Insect egg-induced physiological changes and transcriptional reprogramming leading to gall formation.

Gall-inducing insects and their hosts present some of the most intricate plant-herbivore interactions. Oviposition on the host is often the first cue of future herbivory and events at this early time point can affect later life stages. Many gallers are devastating plant pests, yet little information regarding the plant-insect molecular interplay exists, particularly following egg deposition. We studied the physiological and transcriptional responses of Eucalyptus following oviposition by the gall-inducing wasp, Leptocybe invasa, to explore potential mechanisms governing defence responses and gall development. RNA sequencing and microscopy were used to explore a susceptible Eucalyptus-L. invasa interaction. Infested and control material was compared over time (1-3, 7 and 90 days post oviposition) to examine the transcriptional and morphological changes. Oviposition induces accumulation of reactive oxygen species and phenolics which is reflected in the transcriptome analysis. Gene expression supports phytohormones and 10 transcription factor subfamilies as key regulators. The egg and oviposition fluid stimulate cell division resulting in gall development. Eucalyptus responses to oviposition are apparent within 24 hr. Putative defences include the oxidative burst and barrier reinforcement. However, egg and oviposition fluid stimuli may redirect these responses towards gall development.

A Genome-Wide Association Study for Resistance to the Insect Pest Leptocybe invasa in Eucalyptus grandis Reveals Genomic Regions and Positional Candidate Defense Genes.

The galling insect, Leptocybe invasa, causes significant losses in plantations of various Eucalyptus species and hybrids, threatening its economic viability. We applied a genome-wide association study (GWAS) to identify single-nucleotide polymorphism (SNP) markers associated with resistance to L. invasa. A total of 563 insect-challenged Eucalyptus grandis trees, from 61 half-sib families, were genotyped using the EUChip60K SNP chip, and we identified 15,445 informative SNP markers in the test population. Multi-locus mixed-model (MLMM) analysis identified 35 SNP markers putatively associated with resistance to L. invasa based on four discreet classes of insect damage scores: (0) not infested, (1) infested showing evidence of oviposition but no gall development, (2) infested with galls on leaves, midribs or petioles and (3) stunting and lethal gall formation. MLMM analysis identified three associated genomic regions on chromosomes 3, 7 and 8 jointly explaining 17.6% of the total phenotypic variation. SNP analysis of a validation population of 494 E. grandis trees confirmed seven SNP markers that were also detected in the initial association analysis. Based on transcriptome profiles of resistant and susceptible genotypes from an independent experiment, we identified several putative candidate genes in associated genomic loci including Nucleotide-binding ARC- domain (NB-ARC) and toll-interleukin-1-receptor-Nucleotide binding signal- Leucine rich repeat (TIR-NBS-LRR) genes. Our results suggest that Leptocybe resistance in E. grandis may be influenced by a few large-effect loci in combination with minor effect loci segregating in our test and validation populations.

The Road to Resistance in Forest Trees.

In recent years, forests have been exposed to an unprecedented rise in pests and pathogens. This, coupled with the added challenge of climate change, renders forest plantation stock vulnerable to attack and severely limits productivity. Genotypes resistant to such biotic challenges are desired in plantation forestry to reduce losses. Conventional breeding has been a main avenue to obtain resistant genotypes. More recently, genetic engineering has become a viable approach to develop resistance against pests and pathogens in forest trees. Tree genomic resources have contributed to advancements in both these approaches. Genome-wide association studies and genomic selection in tree populations have accelerated breeding tools while integration of various levels of omics information facilitates the selection of candidate genes for genetic engineering. Furthermore, tree associations with non-pathogenic endophytic and subterranean microbes play a critical role in plant health and may be engineered in forest trees to improve resistance in the future. We look at recent studies in forest trees describing defense mechanisms using such approaches and propose the way forward to developing superior genotypes with enhanced resistance against biotic stress.

Insect Gallers and Their Plant Hosts: From Omics Data to Systems Biology.

Gall-inducing insects are capable of exerting a high level of control over their hosts' cellular machinery to the extent that the plant's development, metabolism, chemistry, and physiology are all altered in favour of the insect. Many gallers are devastating pests in global agriculture and the limited understanding of their relationship with their hosts prevents the development of robust management strategies. Omics technologies are proving to be important tools in elucidating the mechanisms involved in the interaction as they facilitate analysis of plant hosts and insect effectors for which little or no prior knowledge exists. In this review, we examine the mechanisms behind insect gall development using evidence from omics-level approaches. The secretion of effector proteins and induced phytohormonal imbalances are highlighted as likely mechanisms involved in gall development. However, understanding how these components function within the system is far from complete and a number of questions need to be answered before this information can be used in the development of strategies to engineer or breed plants with enhanced resistance.

The Transcriptome and Terpene Profile of Eucalyptus grandis Reveals Mechanisms of Defense Against the Insect Pest, Leptocybe invasa.

Plants have evolved complex defenses that allow them to protect themselves against pests and pathogens. However, there is relatively little information regarding the Eucalyptus defensome. Leptocybe invasa is one of the most damaging pests in global Eucalyptus forestry, and essentially nothing is known regarding the molecular mechanisms governing the interaction between the pest and host. The aim of the study was to investigate changes in the transcriptional landscape and terpene profile of a resistant and susceptible Eucalyptus genotype in an effort to improve our understanding of this interaction. We used RNA-seqencing to investigate transcriptional changes following L. invasa oviposition. Expression levels were validated using real-time quantitative PCR. Terpene profiles were investigated using gas chromatography coupled to mass spectometry on uninfested and oviposited leaves. We found 698 and 1,115 significantly differentially expressed genes from the resistant and susceptible interactions, respectively. Gene Ontology enrichment and Mapman analyses identified putative defense mechanisms including cell wall reinforcement, protease inhibitors, cell cycle suppression and regulatory hormone signaling pathways. There were significant differences in the mono- and sesquiterpene profiles between genotypes and between control and infested material. A model of the interaction between Eucalyptus and L. invasa was proposed from the transcriptomic and chemical data.

Uncovering the defence responses of Eucalyptus to pests and pathogens in the genomics age.

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.