BackgroundEucalyptus species and interspecific hybrids exhibit valuable growth and wood properties that make them a highly desirable commodity. However, these trees are challenged by a wide array of biotic stresses during their lifetimes. The Eucalyptus grandis reference genome sequence provides a resource to study pest and pathogen defence mechanisms in long-lived woody plants. E. grandis trees are generally susceptible to Chrysoporthe austroafricana, a causal agent of stem cankers on eucalypts. The aim of this study was to characterize the defence response of E. grandis against C. austroafricana.ResultsHormone profiling of susceptible and moderately resistant clonal E. grandis genotypes indicated a reduction in salicylic acid and gibberellic acid levels at 3 days post inoculation. We hypothesized that these signaling pathways may facilitate resistance. To further investigate other defence mechanisms at this time point, transcriptome profiling was performed. This revealed that cell wall modifications and response to oxidative stress form part of the defence responses common to both genotypes, whilst changes in the hormone signaling pathways may contribute to resistance. Additionally the expression of selected candidate defence response genes was induced earlier in moderately resistant trees than in susceptible trees, supporting the hypothesis that a delayed defence response may occur in the susceptible interaction.ConclusionThe ability of a host to fine-tune its defence responses is crucial and the responses identified in this study extends our understanding of plant defence, gained from model systems, to woody perennials.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1529-x) contains supplementary material, which is available to authorized users.

Long-lived tree species are subject to attack by various pests and pathogens during their lifetime. This problem is exacerbated by climate change, which may increase the host range for pathogens and extend the period of infestation by pests. Plant defences may involve preformed barriers or induced resistance mechanisms based on recognition of the invader, complex signalling cascades, hormone signalling, activation of transcription factors and production of pathogenesis-related (PR) proteins with direct antimicrobial or anti-insect activity. Trees have evolved some unique defence mechanisms compared with well-studied model plants, which are mostly herbaceous annuals. The genome sequence of Eucalyptus grandis W. Hill ex Maiden has recently become available and provides a resource to extend our understanding of defence in large woody perennials. This review synthesizes existing knowledge of defence mechanisms in model plants and tree species and features mechanisms that may be important for defence in Eucalyptus, such as anatomical variants and the role of chemicals and proteins. Based on the E. grandis genome sequence, we have identified putative PR proteins based on sequence identity to the previously described plant PR proteins. Putative orthologues for PR-1, PR-2, PR-4, PR-5, PR-6, PR-7, PR-8, PR-9, PR-10, PR-12, PR-14, PR-15 and PR-17 have been identified and compared with their orthologues in Populus trichocarpa Torr. & A. Gray ex Hook and Arabidopsis thaliana (L.) Heynh. The survey of PR genes in Eucalyptus provides a first step in identifying defence gene targets that may be employed for protection of the species in future. Genomic resources available for Eucalyptus are discussed and approaches for improving resistance in these hardwood trees, earmarked as a bioenergy source in future, are considered.

Chrysoporthe austroafricana is a fungal pathogen that causes the development of stem cankers on susceptible Eucalyptus grandis trees. Clones of E. grandis that are partially resistant and highly susceptible have been identified based on the extent of lesion formation on the stem upon inoculation with C. austroafricana. These interactions have been used as a model pathosystem to enhance our understanding of interactions between pathogenic fungi and woody hosts, which may be different to herbaceous hosts. In previous research, transcriptomics of host responses in these two clones to C. austroafricana suggested roles for salicylic acid and gibberellic acid phytohormone signaling in defense. However, it is unclear how the pathogen infiltrates host tissue and which pathogenicity factors facilitate its spread in the two host genotypes. The aim of this study was to investigate these two aspects of the E. grandis–C. austroafricana interaction and to test the hypothesis that the pathogen possesses mechanisms to modulate the tree phytohormone-mediated defenses. Light microscopy showed that the pathogen occurred in most cell types and structures within infected E. grandis stem tissue. Notably, the fungus appeared to spread through the stem by penetrating cell wall pits. In order to understand the molecular interaction between these organisms and predict putative pathogenicity mechanisms of C. austroafricana, fungal gene expression was studied in vitro and in planta. Fungal genes associated with cell wall degradation, carbohydrate metabolism and phytohormone manipulation were expressed in planta by C. austroafricana. These genes could be involved in fungal spread by facilitating cell wall pit degradation and manipulating phytohormone mediated defense in each host environment, respectively. Specifically, the in planta expression of an ent-kaurene oxidase and salicylate hydroxylase in C. austroafricana suggests putative mechanisms by which the pathogen can modulate the phytohormone-mediated defenses of the host. These mechanisms have been reported in herbaceous plant–pathogen interactions, supporting the notion that these aspects of the interaction are similar in a woody species. This study highlights ent-kaurene oxidase and salicylate hydroxylase as candidates for further functional characterization.

Induction of systemic defences to fungal pathogens is well described in model plant systems, but not extensively studied in large woody perennials. Systemic induced defences in long‐lived tree species, such as Eucalyptus grandis, could provide resistance against subsequent biotic challenges. Terpenoids are a class of plant‐specialized metabolites implicated in defence against herbivores and pathogens. The aim of this study was to characterize the systemic induction of terpenoids in E. grandis clones challenged with the fungal pathogen Chrysoporthe austroafricana. Ramets of E. grandis clones previously classified as moderately resistant (TAG5) and susceptible (ZG14) were inoculated with C. austroafricana on the main stem, with mock‐inoculated ramets to serve as controls. Leaf tissue was harvested 3 days post‐inoculation and terpenoid levels were measured using gas chromatography coupled with mass spectrometry. Foliar mRNA abundance profiles of four candidate terpenoid biosynthesis pathway genes were compared in inoculated and mock‐inoculated ramets of each genotype. Monoterpene levels differed between clonal genotypes and p‐cymene was induced systemically in TAG5. Expression profiling of geranyl pyrophosphate synthase (EgrGPPS), farnesyl pyrophosphate synthase (EgrFPPS2), myrcene synthase (EgrTPS084) and β‐caryophyllene synthase (EgrTPS038) show induction of transcripts involved in terpenoid biosynthesis in leaves following pathogen challenge on the main stem. Together these results suggest that resistant and susceptible Eucalyptus genotypes may have different constitutive and induced terpenoid profiles. The significance of the systemic induction of terpenoid levels in TAG5 and terpenoid biosynthesis transcripts in both genotypes in response to C. austroafricana challenge in the stem, and their role in defence against secondary biotic attack, remains to be determined.