] or enter the cell and bind directly to nuclear proteins that act as co-repressors of stress-responsive genes []. Studies like this, as well as the assessment of genotypic variation in VOC perception, will help to identify measurable traits associated with plant VOC perception in the future.

13 Heil M.

Karban R. Explaining evolution of plant communication by airborne signals. 32 Freundlich G.E.

Frost C.J. Variable costs and benefits of eavesdropping a green leaf volatile on two plant species in a common garden. 31 Bode R.F.

Kessler A. Herbivore pressure on goldenrod (Solidago altissima L., Asteraceae): its effects on herbivore resistance and vegetative reproduction. 33 Uesugi A.

Kessler A. Herbivore exclusion drives the evolution of plant competitiveness via increased allelopathy. 34 Uesugi A.

Connallon T.

Kessler A.

Monro K. Relaxation of herbivore-mediated selection drives the evolution of genetic covariances between plant competitive and defense traits. 18 Heil M. Herbivore-induced plant volatiles: targets, perception and unanswered questions. 35 Ameye M.

Allmann S.

Verwaeren J.

Smagghe G.

Haesaert G.

Schuurink R.C.

Audenaert K. Green leaf volatile production by plants: a meta-analysis. 36 Picq S.

Alda F.

Bermingham E.

Krahe R. Drift-driven evolution of electric signals in a neotropical knifefish. 37 Irwin D.E.

Thimgan M.P.

Irwin J.H. Call divergence is correlated with geographic and genetic distance in greenish warblers (Phylloscopus trochiloides): a strong role for stochasticity in signal evolution?. 38 Campbell P.

Pasch B.

Pino J.L.

Crino O.L.

Phillips M.

Phelps S.M. Geographic variation in the songs of neotropical singing mice: testing the relative importance of drift and local adaptation. An important remaining question is why plants from herbivore exclusion plots had retained their ability to transmit information at all, albeit only to themselves? After all, the expression of traits associated with sharing and perceiving VOC-mediated information can come with substantial metabolic and ecological costs, not only from eavesdropping neighbors []. Relief from herbivory in S. altissima populations may merely shift the contribution of different selective agents, increasing the relative importance of competition. Previous studies have demonstrated that excluding herbivores selects for genotypes with reduced resistance to herbivores but higher competitive ability [] and even drives the evolution of genetic covariance between defenses and competition []. Given the multifunctional nature of induced VOC emissions [], it is safe to assume that, with the changes in the selective environment, the major function of an information-mediating trait, such as VOC emission, will shift. Furthermore, plants alter the release of VOCs in response to pathogens or abiotic stress [], which were not altered in our selection experiment. It is possible that selectively sharing information within a clone about these stressors served a competitive advantage in the H− populations, which would have favored the observed private channel communication. Alternatively, the breakdown of open communication and divergence in VOCs in the absence of selection by herbivores can be simply the result of genetic drift. In acoustic and electric communication systems, drift has proven to be an important cause of signal diversification []. With selection by herbivores removed, the relative effect of drift may have increased causing the divergence in inducible VOC emission patterns.

16 Karban R.

Shiojiri K.

Ishizaki S.

Wetzel W.C.

Evans R.Y. Kin recognition affects plant communication and defence. Some factors that affect the applicability of our results in other plant and animal communication systems should be acknowledged. Kin selection is likely to be stronger in species with a clonal growth form, such as S. altissima, in which kin selection and selection for within-plant signaling likely act in the same direction. Nevertheless, kin selection can also favor plant-plant communication between individuals of intermediate relatedness []. In contrast, the mutual benefit hypothesis does not require individuals to be related and the respective mechanisms, such as open channel information sharing and risk spreading, should in theory function in clonal and non-clonal systems alike. Thereby, the way receivers respond to the information and the strength and mode of response of herbivores or other antagonists are more important factors for the evolution of plant-plant communication than relatedness within a population.