Infestation of Rice Striped Stem Borer (Chilo suppressalis) Larvae Induces Emission of Volatile Organic Compounds in Rice and Repels Female Adult Oviposition

Abstract

Plants regulate the biosynthesis and emission of metabolic compounds to manage herbivorous stresses. In this study, as a destructive pest, the pre-infestation of rice striped stem borer (SSB, Chilo suppressalis) larvae on rice (Oryza sativa) reduced the subsequent SSB female adult oviposition preference. Widely targeted volatilomics and transcriptome sequencing were used to identify released volatile metabolic profiles and differentially expressed genes in SSB-infested and uninfested rice plants. SSB infestation significantly altered the accumulation of 71 volatile organic compounds (VOCs), including 13 terpenoids. A total of 7897 significantly differentially expressed genes were identified, and genes involved in the terpenoid and phenylpropanoid metabolic pathways were highly enriched. Correlation analysis revealed that DEGs in terpenoid metabolism-related pathways were likely involved in the regulation of VOC biosynthesis in SSB-infested rice plants. Furthermore, two terpenoids, (-)-carvone and cedrol, were selected to analyse the behaviour of SSB and predators. Y-tube olfactometer tests demonstrated that both (-)-carvone and cedrol could repel SSB adults at higher concentrations; (-)-carvone could simultaneously attract the natural enemies of SSB, Cotesia chilonis and Trichogramma japonicum, and cedrol could only attract T. japonicum at lower concentrations. These findings provide a better understanding of the response of rice plants to SSB and contribute to the development of new strategies to control herbivorous pests.

Keywords: plant–insect interactions; rice; rice striped stem borer; transcriptome; volatilomics.

Figure 1

Oviposition preference of SSB in different treated rice plants. (A) A scheme of the oviposition experiments; (B) Number of eggs laid by SSB female adults in uninfested and SSB pre-infested rice plants; (C) Numbers of egg masses by SSB female adults in uninfested and SSB pre-infested rice plants. The experiment was continued for 72 h and repeated 15 times. Statistical significance was calculated using SPSS. Each bar represents the mean ± SE. Data analysed using GLMs with Wald χ2 statistics indicate the overall difference between uninfested rice and SSB pre-infested rice plants. *** p < 0.001 indicate significant differences between comparison groups.

Figure 2

Volatile organic compounds analysis in rice plants. (A) Classification and proportion of 650 VOCs detected in rice plants. (B) Principal component analysis (PCA) among samples of rice plants by HS-SPME-GC-MS in different groups; the X-axis and Y-axis represent the first and second principal components, respectively.

Figure 3

Overall analysis of VOC changes in rice plants’ response to SSB infestation. (A) Hierarchical cluster analysis of differentially accumulated VOCs in three treatment groups (SSB_24 h, SSB_48 h, Control); each group contained three biological replicates. (B) A histogram with VOCs in SSB-infested rice plants compared with the control group. (C) Venn of VOCs in two comparison groups.

Figure 4

K-means plot and KEGG pathways enrichment of VOCs metabolome in SSB-infested rice compared with the control group. (A) VOCs from different samples collected from control, 24 h and 48 h after treatment were used for the K-means plot. KEGG pathways enrichment of VOCs metabolome in SSB_24 h vs. Control (B) and SSB_48 h vs. Control comparison groups (C).

Figure 5

Overall analysis of differentially expressed genes changes in rice plant response to SSB infestation. (A) Principal component analysis of each transcriptome sample; X-axis, Y-axis and Z-axis represent the first, second and third principal components, respectively. (B) A histogram with DEGs in plants infested by SSB compared with control group. (C) Venn of DEGs in two comparison groups. (D) Hierarchical cluster analysis of DEGs in the three groups of SSB-infested; each group contained three biological replicates. (E,F) Bubble plots with KEGG pathways enriched for DEGs in rice infested by SSB at the 24 h and 48 h time-point compared with the control group.

Figure 6

Analysis of terpenoid biosynthesis and differences between the three treatment groups in rice and GC-MS validation. Note: (A) Key structural genes and their expression level involved in terpenoid backbone biosynthesis pathway in rice in treatment groups. (B) Expression levels of differential metabolites in monoterpenoid biosynthesis pathway. DEV represents differentially expressed volatiles; DEG represents differentially expressed genes; red represents high expression levels; green and blue represent low expression levels in volatiles and transcript, respectively. (C) Relative abundance of (−)-carvone compared with SSB_induced to Control group in GC-MS. (The values represent the mean percentages ± SE of the peak area relative to the peak area of the internal standard). (D) Relative abundance of cedrol compared with SSB_induced to Control group in GC-MS. (The values represent the mean percentages ± SE of the peak area relative to the peak area of the internal standard).

Figure 6

Analysis of terpenoid biosynthesis and differences between the three treatment groups in rice and GC-MS validation. Note: (A) Key structural genes and their expression level involved in terpenoid backbone biosynthesis pathway in rice in treatment groups. (B) Expression levels of differential metabolites in monoterpenoid biosynthesis pathway. DEV represents differentially expressed volatiles; DEG represents differentially expressed genes; red represents high expression levels; green and blue represent low expression levels in volatiles and transcript, respectively. (C) Relative abundance of (−)-carvone compared with SSB_induced to Control group in GC-MS. (The values represent the mean percentages ± SE of the peak area relative to the peak area of the internal standard). (D) Relative abundance of cedrol compared with SSB_induced to Control group in GC-MS. (The values represent the mean percentages ± SE of the peak area relative to the peak area of the internal standard).

Figure 7

Behaviour response to significant chemical compounds in monoterpenoid biosynthesis pathway to SSB; natural enemy Cotesia chilonis, Trichogramma japonicum. (A–C) show preference for SSB, C. chilonis, and T. japonicum behaviour response to chemical compounds (−)-carvone, respectively. (D–F) show the preference of SSB, C. chilonis, and T. japonicum behaviour response to chemical compounds cedrol, respectively.