Induced resistance by a long-chain bacterial volatile: elicitation of plant systemic defense by a C13 volatile produced by Paenibacillus polymyxa

Abstract

Background: Some strains of plant growth-promoting rhizobacteria (PGPR) elicit induced systemic resistance (ISR) by emission of volatile organic compounds (VOCs) including short chain alcohols, acetoin, and 2,3-butanediol. The objective of this study was to evaluate whether species-specific VOCs from PGPR strain Paenibacillus polymyxa E681 can promote growth and induce resistance in Arabidopsis.

Methodology/principal findings: The efficacy of induction was strain-specific, with stronger protection against Pseudomonas syringae pv. maculicola ES4326 in plants exposed to VOCs from P. polymyxa E681 versus Arabidopsis plants exposed to VOCs from a reference strain Bacillus subtilis GB03, which was previously shown to elicit ISR and plant growth promotion. VOC emissions released from E681 primed transcriptional expression of the salicylic acid, jasmonic acid, and ethylene signaling marker genes PR1, ChiB, and VSP2, respectively. In addition, strain E681 produced more than thirty low molecular-weight VOCs, of which tridecane was only produced by E681 and not found in GB03 or IN937a volatile blends. These strain-specific VOCs induced PR1 and VSP2 genes.

Conclusions/significance: These results provide new insight into the existence of a long chain VOC signaling molecule produced by P. polymyxa that can serve as a bacterial trigger of induced systemic resistance in planta.

Figure 1. Growth promotion of Arabidopsis seedling by VOCs emitted from Paenibacillus polymyxa E681.

A) Illustration of plant growth promotion by VOCs produced by strain E681 in a 24-well microtitre system. The diameter of each well was 2 cm. The photograph was taken two weeks after inoculation with strain E681. Right plate = water control; Left plate = E681 treatment. The single arrows indicate inoculation site with strain E681 and water control. B) Plant growth at two weeks after exposure to VOCs released by P. polymyxa E681, Bacillus subtilis GB03, and water control in a microtitre system, as indicated by the differences in the total leaf surface areas. C) Plant growth at two weeks after exposure to VOCs released by P. polymyxa E681, Bacillus subtilis GB03, and water control in a microtitre system, as indicated by the differences in the foliar fresh weight in wild type Col-0 and its hormonal mutant lines, transgenic NahG (encodes salicylate hydroxylase and degrades salicylic acid (SA)), ein2.5 (ET-insensitive), coi1 (insensitive to jasmonic acid), and gai2 (insensitive to gibberellic acid). Different letters indicate significant differences between treatments at 2, 4, and 6 cm away from bacteria inoculation in the microtitre system, according to least significant difference at P = 0.05.

Figure 2. Induction of resistance against Pseudomonas syringae pv. maculicola ES4326 in Arabidopsis exposed to bacterial VOCs.

Induced systemic resistance against P. syringae pv. maculicola ES4326 elicited by VOCs of P. polymyxa E681 and a water control, using the microtitre system. Disease severity (0 = no symptom, 10 = severe chlorosis) was recorded seven days after pathogen challenge. Different letters indicate significant differences between treatments, according to least significant difference at P = 0.05. The error bars indicate SEM.

Figure 3. Induced systemic resistance and priming of defense-related genes by tridecane in Arabidopsis.

A) Effect of tridecane on Arabidopsis growth. Plants were exposed to 10 mM and 100 µM tridecane, strain E681, and water for 2 weeks in presence (black bar) and absence of Ba(OH)₂ (white bar). The photograph shows effect of CO₂ on Arabidopsis growth treatments with 10 mM and 100 µM tridecane, strain E681, and water by the addition of Ba(OH)₂ for trapping CO₂ that precipitated as BaCO₃. B) Induced systemic resistance against P. syringae pv. maculicola ES4326 elicited by strain E681 and 10 mM and 100 µM tridecane using the I-plate system. Disease severity (0 = no symptom, 5 = severe chlorosis) was recorded seven days after pathogen challenge at 10⁸ cfu/ml. Inset picture indicates chemical structure of tridecane. Gene expression levels of PR1 for salicylic acid signaling (C and F), ChiB for ET signaling (D and G), and VSP2 for jasmonic acid signaling (E and H), as determined by quantitative reverse transcriptase (qRT)-PCR after tridecane emission (C, D, and E) at 0 and 6 hour post-inoculation (hpi) and after pathogen challenge for detecting defense priming (F, G, and H). The expression ratio (C–H), a ratio of the expression in the strain E681 or tridecane-inoculated treatment relative to expression of Actin gene, is shown as the mean ± SEM. Different letters indicate significant differences between treatments (A and B) according to least significant difference at P = 0.05.

Figure 4. Induced resistance and priming of defense-related genes by long chain VOCs.

A)Induced systemic resistance against P. syringae pv. maculicola ES4326 elicited by strain E681 and 10 mM and 100 µM of decane, undecane, and dodecane using the I-plate system. Disease severity (0 = no symptom, 5 = severe chlorosis) was recorded seven days after pathogen challenge at 10⁸ cfu/ml. Gene expression levels of PR1 for salicylic acid signaling (B and E), ChiB for ET signaling (C and F), and VSP2 for jasmonic acid signaling (D and G), as determined by quantitative reverse transcriptase (qRT)-PCR after tridecane emission (B, C, and D) at 0 and 6 hour post-inoculation (hpi) and after pathogen challenge for detecting defense priming (E, F, and G). The expression ratio (B–G), a ratio of the expression in the strain E681 or tridecane treatment relative to water-treated control, is shown as the mean ± SEM. Different letters indicate significant differences between treatments (A) according to least significant difference at P = 0.05.