Systemic responses in a tolerant olive (Olea europaea L.) cultivar upon root colonization by the vascular pathogen Verticillium dahliae

Abstract

Verticillium wilt of olive (VWO) is caused by the vascular pathogen Verticillium dahliae. One of the best VWO management measures is the use of tolerant cultivars; however, our knowledge on VWO tolerance/resistance genetics is very limited. A transcriptomic analysis was conducted to (i) identify systemic defense responses induced/repressed in aerial tissues of the tolerant cultivar Frantoio upon root colonization by V. dahliae, and (ii) determine the expression pattern of selected defense genes in olive cultivars showing differential susceptibility to VWO. Two suppression subtractive hybridization cDNA libraries, enriched in up-regulated (FU) and down-regulated (FD) genes respectively, were generated from "Frantoio" aerial tissues. Results showed that broad systemic transcriptomic changes are taking place during V. dahliae-"Frantoio" interaction. A total of 585 FU and 381 FD unigenes were identified, many of them involved in defense response to (a)biotic stresses. Selected genes were then used to validate libraries and evaluate their temporal expression pattern in "Frantoio." Four defense genes were analyzed in cultivars Changlot Real (tolerant) and Picual (susceptible). An association between GRAS1 and DRR2 gene expression patterns and susceptibility to VWO was observed, suggesting that these transcripts could be further evaluated as markers of the tolerance level of olive cultivars to V. dahliae.

Keywords: Olea europaea; defoliating pathotype; susceptibility; systemic defense responses; tolerance; vascular pathogen; verticillium wilt.

Figure 1

“Level 3” Gene Ontology (GO) “Biological processes” (BP) and “Molecular function” (MF) terms distribution of 381 unigenes repressed (FD) and 585 induced (FU) in olive (Olea europaea L.) aerial tissues colonized by Verticillium dahliae. Unigenes were categorized using the “Blast2GO” software.

Figure 2

Relative expression (RE) average of seven genes identified in FD and/or FU cDNA libraries from aerial tissues of “Frantoio” olive plants at different time points after Verticillium dahliae inoculation in roots. ACO, 1-aminocyclopropane-1-carboxylate oxidase; DRR2, disease resistant response protein; ACL, acetone cyanohydrin lyase; GRAS1, transcription factor GRAS1; CO-MT, caffeoyl-o-methyltransferase; PR10, pathogenesis-related protein 10; and DEF, defensin protein 1. Error bars represent the SD from at least two independent qRT-PCR experiments. RE values (log2-fold-change values) were calculated according to the 2^−ΔΔCt method (Livak and Schmittgen, 2001).

Figure 3

Relative expression (RE) average of four genes from aerial tissues of inoculated tolerant (“Frantoio” and “Changlot Real”) and susceptible (“Picual”) olive cultivars at two different time points after Verticillium dahliae inoculation in roots. ACO, 1-aminocyclopropane-1-carboxylate oxidase; DRR2, disease resistant response protein; ACL, acetone cyanohydrin lyase; GRAS1, transcription factor GRAS1; Relative expression values (log2-fold-change values) were calculated according to the 2^−ΔΔCt method (Livak and Schmittgen, 2001). Red and blue lines show RE scored for two different plants sampled at the indicated times after inoculation with the pathogen.