Volatile Dimethyl Disulfide from Guava Plants Regulate Developmental Performance of Asian Citrus Psyllid through Activation of Defense Responses in Neighboring Orange Plants

Abstract

Intercropping with guava (Psidium guajava L.) can assist with the management of Asian citrus psyllid (ACP, Diaphorina citri Kuwayama), the insect vector of the huanglongbing pathogen, in citrus orchards. Sulfur volatiles have a repellent activity and physiological effects, as well as being important components of guava volatiles. In this study, we tested whether the sulfur volatiles emitted by guava plants play a role in plant-plant communications and trigger anti-herbivore activities against ACP in sweet orange plants (Citrus sinensis L. Osbeck). Real-time determination using a proton-transfer-reaction mass spectrometer (PTR-MS) showed that guava plants continuously release methanethiol, dimethyl sulfide (DMS), and dimethyl disulfide (DMDS), and the contents increased rapidly after mechanical damage. The exposure of orange plants to DMDS resulted in the suppression of the developmental performance of ACP. The differential elevation of salicylic acid (SA) levels; the expression of phenylalanine ammonia lyase (PAL), salicylate-O-methyl transferase (SMT), and pathogenesis-related (PR1) genes; the activities of defense-related enzymes PAL, polyphenol oxidase (PPO), and peroxidase (POD); and the total polyphenol content were observed in DMDS-exposed orange plants. The emission of volatiles including myrcene, nonanal, decanal, and methyl salicylate (MeSA) was increased. In addition, phenylpropanoid and flavonoid biosynthesis, and aromatic amino acid (such as phenylalanine, tyrosine, and tryptophan) metabolic pathways were induced. Altogether, our results indicated that DMDS from guava plants can activate defense responses in eavesdropping orange plants and boost their herbivore resistance to ACP, which suggests the possibility of using DMDS as a novel approach for the management of ACP in citrus orchards.

Keywords: Asian citrus psyllid; defense response; dimethyl disulfide; eavesdropping; guava; sulfur volatiles; sweet orange.

Figure 1

Determination of sulfur volatiles from guava plants with or without mechanical damage with a proton-transfer-reaction mass spectrometer (PTR-MS). (a) Real-time determination of sulfur volatiles from guava plants. (b–d) The concentration of methanethiol, dimethyl sulfide (DMS), and dimethyl disulfide (DMDS) at each time point. Data are shown as means ± SE (n = 6). Asterisks indicate significant differences between mechanical damaged and control (undamaged) plants based on Student’s t-test (* p < 0.05, ** p < 0.01, and *** p < 0.001); ns, no significant difference.

Figure 2

Exposure of sweet orange plants to sulfur volatiles affected the host selection behavior and population development of Asian citrus psyllid (ACP). (a) The number of settled ACP adults at 24 h after their release in wind tunnels. (b) The number of eggs on each plant after ACP oviposition for 48 h. (c) The percentage of nymphal instars at 10 d after ACP release for oviposition. (d) The number of total emerged adults (columns) and the rate of survivorship (dotted line) from egg to adult. Data are shown as means ± SE (n = 5). Different letters on the columns indicate significant differences among the different treatments based on Tukey’s test (p < 0.05).

Figure 3

The levels of jasmonic acid (JA) and salicylic acid (SA) in orange plants after exposure to sulfur volatiles. (a) JA content; (b) SA content. Data are shown as means ± SE (n = 5). Different letters on the columns indicate significant differences among different treatments based on Tukey’s test (p < 0.05).

Figure 4

The expression of phenylalanine ammonia lyase (PAL), salicylate-O-methyl transferase (SMT), and pathogenesis-related (PR1) genes of orange plants after exposure to sulfur volatiles. Data are shown as means ± SE (n = 3). Asterisks indicate significant differences between exposed and control plants, based on Student’s t-test (* p < 0.05 and ** p < 0.01); ns, no significant difference.

Figure 5

Phenylalanine ammonia lyase (PAL), polyphenol oxidase (PPO), and peroxidase (POD) activities and total phenol content of orange plants after exposure to dimethyl disulfide (DMDS). (a–c) PAL, PPO, and POD activities, respectively; (d) total phenol content. Data are shown as means ± SE (n = 3). Different letters on the columns indicate significant differences among different treatments based on Tukey’s test (p < 0.05).

Figure 6

Volatile profiling of orange plants after exposure to dimethyl disulfide (DMDS). (a) PLS-DA (score plot); and (b) clustering analysis and heat map (average) of orange volatile metabolites. (c) Emission of orange volatiles from different classifications. Data are shown as means ± SE (n = 6). Different letters on the columns indicate significant differences among different treatments based on Tukey’s test (p < 0.05).

Figure 7

Characterization of the differentially expressed semipolar metabolites (DEMs) of orange plants after exposure to dimethyl disulfide (DMDS). (a) PLS-DA (score plot); and (b) clustering analysis and heat map of expression measures of semipolar metabolites. (c) The number of DEMs in different groups (for details, see Supplementary Table S2). (d) Venn diagram showing the distribution and overlap of up- and downregulated DEMs across all treatments. (e) KEGG pathway enrichment of DEMs.