Analysis of plant and fungal transcripts from resistant and susceptible phenotypes of Leptospermum scoparium challenged by Austropuccinia psidii.

Austropuccinia psidii is the causal pathogen of myrtle rust disease of Myrtaceae. To gain understanding of the initial infection process, gene expression in germinating Austropuccinia psidii urediniospores and in Leptospermum scoparium inoculated leaves were investigated via analyses of RNAseq samples taken 24 and 48 hours post inoculation (hpi). Principal component analyses of transformed transcript count data revealed differential gene expression between the uninoculated L. scoparium control plants that correlated with the three plant leaf resistance phenotypes (immunity, hypersensitive response and susceptibility). Gene expression in the immune resistant plants did not significantly change in response to fungal inoculation, while susceptible plants showed differential expression of genes in response to fungal challenge. A putative disease resistance gene, jg24539.t1, was identified in the L. scoparium hypersensitive response phenotype family. Expression of this gene may be associated with the phenotype and could be important for further understanding the plant hypersensitive response to A. psidii challenge. Differential expression of pathogen genes was found between samples taken 24 and 48 hpi, but there were no significant differences in pathogen gene expression that were associated with the three different plant leaf resistance phenotypes. There was a significant decrease in the abundance of fungal transcripts encoding three putative effectors and a putative carbohydrate-active enzyme between 24 and 48 hpi, suggesting that the encoded proteins are important during the initial phase of infection. These transcripts, or their translated proteins, may be potential targets to impede the early phases of fungal infection by this wide-host range obligate biotrophic basidiomycete.

Both Constitutive and Infection-Responsive Secondary Metabolites Linked to Resistance against Austropuccinia psidii (Myrtle Rust) in Melaleuca quinquenervia.

Austropuccinia psidii is a fungal plant pathogen that infects species within the Myrtaceae, causing the disease myrtle rust. Myrtle rust is causing declines in populations within natural and managed ecosystems and is expected to result in species extinctions. Despite this, variation in response to A. psidii exist within some species, from complete susceptibility to resistance that prevents or limits infection by the pathogen. Untargeted metabolomics using Ultra Performance Liquid Chromatography with Ion Mobility followed by analysis using MetaboAnalyst 3.0, was used to explore the chemical defence profiles of resistant, hypersensitive and susceptible phenotypes within Melaleuca quinquenervia during the early stages of A. psidii infection. We were able to identify three separate pools of secondary metabolites: (i) metabolites classified structurally as flavonoids that were naturally higher in the leaves of resistant individuals prior to infection, (ii) organoheterocyclic and carbohydrate-related metabolites that varied with the level of host resistance post-infection, and (iii) metabolites from the terpenoid pathways that were responsive to disease progression regardless of resistance phenotype suggesting that these play a minimal role in disease resistance during the early stages of colonization of this species. Based on the classes of these secondary metabolites, our results provide an improved understanding of key pathways that could be linked more generally to rust resistance with particular application within Melaleuca.

Resistance of New Zealand Provenance Leptospermum scoparium, Kunzea robusta, Kunzea linearis, and Metrosideros excelsa to Austropuccinia psidii.

Resistance to the pandemic strain of Austropuccinia psidii was identified in New Zealand provenance Leptospermum scoparium, Kunzea robusta, and K. linearis plants. Only 1 Metrosideros excelsa-resistant plant was found (of the 570 tested) and no resistant plants of either Lophomyrtus bullata or L. obcordata were found. Three types of resistance were identified in Leptospermum scoparium. The first two, a putative immune response and a hypersensitive response, are leaf resistance mechanisms found in other myrtaceous species while on the lateral and main stems a putative immune stem resistance was also observed. Both leaf and stem infection were found on K. robusta and K. linearis plants as well as branch tip dieback that developed on almost 50% of the plants. L. scoparium, K. robusta, and K. linearis are the first myrtaceous species where consistent infection of stems has been observed in artificial inoculation trials. This new finding and the first observation of significant branch tip dieback of plants of the two Kunzea spp. resulted in the development of two new myrtle rust disease severity assessment scales. Significant seed family and provenance effects were found in L. scoparium, K. robusta, and K. linearis: some families produced significantly more plants with leaf, stem, and (in Kunzea spp.) branch tip dieback resistance, and provenances provided different percentages of resistant families and plants. The distribution of the disease symptoms on plants from the same seed family, and between plants from different seed families, suggested that the leaf, stem, and branch tip dieback resistances were the result of independent disease resistance mechanisms.

Imminent Extinction of Australian Myrtaceae by Fungal Disease.

Myrtle rust is a fungal disease that has spread rapidly across the globe, arriving in Australia in 2010. The tree species Rhodomyrtus psidioides is nearly extinct in the wild as a result of the disease, leading to potential disruption of ecosystem function. Many other Myrtaceae may also be threatened and unprecedented impacts of the disease are predicted.

Comparison of host susceptibilities to native and exotic pathogens provides evidence for pathogen-imposed selection in forest trees.

The extent to which spatial structuring of host resistance in wild plant populations reflects direct pathogen-imposed selection is a subject of debate. To examine this issue, genetic susceptibilities to an exotic and a coevolved native fungal pathogen were compared using two Australian host tree species. Damage to common host germplasm of Corymbia citriodora ssp. variegata (CCV) and Eucalyptus globulus, caused by recently introduced (Austropuccinia psidii) and native (Quambalaria pitereka and Teratosphaeria sp.) pathogens was evaluated in common-garden experiments. There was significant additive genetic variation within host species for susceptibility to both the exotic and native pathogens. However, susceptibility to A. psidii was not genetically correlated with susceptibility to either native pathogen, providing support for pathogen-specific rather than general mechanisms of resistance. Population differentiation (QST ) for susceptibility to the native pathogens was greater than neutral expectations (molecular FST ), arguing for divergent selection. Coupled with lower native, but not exotic, pathogen susceptibility in host populations from areas climatically more prone to fungal proliferation, these findings suggest that pathogen-imposed selection has contributed directly to a geographic mosaic of host resistance to native pathogens.

Lessons from the Incursion of Myrtle Rust in Australia.

Austropuccinia psidii (myrtle rust) is a globally invasive neotropical rust of the Myrtaceae that came into international prominence following extensive damage to exotic Eucalyptus plantations in Brazil in the 1970s and 1980s. In 2005, myrtle rust established in Hawaii (USA), and over the past 12 years has spread from the Americas into Asia, the Pacific, and South Africa. Myrtle rust was detected in Australia in 2010, and the response and ultimately unsuccessful eradication attempt was a lesson to those concerned about the threat of exotic pests and diseases to Australia's environment. Seven years following establishment, we are already observing the decline of many myrtaceous species and severe impacts to native plant communities. However, the recently developed Myrtle rust in Australia draft action plan identified that there is no nationally coordinated response strategy for the environmental dimensions of this threat. Recent reviews have identified a greater need for involvement from environmental agencies in biosecurity preparedness, response, and resourcing, and we believe this approach needs to extend to the management of invasive environmental pathogens once they establish.

Ceratocystis species, including two new species associated with nitidulid beetles, on eucalypts in Australia.

The genus Ceratocystis includes important fungal pathogens of trees, including Eucalyptus spp. Ironically, very little is known regarding the diversity or biology of Ceratocystis species on Eucalyptus species in Australia, where most of these trees are native. The aim of this study was to survey for Ceratocystis spp., and their possible insect associates, on eucalypts in Australia and thus to establish a foundation of knowledge regarding these fungi on the continent. Collections were made in three states of Australia from wounds on trees, as well as from nitidulid beetles associated with these wounds. Ceratocystis spp. were identified based on morphology and multigene sequence comparisons. Of the 54 isolates obtained, two previously unknown species of Ceratocystis were found and these are described here as Ceratocystis corymbiicola sp. nov. and Ceratocystis tyalla sp. nov. Furthermore, the distribution of Ceratocystis pirilliformis is expanded to include Eucalyptus spp. in Tasmania.

Phylogeny of the Quambalariaceae fam. nov., including important Eucalyptus pathogens in South Africa and Australia.

The genus Quambalaria consists of plant-pathogenic fungi causing disease on leaves and shoots of species of Eucalyptus and its close relative, Corymbia. The phylogenetic relationship of Quambalaria spp., previously classified in genera such as Sporothrix and Ramularia, has never been addressed. It has, however, been suggested that they belong to the basidiomycete orders Exobasidiales or Ustilaginales. The aim of this study was thus to consider the ordinal relationships of Q. eucalypti and Q. pitereka using ribosomal LSU sequences. Sequence data from the ITS nrDNA were used to determine the phylogenetic relationship of the two Quambalaria species together with Fugomyces (= Cerinosterus) cyanescens. In addition to sequence data, the ultrastructure of the septal pores of the species in question was compared. From the LSU sequence data it was concluded that Quambalaria spp. and F. cyanescens form a monophyletic clade in the Microstromatales, an order of the Ustilaginomycetes. Sequences from the ITS region confirmed that Q. pitereka and Q. eucalypti are distinct species. The ex-type isolate of F. cyanescens, together with another isolate from Eucalyptus in Australia, constitute a third species of Quambalaria, Q. cyanescens (de Hoog & G.A. de Vries) Z.W. de Beer, Begerow & R. Bauer comb. nov. Transmission electron-microscopic studies of the septal pores confirm that all three Quambalaria spp. have dolipores with swollen lips, which differ from other members of the Microstromatales (i.e. the Microstromataceae and Volvocisporiaceae) that have simple pores with more or less rounded pore lips. Based on their unique ultrastructural features and the monophyly of the three Quambalaria spp. in the Microstromatales, a new family, Quambalariaceae Z.W. de Beer, Begerow & R. Bauer fam. nov., is described.