Phakopsora myrtacearum sp. nov., a newly described rust (Pucciniales) on eucalypts in eastern and southern Africa

Outbreaks of a rust disease in eucalypt forestry plantations and nurseries in Kenya, Mozambique and South Africa occurred between 2009 and 2014. The pathogen was identified using morphology and molecular phylogenetic analyses as an undescribed species in the Phakopsoraceae. A systematic study, based on nuclear ribosomal DNA, showed that it is a species of Phakopsora, herein named Phakopsora myrtacearum sp. nov. This new species of rust is the second validly described species on Eucalyptus, along with Puccinia psidii. Phakopsora myrtacearum is distinguished from P. psidii by leaf symptoms, morphology of the urediniospores and distinct phylogenetic placement. Phakopsora myrtacearum has been found on three species of Eucalyptus in Kenya, Mozambique and South Africa, and it may have future negative implications for commercial forestry in these areas.     

Evidence that Austropuccinia psidii may complete its sexual life cycle on Myrtaceae

The rust fungus Austropuccinia psidii has spread globally and naturalized in areas with naïve species of Myrtaceae. Previous studies have revealed multiple strains of A. psidii within South America and two strains outside of its native range. The rust spreads by windborne mitotic urediniospores, which are the dominant spore stage. Teliospores and basidiospores of A. psidii are also formed; however, the biological role of these stages in the life cycle is unknown. Experiments presented here tested whether basidiospores of A. psidii could infect Syzygium jambos. The sori produced by infection with basidiospores were screened with five microsatellite markers to confirm whether they were a product of recombination. The findings showed that basidiospores of A. psidii could cause infection on species of Myrtaceae and the resulting sori were a product of recombination. This has important implications for programmes that breed for resistance to this aggressive pathogen in commercial eucalypt forestry.     

Eucalyptus rust caused by Puccinia psidii and the threat it poses to Australia1

Puccinia psidii, which causes the disease Eucalyptus rust, poses a threat to biodiversity in Australia and the Eucalyptus forest industry worldwide. It is native to South America and Central America and has spread to North America (Mexico, USA – Florida). In mid‐2005, the rust was reported in Hawaii, USA, which means it is now present in the Pacific region.     

Disease cycle of Austropuccinia psidii on Eucalyptus globulus and Eucalyptus obliqua leaves of different rust response phenotypes

Myrtle rust poses a significant biosecurity threat to Australia with potential for long-term damaging impacts on native flora and plant industries. This study describes the disease cycle of Austropuccinia psidii, the myrtle rust pathogen, in Eucalyptus globulus and Eucalyptus obliqua, two commercially and ecologically important species from different subgenera of Eucalyptus. Ontogeny and morphology of infection structures of A. psidii on plants of both Eucalyptus species with different rust response phenotypes, i.e. completely resistant, hypersensitive and highly susceptible, were investigated. Plants were inoculated with single-uredinium-derived urediniospores and examined by scanning electron microscopy. No differences between rust response phenotypes were observed in germination of urediniospores, formation of appressoria or length of germ tubes. The growth of germ tubes had no affinity towards stomata of either species. Histological observations indicated direct penetration by infection pegs through the leaf cuticle and no penetration beyond the epidermis on rust-resistant E. obliqua. Eucalyptus obliqua plants that were identified as susceptible to A. psidii at 3- and 6-months-old showed no disease when reinoculated with A. psidii at 12-months-old; this indicated possible early acquisition of adult plant resistance to A. psidii in this species. In the susceptible phenotype of E. globulus rust inoculation led to rapid colonization of leaf parenchyma cells with the disease cycle completed within 10 days. These findings provide valuable insights into host–pathogen interactions in the Eucalyptus–A. psidii pathosystem, which might be useful for the development of effective rust control strategies across Eucalyptus subgenera.     

Screening Corymbia populations for resistance to Puccinia psidii

The exotic rust pathogen Puccinia psidii is now widespread along the east coast of Australia from temperate Victoria to tropical far north Queensland, with a current host range exceeding 200 species from 37 myrtaceous genera. To determine the threat P. psidii poses to plantation and native eucalypts, artificial inoculation was used to screen germplasm of spotted gum (Corymbia spp.) for resistance to the biotype of P. psidii that has become established in Australia. The objective was to characterize resistance to P. psidii within the Corymbia species complex so that management strategies for the deployment of germplasm from existing breeding programmes of these spotted gum species could be developed. Symptom development initiated 7 days after inoculation, with resistant and susceptible seedlings identified within all species, provenances and families. Inter‐ and intraspecific variability in rust resistance was observed among spotted gum species. There was no apparent relationship between climatic conditions at the provenance origin and disease resistance. The heritability estimates for all assessments are moderate to high and indicate a significant level of additive genetic variance for rust resistance within the populations. The results of this study clearly identify potential to select for resistance at the family level within the tested populations. While the potential for P. psidii to detrimentally impact upon Corymbia in the nursery and in young plantations was demonstrated, estimations of the heritability of resistance suggest that efforts to enhance this trait through breeding have reasonable prospects for success.     

Susceptibility of native New Zealand Myrtaceae to the South African strain of Austropuccinia psidii: A biosecurity threat

Austropuccinia psidii, cause of myrtle rust, has spread globally where Myrtaceae occur. Multiple strains of A. psidii have been identified, including a unique strain found only in South Africa. The South African strain is a biosecurity concern for species of Myrtaceae worldwide. This is because preliminary testing of South African Myrtaceae suggests it could have a wide host range, and thus has the potential to be invasive. In this study, we assessed the ability of the South African strain to infect other species of Myrtaceae by testing the susceptibility of New Zealand provenance Myrtaceae. Seedlings of four native New Zealand Myrtaceae species (Metrosideros excelsa, Leptospermum scoparium, Kunzea robusta, and Kunzea linearis) were artificially inoculated in South Africa with a single-uredinium isolate of the South African strain. Fourteen days after inoculation, uredinia, and in many cases telia, had developed on the young leaves and stems of all four host species, which led to shoot tip dieback in the more severe cases. When comparisons were made between the species, K. robusta was the least susceptible to the South African strain of A. psidii, while L. scoparium and M. excelsa were the most susceptible. While only a limited number of seed families were tested, only a small proportion of the seedlings showed resistance to infection by the South African strain. This preliminary testing highlights the potential invasive risk the South African strain poses to global Myrtaceae communities, including New Zealand Myrtaceae.     

Simple sequence repeat markers support the presence of a single genotype of Puccinia psidii in Australia

A non‐native rust of Myrtaceae was first detected in Australia in 2010, and was later identified as Puccinia psidii. The presence of many native species of Myrtaceae and a lack of understanding of genetic variability in P. psidii in Australia led to the current study. Low coverage genome sequencing of P. psidii suggested a genome size of c. 142 Mb. A set of 240 simple sequence repeat (SSR) primers was designed based on the genome sequence information generated. Seventeen isolates of P. psidii comprising 14 from Australia, two from Brazil and one from Hawaii were selected to study genetic variation in the pathogen. Out of 240 initially screened markers, 74% showed amplification among P. psidii isolates and 38% were polymorphic. Primers were fluorescently labelled and genotyping revealed three distinct genotypes among the isolates: one comprising Australian isolates and an isolate from Hawaii, and the second and third comprising two Brazilian isolates. Locus USYD_Pp151 produced a fourth genotype for the Hawaiian isolate of P. psidii. Markers revealed that all Australian isolates were genetically similar to the one from Hawaii. There was no genetic variation among the Australian isolates of P. psidii, supporting the hypothesis that only one genotype of P. psidii was introduced into Australia. The SSR markers developed in this study are highly specific to P. psidii and can be used confidently as a new profiling tool to monitor evolution of P. psidii in Australia and elsewhere.     

Armillaria root rot spreading into a natural woody ecosystem in South Africa

Signs and symptoms of a disease similar to those of armillaria root rot have recently been observed on various native woody plants on the foothills of Table Mountain in South Africa, one of the most botanically diverse natural environments globally. This is of concern because the root rot fungus Armillaria mellea has previously been shown to be an alien pathogen of European origin in planted gardens in the City of Cape Town. An aim of this study was to identify the cause of the root rot disease on infected plants. Based on DNA‐sequence phylogeny, it was shown that isolates collected from at least 16 native tree and woody shrub species represented the non‐native A. mellea. Microsatellite markers were then used to determine the genetic diversity and population structure of the A. mellea isolates from Table Mountain and two planted gardens where the pathogen has previously been found. Population genetic analyses revealed low levels of gene diversity and no population differentiation amongst the three populations. The results provide the first firm evidence that A. mellea has escaped the planted environment and invaded a sensitive and ecologically important natural woody environment in South Africa. This is only the second definitive case of a non‐native tree pathogen invading a natural ecosystem in the country.     

Host switching between native and non‐native trees in a population of the canker pathogen Chrysoporthe cubensis from Colombia

The purpose of this study was to test the hypothesis that Chrysoporthe cubensis on native trees in South America could be the source of the pathogen that causes severe stem cankers and often mortality in commercially propagated Eucalyptus trees. This was done by investigating populations originating from two adjacent Eucalyptus (Myrtaceae) plantations in Colombia, and wild Miconia rubiginosa trees (Melastomataceae) growing alongside these stands. Polymorphic microsatellite markers were used to quantify allele sizes in 20 and 39 isolates from the two Eucalyptus stands and 32 isolates from adjacent M. rubiginosa trees. Gene and genotypic diversities were calculated from these data, and population differentiation and assignment tests were performed to ascertain whether the populations were genetically different. Results showed that there were no differences between any of the populations using these techniques, and that they can be treated as a single population. Therefore, the results support the hypothesis that host switching has occurred in C. cubensis in Colombia.     

Description and molecular phylogeny of <Emphasis Type="Italic">Ditylenchus gilanicus</Emphasis> n. sp. (Nematoda: Anguinidae) from northern forests of Iran

Commercial areas containing Eucalyptus plantations have expanded in recent years due to increased demands for pulp, paper and bioenergy. One of the threats that can reduce Eucalyptus production is the eucalyptus rust disease caused by Austropuccinia psidii, a biotrophic fungus that affects a broad range of Myrtaceae. An accurate diagnosis tool for the early detection of rust disease could be useful in breeding programs for selection of resistant plants against rust, in phytosanitary purposes or in rust epidemics studies. The aim of the present work was to develop a SYBR Green-based quantitative real-time PCR (qPCR) assay for the early detection and quantification of A. psidii in Eucalyptus grandis leaves. Three sets of primers based on the A. psidii ribosomal DNA intergenic space region (IGS), beta-tubulin and elongation factor genes were designed and evaluated. The assays using the IGS primer set resulted in the highest detection efficiency, detecting a lower limit of 0.5 pg of A. psidii DNA. Under artificial inoculation in plants, A. psidii was detected immediately after pathogen inoculation until 240 h post-inoculation using qPCR. In field validation of the method, A. psidii was detected using qPCR in naturally infected leaves with or without rust symptoms. This easy and fast method can be used for an efficient detection of A. psidii in E. grandis leaves. The implications of this tool for rust studies are discussed below.     

Projecting the suitability of global and local habitats for myrtle rust (Austropuccinia psidii) using model consensus

Myrtle rust (caused by Austropuccinia psidii) affects more than 500 known host species in the Myrtaceae family. Three different modelling approaches (CLIMEX, MaxEnt and Multi-Model Framework) were used to project the habitat suitability for myrtle rust at both global and local scales. Current data on the global occurrence of myrtle rust were collected from online literature and expert solicitation. Long-term averages of climate data (1960–1990) were sourced from WorldClim and CliMond websites. Recent reports of myrtle rust in New Zealand were used for validation of model outputs but not in model training and testing. The model outputs were combined into a consensus model to identify localities projected to be suitable for myrtle rust according to two or three models (hotspots). In addition to the locations where the pathogen is currently present, all models successfully projected independent occurrence data in New Zealand suitable for establishment of the pathogen. Climate suitability for the pathogen was primarily related to temperature followed by rainfall in MaxEnt and the CLIMEX model. The results confirmed the optimum temperature range of this pathogen in the literature (15–25 °C). Additional analysis of the precipitation variables indicated that excessive rain (more than 2000 mm in warmest quarter of the year) combined with high temperatures (>30 °C) constrain pathogen establishment. The results of the current study can be useful for countries such as New Zealand, China, South Africa and Singapore where the pathogen has not fully spread or established.     

Puccinia psidii in Queensland,Australia: disease symptoms,distribution and impact

Puccinia psidii has long been considered a significant threat to Australian plant industries and ecosystems. In April 2010, P. psidii was detected for the first time in Australia on the central coast of New South Wales (NSW). The fungus spread rapidly along the east coast and in December 2010 was found in Queensland (Qld) followed by Victoria a year later. Puccinia psidii was initially restricted to the southeastern part of Qld but spread as far north as Mossman. In Qld, 48 species of Myrtaceae are considered highly or extremely susceptible to the disease. The impact of P. psidii on individual trees and shrubs has ranged from minor leaf spots, foliage, stem and branch dieback to reduced fecundity. Tree death, as a result of repeated infection, has been recorded for Rhodomyrtus psidioides. Rust infection has also been recorded on flower buds, flowers and fruits of 28 host species. Morphological and molecular characteristics were used to confirm the identification of P. psidii from a range of Myrtaceae in Qld and compared with isolates from NSW and overseas. A reconstructed phylogeny based on the LSU and SSU regions of rDNA did not resolve the familial placement of P. psidii, but indicated that it does not belong to the Pucciniaceae. Uredo rangelii was found to be con‐specific with all isolates of P. psidii in morphology, ITS and LSU sequence data, and host range.     

Dothistroma needle blight: an emerging epidemic caused by Dothistroma septosporum in Colombia

Plantation forestry in Colombia is based mainly on non‐native species of Pinus and Eucalyptus. Since 2008, a disease with symptoms similar to those of dothistroma needle blight (DNB) has been found affecting large areas planted to Pinus spp. The aim of this study was to identify the causal pathogen as well as to document the levels of disease incidence and severity. Isolates from each of three forestry zones, collected from different host species, were compared based on rDNA sequence of the ITS regions. These were conclusively identified as Dothistroma septosporum, one of two Dothistroma spp. known to cause DNB. Susceptibility was greatest on low elevation Pinus tecunumanii followed by Pinus kesiya and Pinus oocarpa. Pinus maximinoi and high elevation P. tecunumanii showed tolerance to D. septosporum. The disease incidence in the different zones varied significantly with the North zone being the most severely affected. This constitutes the first report of disease distribution and susceptibility of hosts, as well as the first consideration of the relative importance of D. septosporum in Colombia.     

Detecting myrtle rust (Austropuccinia psidii) on lemon myrtle trees using spectral signatures and machine learning

Hundreds of species in one of Australia's dominant plant families, the Myrtaceae, are at risk from the invasive pathogenic fungus Austropuccinia psidii. Since its arrival in Australia in 2010, native plant communities have been severely affected, with highly susceptible species likely to become extinct from recurring infections. While severe impact on Australian native and plantation forestry has been predicted, the lemon myrtle industry is already under threat. Commercial cultivars of lemon myrtle (Backhousia citriodora) are highly susceptible to A. psidii. Detecting and monitoring disease outbreaks is currently only possible by eye, which is costly and subject to human bias. This study aims at developing a proof‐of‐concept for automated, non‐biased classification of healthy (naïve), fungicide‐treated and diseased lemon myrtle trees by means of their spectral reflectance signatures. From a lemon myrtle plantation, spectral signatures of fungicide‐treated and untreated leaves were collected using a portable field spectrometer. A third class of spectra, from naïve lemon myrtle leaves that had not been exposed to A. psidii, was collected from a botanical garden. Reflectance spectra in their primary form and their first‐order derivatives were used to train a random forest classifier resulting in an overall accuracy of 78% (kappa = 0.68) for primary spectra and 95% (kappa = 0.92) for first‐order derivative‐transformed spectra. Thus, an optical sensor‐based discrimination, using spectral reflectance signatures of this as yet uninvestigated pathosystem, seems technically feasible. This study provides a foundation for the development of automated, sensor‐based detection and monitoring systems for myrtle rust.     

Glyphosate reduces urediniospore development and Puccinia psidii disease severity on Eucalyptus grandis

BACKGROUND: Recent studies have shown the effects of glyphosate drift on decreasing rust intensity on Eucalyptus grandis plants. However, the effects of the herbicide on Puccinia psidii initial development are unknown. In this study the systemic action of glyphosate on rust severity was evaluated on Eucalyptus plants maintained under greenhouse conditions. Urediniospore germination and apressorium formation on detached leaves and on water agar medium, previously treated with glyphosate, were also evaluated. RESULTS: Rust severity and the number of urediniospores per leaf area were significantly reduced with increasing glyphosate doses, even on branches not directly treated with the herbicide, indicating a systemic effect of glyphosate on pathogen development. Similarly, higher glyphosate doses also reduced germination and apressorium formation on detached Eucalyptus leaves, regardless of the direct application of the product on the leaf limb or on the petiole base. Puccinia psidii urediniospore germination in water agar medium also decreased with increasing herbicide doses. CONCLUSIONS: Reductions in germination and apressorium formation of P. psidii urediniospores with increasing glyphosate dose indicate that a lower severity and intensity of the disease may perhaps be due to blockage of the shikimic acid pathway in the fungal metabolic system. Copyright © 2011 Society of Chemical Industry     

First release of a fungal classical biocontrol agent against an invasive alien weed in Europe: biology of the rust,Puccinia komarovii var. glanduliferae

The rust fungus Puccinia komarovii var. glanduliferae was first identified infecting Impatiens glandulifera in its native range (western Himalayas) between 2006 and 2010. Subsequently, it was imported into quarantine in the UK for evaluation as a classical biocontrol agent. To assess the safety of the rust, plant species relevant to Europe were tested for susceptibility. To confirm the life cycle, all infective spore stages were inoculated on I. glandulifera to follow disease progression. Teliospores were primed using bleaching and low temperatures to break dormancy. Temperature and dew period experiments using urediniospores were conducted to assess the parameters required for infection. Of the 74 plant species tested, only I. balsamina, an ornamental species, was fully susceptible to urediniospore inoculum. The life cycle of the rust – an autoecious, full‐cycled species with five spore stages – was confirmed. Urediniospores were infective between 5 and 25°C, with an optimum at 15°C. A minimum of 8 h dew period was required to achieve consistent infection. Based on a pest risk assessment, the rust poses no threat to native biodiversity within EU Member States; making P. komarovii var. glanduliferae a suitable candidate as the first fungal classical biocontrol agent against an exotic weed in the region.     

Age‐related susceptibility of Eucalyptus species to Phytophthora boodjera

Phytophthora boodjera is a newly described pathogen causing damping off and mortality of Eucalyptus seedlings in Western Australian nurseries. This study evaluated the age‐related susceptibility of several taxa of mallee Eucalyptus to P. boodjera in sterilized washed river sand‐infestation pot trials. Phytophthora cinnamomi and P. arenaria were included for comparison. Seedlings of Eucalyptus taxa were inoculated at 0, 2, 4, 12 and 88 weeks with individual Phytophthora isolates. Pre‐emergent mortality in the presence of Phytophthora was almost 100%. Post‐emergent mortality was 50–100%, depending on isolate, compared to 0% for the control. Mortality was also high for inoculated 1 month‐old seedlings (46–68%) and root length of surviving seedlings was severely reduced. Death from root infection was not observed for seedlings inoculated at 12 and 88 weeks, but they developed root necrosis and reduced root dry weight compared to non‐inoculated controls. Phytophthora boodjera is a pre‐ and post‐emergent pathogen of mallee eucalypts. These eucalypts are susceptible to P. boodjera at all life stages tested, but the mortality rates declined with plant age. Similar results were obtained for P. cinnamomi and P. arenaria. The events leading to its recent appearance in the nurseries remain unknown and further investigations are underway to determine if this is an introduced or endemic pathogen. The approach used here to understand the impact of a Phytophthora species on multiple hosts at different seedling ages is novel and sets a benchmark for future work.     

Characterization of Botryosphaeriaceae from plantation‐grown Eucalyptus species in South China

The Botryosphaeriaceae is a species‐rich family that includes pathogens of a wide variety of trees, including Eucalyptus species. Symptoms typical of infection by the Botryosphaeriaceae have recently been observed in Eucalyptus plantations in South China. The aim of this study was to identify the Botryosphaeriaceae associated with these symptoms. Isolates were collected from branch cankers and senescent twigs of different Eucalyptus spp. All isolates resembling Botryosphaeriaceae were separated into groups based on conidial morphology. Initial identifications were made using PCR‐RFLP fingerprinting, by digesting the ITS region of the rDNA operon with the restriction enzymes CfoI and KspI. Furthermore, to distinguish isolates in the Neofusicoccum parvum/N. ribis complex, a locus (BotF15) previously shown to define these species, was amplified and restricted with CfoI. Selected isolates were then identified using comparisons of DNA sequence data for the ITS rDNA and translation elongation factor 1‐alpha (TEF‐1α) gene regions. Based on anamorph morphology and DNA sequence comparisons, five species were identified: Lasiodiplodia pseudotheobromae, L. theobromae, Neofusicoccum parvum, N. ribis sensu lato and one undescribed taxon, for which the name Fusicoccum fabicercianum sp. nov. is provided. Isolates of all species gave rise to lesions on the stems of an E. grandis clone in a glasshouse inoculation trial and on the stems of five Eucalyptus genotypes inoculated in the field, where L. pseudotheobromae and L. theobromae were most pathogenic. The five Eucalyptus genotypes differed in their susceptibility to the Botryosphaeriaceae species suggesting that breeding and selection offers opportunity for disease avoidance in the future.     

First report of rust disease on ohia and the causal fungus, <Emphasis Type="Italic">Puccinia psidii</Emphasis>, in Japan

In 2007, a rust disease of ohia (Metrosideros polymorpha) was found in Japan. We identified this pathogen as Puccinia psidii based on its morphological characters and the internal transcribed spacer (ITS) sequence of the ribosomal DNA. This pathogen was pathogenic on Eucalyptus amplifolia and E. rudis in addition to previously reported host species. This is the first report in Japan of a rust disease on ohia and the occurrence of its causal fungus, P. psidii.