Volatile DMNT systemically induces jasmonate-independent direct anti-herbivore defense in leaves of sweet potato (Ipomoea batatas) plants

Abstract

Plants perceive and respond to volatile signals in their environment. Herbivore-infested plants release volatile organic compounds (VOCs) which can initiate systemic defense reactions within the plant and contribute to plant-plant communication. Here, for Ipomoea batatas (sweet potato) leaves we show that among various herbivory-induced plant volatiles, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) had the highest abundance of all emitted compounds. This homoterpene was found being sufficient for a volatile-mediated systemic induction of defensive Sporamin protease inhibitor activity in neighboring sweet potato plants. The systemic induction is jasmonate independent and does not need any priming-related challenge. Induced emission and responsiveness to DMNT is restricted to a herbivory-resistant cultivar (Tainong 57), while a susceptible cultivar, Tainong 66, neither emitted amounts comparable to Tainong 57, nor showed reaction to DMNT. This is consistent with the finding that Spodoptera larvae feeding on DMNT-exposed cultivars gain significantly less weight on Tainong 57 compared to Tainong 66. Our results indicate a highly specific, single volatile-mediated plant-plant communication in sweet potato.

Figure 1

Local but no systemic increase of jasmonates after wounding in Ipomoea batatas. (a–j) Jasmonate levels after mechanical wounding by MecW (a,c,e,g,i; n = 10–11) and insect feeding by S. littoralis (b,d,f,h,j; n = 7–12) measured in I. batatas TN57 after 0.5 h, 1 h and 3 h. Phytohormone levels were measured in locally wounded leaves (dark gray bars) and the adjacent unwounded systemic leaf (light gray bars). Leaves from undamaged plants served as controls (black bars). Statistically significant differences between each treatment group after treatment were analyzed for each time point separately using one–way ANOVA. Different letters indicate significant differences among groups for p < 0.05, determined by Tukey’s test. In (a–j), data are presented as mean ± SEM.

Figure 2

Volatile emission and upregulation of (E)-4,8–dimethyl–nonatriene (DMNT) in Ipomoea batatas. (a,b) Gas chromatograms of volatiles emitted by I. batatas TN57 (a) and TN66 (b): Controls without wounding; volatiles induced by mechanical damage (MecW) inflicted over 18 h; volatiles induced by feeding of S. littoralis. All volatiles were collected over 24 h and eluted with internal standard. Asterisks mark contamination by plasticizer or column residuals. Identified compounds are marked as follows: (1) α-Pinene; (2) 1-Butoxy-2-propanol; (3) 2-Ethylhexanal; (4) Benzaldehyde; (5) 5-Ethyl-(5 H)-furan-2-one; (6) 6-Methyl-5-hepten-2-one; (7) Mesitylene; (8) 1-Decene; (9) n-Decane; (10) n-Octanal; (11) (Z)-Hex-3-enyl acetate; (12) Hexyl acetate; (13) (E)-Hex-2-enyl acetate; (14) Limonene; (15) 2-Ethyl-hexanol; (16) (E)-β-Ocimene; (17) unidentified monoterpenoid (93, 136); (18) n-Nonanal; (19) 4,8-Dimethylnona-1,3,7-triene; (20) Phenyl acetonitrile; (21) Naphthalene; (22) (Z)-Hex-3-enyl butanoate; (23) n-Decanal; (24) Indole; (25) n-Tridecane; (26) n-Undecanal; (27) Internal standard (n-bromodecane); (28) (E)-2-Undecenal; (29) α-Copaene; (30) β-Cubebene; (31) 7-epi-Sesquithujene; (32) 1-Tetradecene; (33) (Z)-Jasmone; (34) n-Tetradecane; (35) Dodecanal; (36) (E)-β-Caryophyllene; (37) β-Copaene; (38) (E)-α-Bergamotene; (39) Sesquisabinene; (40) α-Humulene; (41) Geranyl acetone; (42) Germacrene D; (43) β-Ionone; (44) Bicyclogermacrene; (45) n-Pentadecane; (46) Tridecanal; (47) Nerolidol; (48) (3E,7E)-4,8,12-Trimethyltrideca-1,3,7,11-tetraene; (49) n-Hexadecane; (50) n-Heptadecane; (51) n-Pentadecanal; (52) n-Octadecane; (53) Isopropyl tetradecanoate; (54) n-Hexadecanol. Identification of compounds is shown in Supplementary Table S1. (c,d) DMNT emission after mechanical damage and herbivore feeding in I. batatas TN57 (c) and TN66. (d) VOCs were collected over 24 h with 18 h mechanical wounding by MecW (light gray bars; n = 5–8) or 24 h infestation with S. littoralis (dark gray bars; n = 7–12) and the respective control (black bars; n = 5–7). Bars represent the mean ± SEM of emitted DMNT in ng cm⁻² leaf area. Statistically significant differences between each group were analyzed using a Kruskal-Wallis one-way ANOVA on ranks. Different letters indicate significant differences among groups for p < 0.05, determined by Dunn’s test and adjusted p-values according to Benjamini & Hochberg. (c) TN57: p < 0.001. (d) TN66: p = 0.004.

Figure 3

Airborne DMNT increases defense capabilities in Ipomoea batatas TN57. (a,b) Larval weight of S. litura after feeding for 7 d and 10 d on DCM (control) or DMNT- treated TN57 (a, n = 16) or TN66 (b, n = 25) plants. For DMNT treatment, plants were incubated with 3.9 nM for 3 h. (c,d) Trypsin inhibitory activity of TN57 (c, n = 5) and TN66 (d, n = 6) after incubation with 3.9 nM of DMNT for 3 h. Bars represent the mean ± SEM of larval weight or trypsin inhibitory activity for control (DCM, black bars) and DMNT treatment (gray bars). Significance levels are indicated by the asterisks (n.s. = non-significant; *p < 0.05; **p < 0.01). (a) TN57: p (CxDMNT) = 0.002; (b) TN66: p (CxDMNT) = 0.468. (c) TN57: p (CxDMNT) = 0.028; (d) TN66: p (CxDMNT) = 0.207. Asterisks indicate significant differences between control and DMNT treatment, based on a t-test (c,d) and ANOVA followed by a Tukey- adjusted comparison based on a linear model (a,b).

Figure 4

DMNT does not systemically induce jasmonate production. JA and JA-Ile levels of single I. batatas TN57 plants incubated with 1.41 µg of DMNT dissolved in dichloromethane in 2.4 L glass desiccators for 1 h (light gray bar, n = 10). Black bars indicate the control samples treated with dichloromethane (n = 7). S. littoralis feeding (light gray with stripes) was used as a positive control for the visualization of jasmonate induction. Bars represent the mean ± SEM of detected JA and JA-Ile. Significant differences were determined using a Shapiro–Wilk normality test and a subsequent Mann-Whitney rank sum test for the treatment and the respective control (n.s = non-significant); p JA (CxDMNT) = 0.130; p JA-Ile (CxDMNT) = 0.661.

Figure 5

Model of DMNT emission that triggers SPI-dependent resistance enhancement in TN57 after mechanical wounding and Spodoptera herbivory. Upon mechanical wounding (MecWorm) or herbivore (S. littoralis) feeding, jasmonates (JA) are locally upregulated in the treated leaf. Sporamin protease inhibitor (SPI) is upregulated mainly systemically. In parallel, (E)-4,8–dimethyl–nonatriene (DMNT) is emitted to the environment and induces the generation of SPI in leaves of non-treated neighboring I. batatas plants without changes in JA levels. As a consequence, these plants show higher resistance against feeding Spodoptera larvae.