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Ceratocystis wilt and canker disease has devastated Acacia mangium plantations in south-east Asia. Current screening methodologies to identify resistant or tolerant germplasm use potted plants in a greenhouse as a preliminary screening to select material for field trials, but these tests are time-consuming, motivating a search for rapid screening protocols. In this study, inoculation procedures were tested on three species of Acacia, viz. A. mangium, A. crassicarpa and a hybrid of A. auriculiformis and A. mangium, using three isolates of Ceratocystis manginecans. Mycelial plugs were compared with spore suspensions as inoculum to infect artificial wounds on the stems of A. mangium potted plants. The rapid screening protocols involved inoculation of stem segments with mycelial plugs and detached phyllodes with a spore suspension, with susceptibility measured by lesion length on the stems or necrosis length on the phyllodes. Both mycelial plugs and spore suspensions produced a similar level of disease incidence, so either inoculum form can be used for an assay. The stem segments were prone to contamination by other fungi and to desiccation, while results from the potted plant and phyllode protocols showed similar trends of susceptibility among the Acacia clones and species. The ease, rapidity, and reproducibility of the phyllode inoculation protocol makes it a potential replacement for inoculation of potted plants as a preliminary screening protocol to identify disease tolerant A. mangium germplasm prior to field screening.

Three screening trials of clonally replicated Acacia mangium seedlings were evaluated for survival and lesion length following inoculation with locally collected strains of Ceratocystis in Indonesia. Tolerance in the population was low with 6.7% of the 1033 clones represented by more than 4 ramets surviving repeated inoculations. Differences in tolerance among populations were slight; however, populations with consistently higher survival and shorter lesion lengths were from Papua New Guinea rather than Queensland. Estimates of the proportion of the experimental variation attributable to differences among parents (heritability) were low to moderate for both survival and lesion length. Estimates of the proportion of the experimental variation that was attributable to differences among clones (repeatability) were greater but typically similar to the heritability estimates, indicating that initial improvements from selection will primarily be derived from identifying tolerant parents. While genetic correlations among experiments were positive, estimates could not exclude the existence of host–pathogen interactions. Two validation trials of the tolerant clones were assessed 9 months after establishment; these trials verified that one-third of the clones identified in the nursery screening were also tolerant to Ceratocystis in field trials. The experiments confirmed that nursery screening may be used to quickly focus efforts on parents that produce more tolerant progeny, screening additional seedlings to increase selection intensity rather than using clonal replication to increase accuracy would lead to greater improvements in tolerance and field trials are required to verify disease tolerance at later ages.

Polypores with complex double-walled basidiospores with a pigmented endosporium ornamented with columns or ridges and a smooth hyaline exosporium are usually classified in the Ganodermataceae. Although the family presents well-delimited morphological features, recent phylogenetic studies have raised divergent evidences about its monophyly. In order to clarify uncertainties of generic delimitation and classification of the group and test its monophyly, morphological analysis and multilocus phylogenetic analyses based on Internal Transcribed Spacer, nuclear 28S rDNA, and portions of the largest subunit of RNA polymerase II and translation elongation factor 1-alpha were carried out. The main outcomes of the study are (i) Ganodermataceae is recovered as monophyletic, (ii) Amaurodermellus is proposed to accommodate the neotropical Amauroderma ovisporum, (iii) Cristataspora is proposed to accommodate two former species of Ganoderma - G. coffeatum and G. flaviporum - with pale context and truncate basidiospores with endosporic ornamentation as ridges. Comments on other species of Ganodermataceae are also presented.

Amauroderma s.lat. has been defined mainly by the morphological features of non-truncate and double-walled basidiospores with a distinctly ornamented endospore wall. In this work, taxonomic and phylogenetic studies on species of Amauroderma s.lat. are carried out by morphological examination together with ultrastructural observations, and molecular phylogenetic analyses of multiple loci including the internal transcribed spacer regions (ITS), the large subunit of nuclear ribosomal RNA gene (nLSU), the largest subunit of RNA polymerase II (RPB1) and the second largest subunit of RNA polymerase II (RPB2), the translation elongation factor 1-α gene (TEF) and the β-tubulin gene (TUB). The results demonstrate that species of Ganodermataceae formed ten clades. Species previously placed in Amauroderma s.lat. are divided into four clades: Amauroderma s.str., Foraminispora, Furtadoa and a new genus Sanguinoderma. The classification of Amauroderma s. lat. is thus revised, six new species are described and illustrated, and eight new combinations are proposed. SEM micrographs of basidiospores of Foraminispora and Sanguinoderma are provided, and the importance of SEM in delimitation of taxa in this study is briefly discussed. Keys to species of Amauroderma s.str., Foraminispora, Furtadoa, and Sanguinoderma are also provided.

Ceratocystis wilt and canker disease has severely compromised the profitability of Acacia mangium plantations in Southeast Asia. The focus of this review is on Ceratocystis wilt and canker disease ...

Recent advances and commercialisation of unmanned aerial vehicle/red blue green (RGB) camera systems and digital photogrammetric techniques now provide a cheap and flexible alternative to higher-co...

Inoculations of two Acacia populations were used to estimate the level of genetic control for tolerance to Ceratocystis canker and wilt disease following inoculation with the causal pathogen, Ceratocystis manginecans. The first screening utilised a range of clones commonly established in Vietnam along with new candidate clones and provided evidence that variation in wilt symptoms is under a moderate level of genetic control in current deployment populations. Subsequent assessments of tolerance in a pedigreed Acacia auriculiformis progeny trial indicated that internal lesion length assessments were less influenced by seedling height and produced higher heritability estimates than external lesion length assessments. Heritability estimates for damage to detached phyllodes following inoculation increased from 2 to 4 weeks, at which point many leaves were entirely damaged. Assessments of phyllode damage were more heritable than assessments of stem damage, and positive genetic correlations between stems and phyllode damage indicate that families will rank similarly for both traits. If validation trials confirm the results of this study in larger trees, screening as described herein may be used to develop acacia breeds with greater disease tolerance.

The present study investigated the axenic basidiospore germination of Ganoderma australe, G. mastoporum and G. philippii at three spore densities in five nutrient media encompassing a range of carbohydrate complexity, in combination with sawdust and/or ethanol as medium additives. Five incubation temperatures ranging from 10 to 35 °C were used, totalling 300 treatment combinations. AWeibull model, with the asymptote occurring at 50 to 75 h, provided good fit to percentage spore germination data at each treatment combination for all three Ganoderma species. Ganoderma australe and G. mastoporum basidiospores germinated on all media, whereas G. philippii basidiospores required media that contained 2% ethanol. The best medium for G. australe and G. mastoporum basidiospore germination was rice dextrose agar with amixture of Eucalyptus and Acacia sawdust, whereas for G. philippii it was 1% malt extract agar plus ethanol, with or without sawdust. Spore density was also critical to achieving the best germination rate, with ~400 spores/cm2 optimal for all three species. As Ganoderma root rot disease affects commercial Acacia mangium and Eucalyptus pellita plantations, the greater understanding of basidiospore germination gained from the current study should assist in developing strategies to contain the dispersal and spread of root rot in Indonesia and other southeast Asian countries where these species have been planted.

Summary Structural host responses of young Acacia mangium and Eucalyptus pellita to infection with the root rot pathogen Ganoderma philippii were described. To our knowledge, this is the first report investigating the histology of root responses infected with G. philippii. The infected roots of A. mangium were characterized by the presence of a continuous multilayered mycelial sheath (~100–200 μm thick), fungal outgrowths and the production of a wound periderm. In contrast, roots of E. pellita were characterized by a mycelial sheath and fungal outgrowths and a wound periderm comprising of only one to four rows of cells.

Climate change could significantly alter forest productivity and climax states. Hence modelling productivity under climate change will need to account for many alternative ecosystem states. We apply qualitative modelling to identify the most likely ecosystem representations for a well-researched Tasmanian forest. Its main ecosystem is a tiered forest with rainforest, wet sclerophyll and myrtaceae components. Interactions between these components are uncertain, especially under additional pressures from climate change. Qualitative modelling is a structured method to analyse these uncertainties. We identify the most appropriate models and research efforts for model development. Further, we identify research needs for interactions between root pathogens and forest components, with research on some impacts of system components on fire being ruled out. The qualitative modelling approach applied here was useful in identifying research priorities for modelling complex ecosystems, even under uncertain system understanding or deficiencies in quantitative data. Qualitative modelling provides a way of conceptualising and addressing uncertainty in ecosystems under climate change.Limited knowledge of ecosystem interaction requires alternative ecosystem models, which results in multiple research avenues.Testing model predictions against known system dynamics enables excluding less likely models and model component linkages.Identifying less likely models and linkages focuses research efforts on critical knowledge gaps.

Needle fungi have been extensively studied in conifers but rarely in Pinus radiata. Previous studies of P. radiata have been based on fungal isolation and not direct PCR detection from needles. This research was a component of a study examining factors linked to spring needle cast (SNC) in Tasmanian P. radiata plantations and aimed to identify as many as possible of the fungal species commonly associated with P. radiata needles in Tasmanian plantations. Needle samples were collected from 13 sites representative of the range of sites in which P. radiata is grown in Tasmania. Fungi were detected by a direct PCR approach and identified using barcode sequences from a reference collection of isolates from Tasmania, mainland Australia and New Zealand as well as sequences from the international nucleotide sequence databases. The total number of molecular operational taxonomic units (MOTUs) was 152, with 127 detected by direct PCR and sequencing, and only 35 operational taxonomic units (OTUs) isolated by culture-...

Plant‐soil feedbacks (PSFs) are important drivers of plant community structure and diversity, with species varying in the way they both condition soils and respond to them. While plant phylogenetic relationships alone can predict this variation in some instances, trait conservatism across phylogenies may provide more reliable predictions. Using integrated common garden and glasshouse inoculation experiments including 13 Eucalyptus species across two subgenera, we specifically investigated soil microbial conditioning and root chemical traits as underlying drivers of phylogenetic differences in PSF. We found that eucalypt species responded variably to soils conditioned by closely related species, depending on their phylogenetic lineage, which was further related to root terpene concentrations and the presence/absence of specific fungal taxa in conditioned soils. Overall, these findings show that trait conservatism in root chemical traits and the subsequent conditioning of soil microbial communities can explain whether or not plants show phylogenetic patterns in PSF.

Ecosystem change is predicted to become more prevalent with climate change. Widespread dieback of cushion plants and bryophytes in alpine fellfield on Macquarie Island may represent such change. Loss of the keystone endemic cushion plant, Azorella macquariensis, was so severe that it has been declared critically endangered. We document the dieback and its extent. Due to the rapidity of the event, we sought to infer causes by testing two mechanistic hypotheses: (i) that extensive dieback was due to a pathogen and (ii) that dieback was a consequence of a change in climate that induced stress in several susceptible species. We searched for pathogens using both conventional and next-generation sequencing techniques. We examined plant functional morphology in conjunction with a long-term climate record of plant-relevant climate parameters to determine whether environmental conditions had become inimical for A. macquariensis. Dieback was found across the entire range of A. macquariensis. A survey found 88% of 115 stratified/ random sites contained affected cushions and 84% contained dead bryophytes. Within-site dieback increased over time. No conclusive evidence that A. macquariensis deaths were caused by a definitive disease-causing pathogen emerged. However, the presence of bacterial, fungal and oomycete taxa, some potentially pathogenic, suggested that stressed cushions could become susceptible to infection. The primary cause of collapse is suspected failure of A. macquariensis and other fellfield species to withstand recent decadal changes in summer water availability. Increased wind speed, sunshine hours and evapotranspiration resulted in accumulated deficits of plant available water spanning 17 years (1992-2008). High vulnerability to interrupted water supply was consistent with functional morphology of A. macquariensis, and climate change has altered the species' environment from wet and misty to one subject to periods of drying. Synthesis and applications. With alpine fellfield dieback baseline data on Macquarie Island established, future monitoring will determine whether this event represents a transient, decadal-level change in the ecosystem or the initiation of a climate-related, transformation away from an Azorella-dominated fellfield ecosystem. That mechanisms driving ecosystem collapse were complex and multiple stressors appeared to be impacting cumulatively may be relevant to other locations. With alpine fellfield dieback baseline data on Macquarie Island established, future monitoring will determine whether this event represents a transient, decadal-level change in the ecosystem or the initiation of a climate-related, transformation away from an Azorella-dominated fellfield ecosystem. That mechanisms driving ecosystem collapse were complex and multiple stressors appeared to be impacting cumulatively may be relevant to other locations.

Puccinia psidii is an important pathogen considered a threat to plants of the Myrtaceae around the world. Morphological variability has created misunderstanding around the classification of this pathogen. Due to the obligately biotrophic nature of P. psidii, the age of the material on which the original description was based, and the poor condition of its DNA, there has been no clear genetic characterization of the species. An epitype is therefore designated for P. psidii for precise application of this taxon name. A morphological description and DNA characterization are provided from the epitype, obtained from the same location and host of the lectotype, also designated herein. Morphologically, the epitype is identical to the original material and phylogenetic and microsatellite analyses show that the epitype is representative of P. psidii from guava in Brazil.

Summary Spring needle cast (SNC) in Tasmania is defined by the early casting of pine needles and appears in 6- to 7-year-old pine plantations after canopy closure. The severity of SNC can vary significantly among trees growing within the same plantation. Fungal communities were surveyed in a Pinus radiata family trial, using DNA extraction from needles, PCR with fungal-specific primers and sequencing to detect the presence of fungal species. Samples were taken at the same time as the trees were scored for disease severity. Trees with contrasting levels of SNC disease severity have significantly different needle fungal communities, but family pedigree and different ages of needle are not clearly distinguished by their fungal communities. All common fungal pathogens previously implicated in causing SNC were identified from the study, but of these, only Lophodermium pinastri was correlated with high levels of disease. Several species of Teratosphaeriaceae were detected, and one of these was also strongly associated with needle cast.

Two Acacia seed orchards, one each of A. mangium and A. auriculiformis , in Central Java have suffered high losses from root rot associated with a species of Ganoderma that is distinct from G. philippii , which causes extensive losses in A. mangium plantations in Sumatra and Kalimantan. Ganoderma steyaertanum was consistently isolated from roots of affected trees in the Javanese orchards and pathogenicity was confirmed by satisfying Koch's postulates. The mode of spread of the pathogen was examined by analysing the distribution of tree deaths in the A. mangium seed orchard and conducting somatic incompatibility tests among G. steyaertanum isolates from diseased trees. Spatial analyses indicate that, after an initially random disease distribution, the pattern of tree deaths was clustered. This is consistent with spore colonisation of thinned stumps followed by vegetative spread through root-to-root contact, as has been observed for other root pathogens such as Heterobasidion annosum . Somatic incompatibility tests demonstrated high genetic variability of the pathogen with root-to-root spread also confirmed.

The rapid emergence of Acacia mangium as the key industrial plantation species in Indonesia has been followed by the equally rapid emergence of red root rot ( Ganoderma philippii ) as its potential nemesis. As a consequence, and on severely affected mineral soils in equatorial tropical environments in particular, A . mangium may no longer be capable of producing commercial yields after three rotations. In this experiment, 100-tree plots were established in commercial plantings of A . mangium at five sites which represented the range of growing conditions used for plantation establishment in Indonesia. Repeated monitoring at approximately 6-month intervals of above- and below-ground variables was used to explore relationships between measures of tree health and root rot. Crown colour and density were poor predictors of either the presence or severity of infections. Tree mortality increased more or less linearly with time and led to the progressive coalescence of initially discrete disease gaps. The average rate of disease development was about 0.3% per month, and average time from infection to tree death was conservatively estimated at around 1 year. Trees with more than four dead/missing neighbours had a >50% chance of being dead by the following monitoring. By the end of the monitoring period >40% of trees were classified as dead/missing, although this value varied from 20 to 70% depending on site, tree age and rotation. Canonical discriminant analysis correctly allocated >90% trees into their correct group on the first monitoring and similarly classified whether trees would be dead or missing in the following monitoring.

DNA sequence variation in the rDNA internal transcribed spacers (ITS) was exploited to design species-specific primers for Ganoderma philippii and Ganoderma mastoporum , two fungi associated with red root rot in Acacia mangium and other perennial crops. Specificity was verified against five other Ganoderma species, 34 other basidiomycete species and two ascomycete species. The PCR tests assisted in the identification of 822 fungal isolates obtained from A. mangium and Eucalyptus pellita plantations in Indonesia. These tests have provided a high level of confidence in the identification of the major root rot pathogens, information that can be used to support the development of management options for their control in A. mangium plantations.

Summary This study describes and identifies the fungi associated with root-rot disease in Eucalyptus pellita, associated anecdotally with a species of Phellinus. Macro- and microscopic morphological observations and DNA analysis were used to identify sporocarps and fungal cultures growing from symptomatic root samples. Investigation of 12 sites in Central Sumatra revealed that red root-rot disease caused by species of Ganoderma (most commonly Ganoderma philippii) was as prevalent as root disease caused by Phellinus noxius. The study also detected the presence of several potential fungal agents of root rot at any one site. Fungal signs such as absence/presence of sporocarps were a poor indicator of root-rot incidence in stands. Instead, fungal isolation from infected roots was essential to obtain accurate identification of active pathogens.

An investigation of root rot in Acacia mangium plantations in Indonesia generated over 1000 fungal isolates from field surveys at six locations. The majority of isolates were identified as Ganoderma philippii or Ganoderma mastoporum by species-specific PCR, but this still left 274 isolates to be identified. As barcoding initiatives for fungi are still in the early stages, to assist identification of these isolates in this way we developed a database of sequences based on identified sporocarps and also searched public DNA sequence databases for sequences with high similarity. The database of sequences from sporocarp collections is more limited than public DNA databases. This resulted in more reliance on public DNA databases for a high proportion of taxa. Several pathogenic species were identified, with Phellinus noxius occurring at several locations and on both A. mangium and Eucalyptus pellita . Ganoderma steyaertanum was isolated from A. mangium roots and Rigidoporus microporus from E. pellita roots, each at a single location only. Potential pitfalls of using public DNA databases for fungal identification are discussed and methods to avoid these and increase confidence in the identification are presented.

Ganoderma philippii causes root-rot disease in a wide range of commercial perennial woody crops such as rubber and tropical acacias. In terms of productivity loss, the disease is considered as the most economically damaging disease of Acacia mangium . Many strategies to manage root rot have been developed and applied, including application of biological control which has focused on species and strains of the ascomycete genus Trichoderma . This research aimed to test the potential of a wood-rotting basidiomycete as a biocontrol agent for root rot caused by Ganoderma philippii . This basidiomycete was isolated from a Eucalyptus pellita stump in a coppicing trial in Riau province, central Sumatra. DNA sequence analysis indicated that this isolate is most likely a Phlebiopsis sp. In vitro tests and microscopic morphological description of mycelial interactions between the Phlebiopsis and G. philippii isolates showed a mycoparasitic reaction. This result offered an alternative potential fungal isolate for root-rot management in Indonesia as Phlebiopsis gigantea is already successfully used as a biocontrol agent for managing root rot caused by Heterobasidion annosum throughout pine forests in Europe and North America.

USDA-Forest Service, RMRS-Forest and Woodlands Ecosystem Program, Western Wildlands Environmental Threat Assessment Center, Special Technology Development Program, State and Private Forestry, Forest Health Protection-Region 5; Conselho Nactional de Desenvolvimento Cientifico e Tecnologico, Brasil (CNPq); Fundacao de Amparo a Pesquisa do Estado de Minas Gerais (FAPEMIG); Research Joint Venture Agreements RMRS 15-JV-11221633-160 (Jane Stewart, Colorado State University) and RMRS 14-JV-11221633-117 (Western Forest Conservation Association).

Fungal rot of sweet cherry fruit leads to reduced yield and quality at harvest. This study investigated a range of host factors related to cherry rot and also identified the main species present in latently infected-fruit and fruit with visible rot at harvest in a Tasmanian orchard. There was a significant effect of tree cultivar on total rot found at harvest, with the cultivar “Sweetheart” associated with almost double (approx. 25 % infection) the amount of rotten fruit compared to “Regina” and “Simone” (approx. 13 %). Harvest date may have had some influence on this result, as Sweetheart fruit were harvested 2 weeks later than the other cultivars, and there was a large rainfall event just before Sweetheart harvest (rainfall can increase disease spread and infection). Crop load did not influence the amount of rotten fruit found at harvest for any cultivar, although crop load was low in this season at this site. Removing fungicide application from the point of 50 % full bloom onwards did not significantly alter disease incidence. The study found that Botrytis cinerea was the dominant rot pathogen in cherry fruit at this site and season. No evidence of Monilinia species was found in this study with the methods used, despite it being typically associated with rot of stone fruit. Fruit quality characteristics were not indicators of disease susceptibility, as very few characteristics observed to correlate with disease at harvest.

Cyclaneusma Needle Cast is associated with the fungal pathogen Cyclaneusma minus . The disease has led to significant losses of Pinus radiata production in New Zealand, but the disease appears less serious in Australia. Two cultural morphotypes of C. minus have been discriminated in New Zealand, and isolates from Australia also correspond to these two morphotypes. To examine if the two morphotypes of Cyclaneusma are different species, phylogenetic analysis using sequences from five gene regions (rDNA ITS, mt-LSU, n-LSU, tef–1 and βT-1) was conducted. This study revealed that C. minus consists of two distinct clades which correspond to the two morphotypes. One morphotype (‘verum’) was more closely related to Cyclaneusma niveum than to the other morphotype (‘simile’). A morphological description is required to formalise the new species represented by the ‘simile’ clade; an investigation of the individual pathogenic behaviours of both morphotypes is also required.

Cyclaneusma needle cast (CNC) is a pine disease caused by the ascomycetous fungus Cyclaneusma minus (Butin) DiCosmo, Peredo and Minter. The pathogen occurs worldwide but is of particular significance in New Zealand where it infects Pinus radiata D. Don plantations. There are two morphological types of C. minus, termed C. minus ‘simile’ and C. minus ‘verum’, recently shown by multigene phylogenetic analysis to belong to distinct clades and therefore proposed to be two separate species. It is currently unknown whether one or both of these molecular operational taxonomic units (MOTUs) are responsible for CNC. In this study, DNA analysis of the rDNA internal transcribed spacer (ITS) region was carried out on 120 isolates of C. minus collected in New Zealand since 1969 to distinguish C. minus simile from C. minus verum. Specific primers for C. minus simile and C. minus verum were developed for molecular differentiation from pure cultures and direct amplification from infected needles. Using these specific primers, the distribution of C. minus simile and verum was determined from isolates collected throughout New Zealand. The C. minus simile and C. minus verum primers developed were specific to these morphotypes only and were successfully used to identify simile and verum MOTUs from culture. Morphological typing of cultures was consistent with specific primer results in only 56% of isolates tested, demonstrating the high level of skill and experience required for accurate morphotyping from culture alone. C. minus simile was more common throughout New Zealand including regions where CNC is known to be more prevalent. From a collection of 120 isolates, 89% of North Island isolates and 59% of the South Island isolates were C. minus simile. The C. minus simile and verum specific primers were used to detect Cyclaneusma DNA in symptomatic P. radiata needles. C. minus simile was detected in 39% of the needle samples and C. minus verum was only detected in 11%. However, Cyclaneusma DNA was not detected by PCR in 50% of the sampled needles. The molecular methods developed for differentiation of C. minus simile and C. minus verum can be used to rapidly identify these morphotypes from cultures or from infected needles, which reduces the time otherwise required for morphological identification. Analysis using these tools for characterisation of C. minus sensu lato, isolated from P. radiata plantations in New Zealand since 1969, revealed that C. minus simile was more common than C. minus verum.

Eucalypt forest decline has a complex aetiology often linked to altered soil chemistry caused by environmental disturbances. Forest decline has also been linked to alterations in ectomycorrhizal (ECM) fungal communities, which are imperative for nutrient transfer and affect ecosystem productivity and health. Our aim was to determine the influence of soil chemistry on ECM fungal communities and tree health in declining temperate eucalypt forests. We hypothesise that forests with changed soil chemistry, in particular altered nitrogen cycling associated with forest decline, supports unique ECM fungal communities. ECM communities from twelve Eucalyptus delegatensis forest plots were characterised by DNA sequencing of root tip and sporocarp samples. Tree health and nutrient concentrations from soil and foliage samples were quantified for each plot. Multivariate and regression analyses and t-tests were used to determine ECM fungal community differences between forest health classes, and identify which soil variables were important for defining these communities. Elevated available soil nitrogen and soil acidity were associated with severely declining forest. Soil pH, nitrate and organic carbon significantly explained the majority of variation in ECM fungal community composition and structure, which differed between moderately and severely declining forest. Russulaceae species richness was greatest in acidic soils (severely declining forest) while Cortinariaceae species richness was greatest in soils with lower concentrations of soil nitrate (moderately declining forest). Total ECM fungal richness was inversely related to available soil phosphorus and soil nitrate. Thus, altered soil chemistry associated with eucalypt forest decline mediates changes in the ECM fungal community. Forest management must consider the role of disturbance in maintaining suitable soil conditions for symbiotic fungi which are important for maintaining healthy eucalypt forest and restoring declining forest ecosystems.

A device comprising a filter attached to a vacuum cleaner was purpose-built to sample rust spores from three potentially high-risk pathways in Australia: passengers, fresh flowers, and sea cargo. The proportion of spores recovered from eight surfaces comparable with those on each pathway (cotton, denim, roses, carnations, chrysanthemums, wood, plastic, and metal) was estimated in the laboratory. Spore recovery percentages were highest for denim clothing (61% Puccinia triticina Erikss. and 62% Uromycladium tepperianum) and lowest for carnations (4% P. triticina Erikss. and 5% U. tepperianum). Subsequently, the device was tested at several locations on the Central Coast of New South Wales, Australia, recently affected by a “myrtle rust” outbreak. Symptomatic and asymptomatic myrtle rust hosts, myrtle rust nonhosts, and inanimate objects (e.g., clothing and vehicles) were sampled in conjunction with the emergency response to the outbreak. A polymerase chain reaction (PCR) assay developed for P. psidii established the presence of myrtle rust, and visual inspections provided spore count estimations. All samples from symptomatic myrtle rust hosts produced positive PCR results and spore count estimations were generally much greater. Several samples from asymptomatic myrtle rust hosts, myrtle rust nonhosts, and inanimate objects also produced positive PCR results; however, there were discrepancies between PCR results and spore count estimations in some of these samples, all of which had

The possible drivers and implications of an observed latitudinal cline in disease resistance of a host tree were examined. Mycosphaerella leaf disease (MLD) damage, caused by Teratosphaeria species, was assessed in five Eucalyptus globulus (Tasmanian blue gum) common garden trials containing open-pollinated progeny from 13 native-forest populations. Significant population and family within population variation in MLD resistance was detected, which was relatively stable across different combinations of trial sites, ages, seasons and epidemics. A distinct genetic-based latitudinal cline in MLD damage among host populations was evident. Two lines of evidence argue that the observed genetic-based latitudinal trend was the result of direct pathogen-imposed selection for MLD resistance. First, MLD damage was positively associated with temperature and negatively associated with a prediction of disease risk in the native environment of these populations; and, second, the quantitative inbreeding coefficient (QST) significantly exceeded neutral marker FST at the trial that exhibited the greatest MLD damage, suggesting that diversifying selection contributed to differentiation in MLD resistance among populations. This study highlights the potential for spatial variation in pathogen risk to drive adaptive differentiation across the geographic range of a foundation host tree species.

The rust Puccinia psidii infects many species in the family Myrtaceae. Native to South America, the pathogen has recently entered Australia which has a rich Myrtaceous flora, including trees of the ecologically and economically important genus Eucalyptus. We studied the genetic basis of variation in rust resistance in Eucalyptus globulus, the main plantation eucalypt in Australia. Quantitative trait loci (QTL) analysis was undertaken using 218 genotypes of an outcross F2 mapping family, phenotyped by controlled inoculation of their open pollinated progeny with the strain of P. psidii found in Australia. QTL analyses were conducted using a binary classification of individuals with no symptoms (immune) versus those with disease symptoms, and in a separate analysis dividing plants with disease symptoms into those exhibiting the hypersensitive response versus those with more severe symptoms. Four QTL were identified, two influencing whether a plant exhibited symptoms (Ppr2 and Ppr3), and two influencing the presence or absence of a hypersensitive reaction (Ppr4 and Ppr5). These QTL mapped to four different linkage groups, none of which overlap with Ppr1, the major QTL previously identified for rust resistance in Eucalyptus grandis. Candidate genes within the QTL regions are presented and possible mechanisms discussed. Together with past findings, our results suggest that P. psidii resistance in eucalypts is quantitative in nature and influenced by the complex interaction of multiple loci of variable effect.

Fungal diversity of Australian eucalypt forests remains underexplored. We investigated the ectomycorrhizal (EcM) fungal community characteristics of declining temperate eucalypt forests in Tasmania. Within this context, we explored the diversity of EcM fungi of two forest types in the northern highlands in the east and west of the island. We hypothesised that EcM fungal community richness and composition would differ between forest type but that the Cortinariaceae would be the dominant family irrespective of forest type. We proposed that EcM richness would be greater in the wet sclerophyll forest than the dry sclerophyll forest type. Using both sporocarps and EcM fungi from root tips amplified by PCR and sequenced in the rDNA ITS region, 175 EcM operational taxonomic units were identified of which 97 belonged to the Cortinariaceae. The Cortinariaceae were the most diverse family, in both the above and below ground communities. Three distinct fungal assemblages occurred within the wet and dry sclerophyll forest types and two geographic regions that were studied, although this pattern did not remain when only the root tip data were analysed. EcM sporocarp richness was unusually higher than root tip richness and EcM richness did not significantly differ among forest types. The results are discussed in relation to the importance of the Cortinariaceae and the drivers of EcM fungal community composition within these forests.

Free air CO2 enrichment (FACE) experiments are considered the most reliable approach for quantifying our expectations of forest ecosystem responses to changing atmospheric CO2 concentrations [CO2]. Because very few Australian tree species have been studied in this way, or are likely to be studied in the near future because of the high installation and maintenance costs of FACE, there are no clear answers to questions such as: (1) which species will be the winners in Australia's natural forests and what are the implications for biodiversity and carbon (C) sequestration; and (2) which will be the most appropriate species or genotypes to ensure the sustainability of Australia's plantation forests. We examined possible experimental approaches that may provide insights into, and more rapid assessment of, responses to elevated [CO2]. Our main conclusions were: (1) better understanding the extent to which species are C-limited could indicate when elevated [CO2] might be expected to increase photosynthesis and biomass production. Plant tissue carbohydrate concentrations can be used to assess any C limitation. Consistently high levels of carbohydrates indicate that plants are not C limited, but rather that growth is determined by other limiting resources or by rates of cell development and expansion; (2) historical examination of forest responses to increasing atmospheric [CO2] using stable isotopes in wood cores can provide clues as to which species may respond favourably to increasing [CO2], although it may remain difficult to distinguish between the environmental conditions under which favourable responses occurred. Undertaking stable isotope studies close to anthropogenic CO2 sources has the potential to provide insights into how species may respond to the higher [CO2] that is predicted during this century; (3) by focusing on genetic and metabolomic regulation of source and sink activity, selection for greater biomass production under elevated [CO2] is possible.

Acacia mangium is a fast-growing tree species. It is mainly planted in large monocultures for pulpwood in South-East Asia. Root rot has become the most economically damaging disease of this species with high tree mortality rates observed during second and third rotations. Two main types of root rots have been found in A. mangium, viz. brown root-rot and red-root disease caused by Phellinus spp. and Ganoderma spp., respectively. To assess the future management options for root rot of A. mangium, we review past and current disease-management strategies for root rot in different temperate and tropical industrial tree crops. The efficacies of a wide range of silvicultural, chemical and biological options are detailed, and their potential utilization in managing root rot of A. mangium is discussed. We conclude that the current gaps in knowledge regarding identification, biology and disease epidemiology of the root-rot pathogens will need to be addressed so that effective management options can be developed

A species-specific, nested polymerase chain reaction (PCR)-based detection assay, using two primer sets designed from the rRNA ITS region, was developed for Puccinia psidii. Detection sensitivities of one urediniospore alone, or one or two urediniospores in the presence of pollen or leaf tissues, respectively, were observed. The assay reliably, accurately and sensitively detected the rust from naturally infected, geographically widespread eucalypt and fruit tree plantation and nursery species from diverse tissues types (e.g. leaves, flowers, fruits, pollen, seeds and woody material) including symptomless or cryptically contaminated plants or plant tissue. Independent testing in Brazil and Australia demonstrated the international inter-laboratory transferability of the P. psidii assay required for germplasm screening, disease monitoring and quarantine and incursion management, towards which the assay has already been employed.

Species richness and species composition of ectomycorrhizal (EM) fungi were compared among rehabilitated mine sites and unmined jarrah forest in southwest Western Australia. Species richness, measured in 50 m × 50 m plots, was high. In the wetter, western region, mean species richness per plot in 16-year-old rehabilitated mine sites (63.7 ± 2.5, n = 3) was similar to that of unmined jarrah forest (63.6 ± 9.6, n = 9). In the drier, eastern region, species richness in 12-year-old rehabilitated mine sites (40.3 ± 2.1, n = 3) approached that of nearby forest (52.4 ± 9.3, n = 9). Species composition was analysed by detrended correspondence analysis. Rehabilitated sites of similar age clustered together in the analysis and species composition was closer to the native jarrah forest in the older rehabilitated plots. In unmined forest, species composition of fungal communities in the wetter, western region was different from communities in the drier, eastern region.

Puccinia psidii is considered a biosecurity threat in Australia because of its broad host range that includes many species of Myrtaceae which dominate Australian ecosystems. Since it was first reported in this country, in April 2010, there has been little information about the population structure of the pathogen. In this study, six microsatellite loci were analysed to determine the genetic relationship among rust specimens from different hosts and locations in Australia, New Caledonia, Hawaii and China. The Chinese and New Caledonian specimens share a multi-locus genotype with the majority of the Australian specimens. The results also indicated a close relationship between Australian and Hawaiian samples. At present, the P. psidii population in Australia is genetically uniform with no evidence of sexual recombination. Five of the 104 collections varied by one allele at single loci, indicating that mutations are common but persistence of the mutants in the population may be less common.

A molecular survey of basidiomycete ectomycorrhizal fungi colonising root tips at a site in Eucalyptus marginata (jarrah) forest revealed the presence of many fungal species which could not be identified from a database of ITS-PCR-RFLP profiles from morphologically identified species. Three of these unidentified taxa were among the six most frequently encountered profiles. Phylogenetic analyses of ITS and nuclear LSU sequences revealed a close relationship among the three fungi and that they belong to the family Sebacinaceae (sensu Weiss and Oberwinkler 2001). The possibility that DNA of non-ectomycorrhizal rhizosphere or endophytic fungi had been amplified selectively by the basidiomycete-specific primers was tested by amplification with fungal-specific primers. A single PCR fragment was amplified in all but two of the 24 samples tested and digestion with two restriction enzymes produced RFLP profiles which matched those from the Sebacinoid sequence. We conclude, therefore, that at least three species of Sebacinaceae are common ectomycorrhizal associates of E. marginata.

Summary In Vietnam, the productivity of Acacia hybrid (Acacia mangium x A. auriculiformis) plantations is being threatened by an aggressive canker pathogen, Ceratocystis manginecans, and selection for tolerance is the main control strategy. A pot trial was established in Binh Duong province to screen for the host response of nine Acacia genotypes (six Acacia hybrid clones, two A. auriculiformis clones and mixed provenance seedlings of A. mangium) to artificial inoculation with three isolates of C. manginecans. Lesion lengths as measured on the inner bark suggested that the two A. auriculiformis clones were relatively more tolerant to C. manginecans than the A. mangium genotype. In contrast, the lesion lengths of all six Acacia hybrid clones fell between the A. auriculiformis and A. mangium genotypes. The results of this study indicate that among the Acacia hybrid clones, BV10 showed the most tolerance to C. manginecans. Chemical analysis of crude sapwood extracts sampled from the lesion provided some evidence that induced phenolic compounds, particularly tetrahydroxyflavanone and condensed tannins may have a defensive role in the Acacia—C. manginecans pathosystem. However, results were not consistent across individual Acacia hybrid clones and A. mangium genotypes.

Gondwanan vegetation, and the Australian region in particular, is species rich for ectomycorrhizal fungi in epigeous and hypogeous forms with over 100 species recorded in small (1 ha) patches of forests. Distinguishing co-occurring ectomycorrhizal fungi as root associations in native (natural or wildlands) vegetation or plantations and discriminating them from other larger basidiomycetes, e.g. wood and leaf litter decomposer fungi, places large demands on molecular identification, especially if interspecific similarities and intraspecific variation occur in target sequences. One hundred and nine species of larger basidiomycetes from a single forest location were characterised by PCR-RFLP profiles of two genomic regions (nuclear rDNA ITS and mtLSU). Over one-third of the species for which multiple isolates were tested showed intraspecific variation in either one or both genomic regions. This remarkably high variation questions previous assumptions about intraspecific ITS sequence variation and highlights the value of integrated molecular and morphological databases including voucher specimens. It also emphasises the value of molecular investigations that use more than one genomic region. Interspecific similarities were common among the Cortinariaceae, especially in the ITS region. Discrimination of most Cortinariaceae species was achieved using variation in the mtLSU region in conjunction with the ITS. This new information raises the possibilities that the ITS sequence is more conserved and the mtLSU more variable than among species of the other 23 families. In the other families, interspecific ITS variation was greater and the mtLSU profiles grouped species within families. The high variation in the two genomic regions indicated possible differences in the fungal population structure between two adjacent, differently managed blocks of Eucalyptus marginata forest. The significance of this variation to ecology, biodiversity assessment and ecosystem management are discussed.

Techniques to rapidly identify the basidiomycete fungal partner of ectomycorrhizal associations would be a major advantage for ecological, fungal population dynamics and life history studies of epigeous and hypogeous forms in plantations, forests, wild lands and other native or natural vegetation. PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism) identification of DNA regions is an available technique; however, primers which have a high probability of amplifying only the basidiomycete DNA are needed. Here we have assessed the specificity, sensitivity and discrimination of six different primer pairs, three targeting nuclear and three mitochondrial regions, for use in identification of Australian basidiomycete fungi from Eucalyptus forests by matching PCR-RFLP patterns to morphologically defined species. Two sets of primers, one newly designed and targeting the nuclear ribosomal DNA internal transcribed spacers (ITS) and the other amplifying a fragment of mitochondrial large subunit ribosomal DNA met the requirements of high specificity and sensitivity, amplifying DNA from a broad range of larger basidiomycetes, with no amplification of plant, bacterial or ascomycete DNA. The specificity of the ITS primer pair was compared with that of ITS1-F/ITS4-B. PCR-RFLP of the two regions discriminated fungi to species level for 91 fungal species from 28 families. Hence these two DNA regions and the specific primers are a potential practical PCR-RFLP tool for identifying basidiomycetes associated with plants from field samples.

There is controversy surrounding the described life cycle of the rust fungus Puccinia psidii sensu lato, which causes disease on several plant species in the family Myrtaceae. The objective of this study was to determine whether P. psidii s.l. is autoecious by performing basidiospore inoculations, and microscopically examining the fate of basidiospores on the leaf surface and nuclear condition at different stages of rust development. No spermogonia developed on leaves of Agonis flexuosa inoculated either with a teliospore suspension or basidiospores naturally discharged from telia. Uredinial sori that developed in all three inoculations with teliospore suspensions and in one of the five inoculations with naturally-discharged basidiospores from telia were most likely the result of urediniospore infections. Microsatellite analysis revealed that isolates made from these uredinial sori had the same multilocus genotype as that of the original isolate. No signs of penetration of plant cells by basidiospores were observed on A. flexuosa and Syzygium jambos. The nuclear condition of mycelia of uredinial sori, urediniospores, teliospores, and four-celled metabasidia was typical of that in many rust fungi. Our study could not provide unequivocal proof that P. psidii s.l. is autoecious. While it is possible that it could be heteroecious, with an unknown alternate aecial host, it is also possible that basidiospores have lost the ability to infect Myrtaceae or are infrequently operational.

Summary Acacia hybrid (Acacia mangium × A. auriculiformis) is widely planted in Vietnam, and part of the estate is managed for solid timber products. This requires pruning and thinning – practices that through mechanical wounding can facilitate the entry of fungal organisms, leading to stem defects. The extent to which this happens in Acacia hybrid has not been previously studied in Vietnam. A destructive survey was conducted in a 3-year-old Acacia hybrid plantation at Nghia Trung in Binh Phuoc province, 18 months after the imposition of pruning and thinning treatments. Pruned trees had a higher incidence and severity of discoloration and decay in the stem than unpruned trees; thinning increased the incidence of stem decay and the severity of discoloration but not the severity of decay. An interaction between pruning and thinning did not influence the incidence of stem decay but did increase the severity of discoloration in thinning treatments. Across treatments, levels of discoloration and decay were

Mycosphaerella leaf disease (MLD) is a serious disease of two of the major eucalypt species grown in temperate regions worldwide, Eucalyptus globulus and E. nitens. More than 30 species of Mycosphaerella have been reported on eucalypts worldwide. Accurate, rapid, and early discrimination of Mycosphaerella spp. causing crown damage to E. globulus and E. nitens will assist the development of sustainable management strategies. This study describes the development, and incorporation in a nested polymerase chain reaction (PCR) approach, of specific primers for the detection and identification of Mycosphaerella spp. commonly reported from leaf lesions of E. globulus and E. nitens in Australia. Primer design was assisted by sequence alignment and phylogenetic analysis of 165 nonredundant sequences from the nuclear ribosomal DNA internal transcribed spacer regions of Mycosphaerella and related species. Phylo-genetic analysis revealed very high sequence similarity for two taxon groups, Mycosphaerella grandis and M. parva, and M. vespa, M. ambi phylla, and M. molleriana, and primers were designed to differentiate each of the two groups. Three other species, M. cryptica, M. nubilosa, and M. tasmaniensis, were distinct and distinguished by species-specific primers. In double-blind trials, the detection test accurately and rapidly identified Mycosphaerella spp. in cultures and discriminated against other pathogens that co-occur in or on Eucalyptus leaves, thereby verifying its reliability. The detection test has an internal amplification control in the first-round PCR with fungal-specific primers to raise confidence in test results, particularly to highlight negative results due to PCR inhibition. When applied to DNA extracted from leaf or stem samples either as multiple or single lesions, it detected and identified up to five Mycosphaerella spp. or taxon groups in both positively identified and in young (putative) MLD lesions. The samples were 20 mm2or larger in surface area and were collected while undertaking disease rating assessments in an experimental investigation of Eucalyptus plantations and regrowth forest. Using nested PCR detection, Mycosphaerella spp. were positively identified in 2 days, 1 to 5 months earlier than by classical methods, demonstrating the potential application of this detection test to the early discrimination of MLD components in ecological, epidemiological, and genetic investigations.

Uredo rangelii (myrtle rust) is reported for the first time in Australia — detected on 22 April 2010 &#x2014 from Agonis flexuosa, Callistemon viminalis and Syncarpia glomulifera. This taxon is morphologically distinct from Puccinia psidii, the cause of guava rust, but DNA sequence data place it in the P. psidii complex. Surveys up to the end of May 2010 have detected U. rangelii on cultivated shrubs and trees at four properties (two cut flower farms and two wholesale nurseries) on the Central Coast of New South Wales, with no records in native forest thus far. Containment operations are currently underway on infected properties.

Fungal sporocarps and cultures associated with signs and symptoms of root-rot disease were collected from Acacia mangium and other tropical hardwood species. The collections were identified by either morphological characters and/or by phylogenetic analysis based on DNA sequences as Ganoderma philippii, G. mastoporum, G. aff. steyaertanum, G. australe and Amauroderma rugosum. Phylogenetic analysis unequivocally placed in the G. philippii clade four sequences amplified from A. mangium root and butt tissue showing clear signs of red root-rot disease (roots are covered by a red rhizomorphic skin). Whereas G. philippii was the most frequently encountered fungal species in A. mangium with red root-rot disease, this study indicates that other fungal species related to G. mastoporum may cause root-rot disease with very similar symptoms. An isolate (FRIM 138) that had caused red root-rot disease in artificial inoculations carried out before this study and was presumed to be G. philippii, is here determined to be closely related to G. mastoporum, G. cupreum and G. sinense. A Ganoderma species associated with a yellow-brown root-rot disease killing trees in an A. mangium plantation in Central Java, previously identified as G. lucidum, is shown by phylogenetic analysis to be closely related to G. steyaertanum, though some morphological characters vary from the original description of that species.

Puccinia psidii causes a rust disease on a broad range of hosts in the Myrtaceae and Heteropyxidaceae. It is native to South America where it can cause severe disease in eucalypt plantations and other introduced Myrtaceae. The pathogen has recently expanded its geographical range to Hawaii, increasing concerns about the potential for an incursion in Australia. This paper reviews the taxonomy, biology, impact and options for control of P. psidii. It also discusses the probable impact if an incursion were to occur in Australia and the preparations that must be made to mitigate adverse consequences.

Needle fungi in conifers have been extensively studied to explore their diversity, but environmental factors influencing the composition of fungal communities in Pinus radiata D.Don needles have received little attention. This study was conducted to examine the influence of the environment as defined by rainfall, elevation and temperature on the composition of fungal communities in pine needles at an age prior to that at which spring needle cast (SNC) is generally observed. Elucidating the entire fungal community in the needles is a first step towards understanding the cause of the disease. Needle samples were collected from 5-year-old P. radiata trees, their age predating the onset of SNC, from 12 plantations in Tasmania. Interpolated data for the climate variables, including seasonal components for rainfall and temperature, were obtained from an enhanced climate data bank. Pooled needle samples were examined for the fungi they contained using DNA sequencing of cloned polymerase chain reaction (PCR) products. Clones were grouped into operational taxonomic units (OTUs) and identified to their lowest possible taxonomic level by comparison with reference isolates and public DNA databases. DNA sequencing revealed that needle fungal communities differed greatly, depending upon the total annual rainfall and needle age. Needle fungi that have been previously associated with pathogenic behaviour, such as Cyclaneusma minus, Dothistroma septosporum, Lophodermium pinastri, Strasseria geniculata and Sydowia polyspora, were all found in the needles in this study. Also, of 70 ascomycetous OTUs that were discriminated by their DNA sequences, 21 were identified as belonging to the Teratosphaeriaceae. An overview of the endophytic fungi present in the needles of P. radiata has been obtained from trees whose age precedes the onset of SNC. All of the previously known pathogenic species for needle cast diseases were present, in addition to a range of unidentified OTUs belonging to the Teratosphaeriaceae.