The role of Phytophthora multivora in Eucalyptus gomphocephala (tuart) woodland decline

Since the 1990’s Eucalyptus gomphocephala (tuart) has been suffering a significant decline in Yalgorup National Park, approximately 100 km south of Perth Western Australia. Symptoms range from chronic deterioration to sudden mass collapse. The role of Phytophthora pathogens was investigated because the progressive canopy thinning, dieback and heterogeneous distribution of the decline were similar to other forest declines caused by a range of Phytophthora species which are widespread throughout south-west Western Australia and worldwide. In combination with sampling for Phytophthora isolation, an initial diagnostic trial tested the effect of trunk applied phosphite, nutrients and combined phosphite and nutrients on natural stands of declining E. gomphocephala. Phosphite injection was used as a diagnostic tool to identify the possible role of Phytophthora pathogens because the chemical specifically suppresses Phytophthora pathogens and has no known direct fertilizer effect on the host. A range of nutrient treatments was also applied as a diagnostic tool to indicate what nutrient deficiencies may be involved in the decline. Initially no Phytophthora species were isolated from the treatment sites. However, individual and combined injection treatments of phosphite and nutrients improved the crown health over four years with the greatest improvement from treatments of phosphite and zinc sulphide. In combination with further rhizosphere sampling for Phytophthora species, the response of declining trees to phosphite application was further investigated in a second injection trial. Phosphite concentrations from 75 to 375 g phosphite/L improved crown health compared to the control, with the best improvement at 150 g phosphite/L. The positive response of declining trees to phosphite injection implicated a Phytophthora pathogen, despite no Phytophthora species being isolated at this time. Consequently further work was undertaken to determine the involvement of Phytophthora species. Concurrently to both injection trials, several seedling bioassays were conducted. The first bioassay tested the effect of pasteurising soil from a declining site within Yalgorup and healthy sites outside the Yalgorup woodland on E. gomphocephala seedlings grown ex situ. Seedling growth ex situ was not significantly reduced in nonpasteurised soil compared to the pasteurised soils from declining sites, and no Phytophthora species was isolated. To further investigate the disease the fine root and ectomycorrhizal systems of the largest main lateral root of 18 declining E. gomphocephala trees within Yalgorup were exposed using an air spade. Necrotic roots were sampled and the crown, fine root and ectomycorrhizal health were assessed. No Phytophthora species was isolated from necrotic roots; however, crown health of the declining trees was significantly correlated with the fine root and ectomycorrhizae density, suggesting that below ground damage could be involved in the decline. The relationship between the above and below ground health of the air spaded trees was investigated further using an in situ and ex situ seedling bioassay. In the in situ bioassay, seedlings were planted within the exposed root mats of the air spaded trees. In the ex situ bioassay, seedlings were grown within a glasshouse in pasteurised and non-pasteurised soil collected from the air spaded root mats. No Phytophthora species was identified in these bioassays, and seedlings grown ex situ in non-pasteurised soil showed no clear decline symptoms, but the health of the woodland trees was significantly correlated with seedling survival, foliar health and height of the seedlings An additional 32 sites throughout the E. gomphocephala range were sampled for the presence of Phytophthora pathogens using a modified sampling and isolation procedure. From this survey a new Phytophthora species was isolated from five sites from the roots of declining E. gomphocephala, E. marginata and Agonis flexuosa at Yalgorup National Park. Morphologically similar to P. citricola, the new Phytophthora species is unique based on phylogenetic analysis of the ITS and Cox1 gene regions and was named P. multivora. Phytophthora multivora has subsequently been isolated from all experimental sites showing tuart decline. Two experiments tested the pathogenicity of P. multivora to E. gomphocephala and E. marginata. The first experiment examined ex situ the pathogenicity of five P. multivora isolates and one P. cinnamomi isolate on the root systems of E. gomphocephala and one P. multivora isolate on the root system of E. marginata. In the second experiment, the pathogenicity of P. multivora to E. gomphocephala and E. marginata saplings was measured in situ using under-bark stem inoculation. Phytophthora multivora isolates caused significant fine root loss and lesion extension in under-bark inoculated stems of both E. gomphocephala and E. marginata. Phytophthora multivora was also reisolated from necrotic fine roots and lesions of inoculated saplings of both E. gomphocephala and E. marginata, thus satisfying Koch’s postulates. No seedlings died in these pathogenicity trials and P. multivora was not reisolated from beyond the fine roots. There was evidence that P. multivora significantly contributes to E. gomphocephala decline by episodically causing fine root damage leading to chronic deterioration of the trees.