Genetics of myrtle rust resistance in Eucalyptus

© 2018 Dr. Wilson Thau Lym YongMyrtle rust, also known as eucalyptus rust or guava rust, caused by the pathogenic fungus Austropuccinia psidii Beenken (formerly Puccinia psidii Winter), is a major threat to Australia's natural vegetation and to Eucalyptus dependent industries worldwide. However, the presence of specific resistance to this introduced pathogen in Australian myrtaceous species raises intriguing questions relating to the underlying mechanisms of resistance and the evolutionary trajectories resulting in the observed variations of susceptibility. This thesis aims to identify rust resistance mechanisms in two key Eucalyptus species i.e. Eucalyptus globulus Labill. ssp. globulus and Eucalyptus obliqua L'Hér. that predate contemporary contact between the host and pathogen, using phenotypic and molecular analyses. Rust screening was carried out to examine the genetic variation of susceptibility to A. psidii in geographic races (and sub-races) of E. globulus ssp. globulus and regional populations of E. obliqua. Analysis revealed genetic variation between and within geographic populations of both species that will allow natural adaptation to the new disease threat, and divergence between the two species representative of the two most species-rich Eucalyptus subgenera i.e. Symphyomyrtus and Eucalyptus. Detailed observations of the rust infection process by scanning electron microscopy did not indicate any distinct differences in either ontogeny or morphology of A. psidii infection structures between different rust response phenotypes i.e. completely resistant, hypersensitive and highly susceptible in both Eucalyptus species. Further histological examination revealed direct (non-stomatal) penetration of germinating urediniospores through the leaf cuticle in susceptible phenotypes while the infection was stopped at the leaf surface, without host penetration, in completely resistant phenotypes. Non-host/non-pathogen interactions, i.e. the growth of variable length germ tubes in random directions and reduced probability of germ tubes in locating stomata in Eucalyptus and A. psidii may be due to the lack of basic compatibility or specific recognition events between the plant and the pathogen as has been observed in other non-host pathosystems. Foliar terpenes were analysed as possible preformed resistance compounds associated with rust resistance in both Eucalyptus species. While terpene components were found to be significantly associated with susceptibility differences between rust response phenotypes, the components were not consistent between E. globulus and E. obliqua and quite different to the previously identified resistance biomarkers in Eucalyptus hybrids. Less specificity between terpene composition and rust resistance may be due to the complexity of terpene defensive chemistry between Eucalyptus species and the synergistic effects of terpene compounds in combination with other metabolites in producing similar defensive responses against the same pathogen. Finally, a genome wide association study (GWAS) of rust resistance in E. obliqua identified genomic regions including those corresponding to the reported major QTLs, i.e. Ppr1 to Ppr5, for rust resistance in E. grandis and E. globulus from subgenus Symphyomyrtus. Functional annotations of positional candidate genes also revealed some hypothetical proteins related to defences against pathogens such as an enzymatic class of R genes, and genes annotated as nucleotide-binding (NB) proteins, leucine-rich repeat (NB-LRR) domains and transcription factors. The co-location of rust resistance loci between subgenera Eucalyptus and Symphyomyrtus is consistent with the hypothesis that the host and pathogen had an ancient evolutionary connection dating perhaps to 30 million years ago, and the presence of specific recognition to A. psidii in modern Australian Eucalyptus may be due to the retention of ancient R genes. Alternatively, the resistance genes may be effective against and maintained by the current native pathogens but happen to be partly effective against A. psidii between resistant myrtaceous plants across the Australian landmass. The present study contributes to our understanding of the complex Eucalyptus – A. psidii pathosystem and the underlying mechanisms of rust resistance at both a fundamental and applied level for sustainable disease management in Eucalyptus forests and commercial breeding for rust resistance