Re-evaluation of Phytophthora citricola isolates from multiple woody hosts in Europe and North America reveals a new species, Phytophthora plurivora sp. nov.

During large-scale surveys for soilborne Phytophthora species in forests and semi-natural stands and nurseries in Europe during the last decade, homothallic Phytophthora isolates with paragynous antheridia, semipapillate persistent sporangia and a growth optimum around 25 °C which did not form catenulate hyphal swellings, were recovered from 39 host species in 16 families. Based on their morphological and physiological characters and the similarity of their ITS DNA sequences with P. citricola as designated on GenBank, these isolates were routinely identified as P. citricola. In this study DNA sequence data from the internal transcribed spacer regions (ITS1 and ITS2) and 5.8S gene of the rRNA operon, the mitochondrial cox1 and β-tubulin genes were used in combination with morphological and physiological characteristics to characterise these isolates and compare them to the ex-type and the authentic type isolates of P. citricola, and two other taxa of the P. citricola complex, P. citricola I and the recently described P. multivora. Due to their unique combination of morphological, physiological and molecular characters these semipapillate homothallic isolates are described here as a new species, P. plurivora sp. nov

Kirramyces destructans in Australia: biosecurity threat or elusive native pathogen?

Kirramyces destructans was first described in 1996 from north Sumatra, Indonesia, where it caused severe leaf and shoot blight on Eucalyptus grandis in nurseries and young plantations. Since then it has been reported in nurseries and plantations in Vietnam, Thailand and China, with its host range extending to include E. camaldulensis and E. urophylla. K. destructans has also been reported from native E. urophylla in East Timor and was considered a significant biosecurity threat to Australia’s native eucalypt forests and plantations. A study on the population diversity of K. destructans isolates throughout south-east Asia in which 8 gene regions were sequenced (four nuclear genes, one mitochondrial gene and three microsatellite markers) detected very low nucleotide polymorphism. This genetic uniformity is indicative of an introduced population which has subsequently spread throughout Asia via human-mediated movement of germplasm. Surveys of sentinel plantings in northern Australia revealed a complex of Kirramyces spp. among which K. destructans was detected. The same gene regions and markers were sequenced as for the Asian study and diversity among the K. destructans isolates in Australia was found to be much greater than that in Asia. We believe that K. destructans is native to Australia where is resides symptomlessly within the native vegetation. The disease is only expressed when non-endemic eucalypts are planted. As such the pathogen is a major encumbrance to the establishment of commercial eucalypt plantations in Northern Australia. The disease has not been observed in native ecosystems, but the effect of inoculum build up within plantations on adjacent native eucalypt remnants is not known

Molecular characterization of natural hybrids formed between five related indigenous clade 6 Phytophthora species

Most Phytophthora hybrids characterized to date have emerged from nurseries and managed landscapes, most likely generated as a consequence of biological invasions associated with the movement of living plants and germplasm for ornamental, horticultural and agricultural purposes. Presented here is evidence for natural hybridization among a group of five closely related indigenous clade 6 Phytophthora species isolated from waterways and riparian ecosystems in Western Australia. Molecular characterization of hybrids consisted of cloning and sequencing two nuclear genes (ITS and ASF), sequencing of two further nuclear loci (BT and HSP) and of two mitochondrial loci (COI and NADH). Additionally, phenotypic traits including morphology of sporangia and optima and maxima temperatures for growth were also determined. In most cases the nuclear genes were biparentally and in all cases the mtDNA were uniparentally inherited, indicating hybrid formation through sexual crosses. Some isolates bear the molecular signature of three parents suggesting additional hybrid events, although it cannot be determined from the data if these were sequential or simultaneous. These species and their hybrids co-exist in riparian ecosystems and waterways where their ability for rapid asexual proliferation would enable them to rapidly colonize green plant litter. The apparent ease of hybridization could eventually lead to the merging of species through introgression. However, at this point in time, species integrity has been maintained and a more likely scenario is that the hybrids are not stable evolutionary lineages, but rather transient hybrid clones

Comparison of primers for the detection of Phytophthora (and other oomycetes) from environmental samples

Many oomycetes are important plant pathogens that cause devastating diseases in agricultural fields, orchards, urban areas, and natural ecosystems. Limitations and difficulties associated with isolating these pathogens have led to a strong uptake of DNA metabarcoding and mass parallel sequencing. At least 21 primer combinations have been designed to amplify oomycetes, or more specifically, Phytophthora species, from environmental samples. We used the Illumina sequencing platform to compare 13 primer combinations on mock communities and environmental samples. The primer combinations tested varied significantly in their ability to amplify Phytophthora species in a mock community and from environmental samples; this was due to either low sensitivity (unable to detect species present in low concentrations) or a lack of specificity (an inability to amplify some species even if they were present in high concentrations). Primers designed for oomycetes underestimated the Phytophthora community compared to Phytophthora-specific primers. We recommend using technical replicates, primer combinations, internal controls, and a phylogenetic approach for assigning a species identity to OTUs or ASVs. Particular care must be taken if sampling substrates where hybrid species could be expected. Overall, the choice of primers should depend upon the hypothesis being tested

An overview of Australia’s Phytophthora species assemblage in natural ecosystems recovered from a survey in Victoria

Although Phytophthora species cause serious diseases worldwide, until recently the main focus on disease in natural ecosystems in southern Australia has been on the distribution and impact of P. cinnamomi. However, new Phytophthora pathogens have emerged from natural ecosystems, and there is a need to better understand the diversity and distribution of these species in our natural forests, woodlands and heathlands. From a survey along a 70 km pipeline easement in Victoria, Phytophthora species were isolated from 249 rhizosphere samples and 25 bait bags deployed in 21 stream, river, or wetland locations. Of the 186 Phytophthora isolates recovered, 130 were identified to species based on ITS sequence data. Ninety-five isolates corresponded to 13 described Phytophthora species while additionally 35 isolates were identified as Clade 6 hybrids. Phytophthora cinnamomi was the most common species isolated (31 %), followed by P. elongata (6 %), both species were only recovered from soil. Samples from sites with the highest soil moisture at the time of sampling had the highest yield of isolates. Consistent with other studies throughout the world, Clade 6 species and their hybrids dominated water samples, although many of these species were also recovered less frequently from soil samples. Many of the species recovered in this study have not previously been reported from eastern Australia, reinforcing that Phytophthora species are widespread, abundant and diverse in natural ecosystems. We have probably been underestimating Phytophthora diversity in Australia

Phytophthora boodjera sp. nov., a damping-off pathogen in production nurseries and from urban and natural landscapes, with an update on the status of P. alticola

A new homothallic Phytophthora species, isolated in Western Australia (WA), is described as Phytophthora boodjera sp. nov. It produces persistent, papillate sporangia, oogonia with thick-walled oospores, and paragynous antheridia. Although morphologically similar to P. arenaria, phylogenetic analyses of the ITS, cox1, HSP90, β-tubulin and enolase gene regions revealed P. boodjera as a new species. In addition, P. boodjera has a higher optimal temperature for growth and a faster growth rate. Phytophthora boodjera has only recently been found in Western Australia and has mostly been isolated from dead and dying Eucalyptus seedlings in nurseries and from urban tree plantings, and occasionally from disturbed natural ecosystems. It is found in association with declining and dying Agonis flexuosa, Banksia media, B. grandis, Corymbia calophylla, Eucalyptus spp,. and Xanthorrhoea preissii. The status of P. alticola was also reviewed. The loss of all isolates associated with the original description except one; discrepancies in both sequence data and morphology of the remaining isolate with that presented the original description, and inconclusive holotype material places the status of this species in doubt

The use of sentinel plantings in forest biosecurity; results from mixed eucalypt species trails in South‐East Asia and Australia

Many diseases of Eucalyptus species have emerged as pathogens in exotic plantations. Guava rust (Puccinia psidii), cryphonectria canker (Crysoporthe cubensis) coniotherium canker (Colletogloeopsis zuluensis) and Kirramyces leaf blight (Kirramyces destructans) are all serious pathogens that have not been found in native forests or in plantations in Australia (Burgess & Wingfield 2002; Cortinas et al. 2006; Glen et al. 2007; Wingfield et al. 2001). The susceptibility to these pathogens of Eucalyptus spp. commonly used in exotic plantations is known; however the susceptibility of many Eucalyptus spp. found only in natural ecosystems in Australia is unknown. There are two main uses of sentinel plantations. Firstly, tree species known to be susceptible to different pathogens can be planted within the natural environment to try and trap pathogens from their surroundings. In Australia, taxa trials planted in different environments act as sentinel plantings. By surveying these taxa trials we have collected and described a number of new eucalypt pathogens and reported the presence in Australia of Kirramyces destructans. The second use for sentinel planting is where many tree species are planted in a region known to harbour certain pathogens. In this manner the susceptibility of the different tree species can be determined

Calcium chelate is as effective as phosphite in controlling Phytophthora root rot in glasshouse trials

Species in the genus Phytophthora cause significant economic losses in crops and damage to forests and natural ecosystems worldwide. Currently, phosphite is the most effective chemical for disease management, but excessive phosphite concentrations can result in phytotoxicity in plants and the development of tolerance by the pathogen. Two newly developed metal chelates and phosphite (alone and in combination) were tested for their in vitro and in planta efficacy against Phytophthora cinnamomi. In glasshouse trials, 0.25% and 0.5% of each chemical treatment (phosphite, Ca chelate, Zn chelate) and Ca chelate + phosphite were used as a foliar application on 3-month-old seedlings of Banksia grandis (experiment not repeated) and Eucalyptus marginata, prior to inoculation with P. cinnamomi. All noninoculated control plants remained healthy, while significant root damage and reduction of dry root weights were observed for inoculated untreated plants. Individually, phosphite and Ca chelate significantly reduced root lesion development of P. cinnamomi compared to the control, with Ca chelate attaining superior results to phosphite at the same concentration. In combination, Ca chelate + phosphite had the largest reduction in root lesion development in both plant species; however, this result has not yet been replicated but did reflect previous in vitro results. The Zn chelate applications were not effective. Ca chelate has the potential to be developed as a fungicide to control Phytophthora species