First Report of Puccinia modiolae Causing Rust Disease on Malva verticillata in Korea.

Malva verticillata (Malvaceae), commonly called Chinese mallow or whorled mallow, is an annual herb native to East Asia and is currently distributed worldwide. In Korea, this plant is cultivated as a leafy vegetable and cooked like spinach or used in soups and also as a medicine material. In March 2022, typical symptoms of rust disease were observed on M. verticillata in a plastic house (37°22'12″ N, 127°34'30" E) in Yeoju, Korea. Yellow or light green round chlorotic spots appeared on the upper surface of infected leaves, while reddish-brown or dark brown rust pustules formed on its lower surface. Infection occurred in 10% of M. verticillata plants surveyed, and disease severity ranged between 30-90%. A representative sample was deposited in the Kunsan National University Herbarium (KSNUH1762). Telia were mostly hypophyllous, reddish-brown to dark brown, round, mostly grouped, and 0.3-0.7 mm in diameter. Teliospores were mostly two-celled, but rarely one or three-celled, yellowish to light brown, fusoid, and 42.9-101 × 10.8- to 18.8 µm (average 72.7 ± 12.3 × 14.2 ± 1.92 µm [mean ± SD]; n = 50), with a smooth, hyaline to yellowish wall of 1.0-2.5 µm thickness. The morphological characteristics were similar to those reported for Puccinia modiolae (Aime and Abbasi 2018; Albu et al. 2019). To confirm the morphological identification, genomic DNA was extracted from the teliospores of an infected leaf. The internal transcribed spacer (ITS) with primers ITS5-u and ITS4rust (Pfunder and Schürch 2001) and the large subunit (LSU) rDNA with primers LRust1R and LRust3 (Beenken et al. 2012) were amplified for sequencing. The resulting sequences were deposited in GenBank with accession numbers ON631218 for ITS and ON631226 for LSU. BLASTn search showed that the Korean sample was identical to the ITS sequences of P. modiolae from Modiola caroliniana (MK458693-MK458697) and the LSU sequences from M. caroliniana, Malva sylvestris, and Alcea rosea (MH742976, MH742977, and MH742978). In the phylogenetic trees of the ITS and LSU sequences, the Korean sample was grouped with the reference sequences of P. modiolae, with the maximum supporting value. For the pathogenicity test, rust-infected leaf discs were placed on the upper or lower surfaces of leaves of three healthy M. verticillata. Three non-inoculated plants served as controls. Inoculated and non-inoculated plants were maintained in a growth chamber at 22°C, a 16/8 h light cycle, and 80% humidity. After three weeks, all inoculated plants developed evident rust symptoms on the upper and lower surfaces of the leaves on which the leaf discs were placed, whereas the control plants remained symptomless. The pathogen present on the inoculated plants was confirmed to be the same pathogen as that observed in the field, fulfilling Koch's postulates. Based on the morphological investigation, sequence analysis, and pathogenicity tests, P. modiolae was identified as the causal agent of rust disease on M. verticillata. To date, this pathogen has been reported on seven Malvaceae plants, including Alcea rosea, Althaea officinalis, Lavatera arborea, Malva parviflora, Malva sylvestris, Modiola caroliniana, and Modiola sp., in North and South America (Farr and Rossman 2022). However, it has not been reported in Asia or Korea. This study is the first report of rust disease on M. verticillata worldwide. Considering its high incidence rate and severe damage, this pathogen is a potential concern for the cultivation of M. verticillata in Korea. This finding could contribute to developing phytosanitary and control treatments for this disease.

First Report of Powdery Mildew Caused by Golovinomyces ambrosiae on Leucanthemum vulgare in Korea.

Leucanthemum vulgare Lam. (Asteraceae), known as ox-eye daisy, is a perennial herb native to Europe and western Asia (Clements et al. 2004, McDougall et al. 2018). In Korea, this plant was introduced for ornamental purposes but has been naturalized as a widespread invasive species. In June 2015, symptoms of a powdery mildew disease were observed on L. vulgare in a public garden in Goseong (38°14'18"N, 128°32'56"E), Korea. Since then, its findings have continued throughout the country, including Mokpo and Seogwipo (in 2018), Hongcheon and Seoul (in 2020), Boeun, Gunsan, and Namwon (in 2022), where the disease incidence was often higher than 80%. Symptoms first appeared as circular to irregular white powdery patches covering leaves and stems. Affected plants became distorted, eventually losing their aesthetic and ornamental value. A total of sixteen samples were deposited in the herbarium of Korea University (KUS-F), Korea. Microscopic observations showed that hyphal appressoria were nipple-shaped. Conidiophores were cylindrical, 98 to 157 × 9 to 12 µm, and produced 2 to 5 immature conidia in chains with a sinuate outline. Foot cells were cylindrical, straight, and 37 to 65 µm long. Conidia were ellipsoid to barrel-shaped, 23 to 39 × 12 to 19 µm, with a length/width ratio of 1.4 to 2.3 and devoid of fibrosin bodies. Germ tubes were produced in the perihilar position of the conidia. Primary conidia were apically rounded and basally subtruncated. No chasmothecia were found until the plants died in winter. The morphological characteristics were typical for anamorph of the genus Golovinomyces. To identify the fungus, genomic DNA was extracted from the four herbarium specimens (KUS-F 28650, 30839, 31728, and 31787). PCR products were amplified using the primer sets PM10/ITS4 for internal transcribed spacer (ITS) and PM3/TW14 for the large subunit (LSU) of the rDNA (Mori et al. 2000, Bradshaw and Tobin 2020). Sequences obtained in the present study were deposited at GenBank (accession numbers ON834488-91 for ITS and ON834494-7 for LSU). A BLASTn search of the Korean specimens showed 100% identity with reference sequences of G. ambrosiae in GenBank (KX98730, MK452580, and MK452588 for ITS and MF612182, MK452653, and MK452661 for LSU). In phylogenetic trees of a concatenated dataset of the ITS and LSU sequences, the Korean specimens formed a well-supported clade with the reference sequences of G. ambrosiae. Pathogenicity tests were carried out by touching and dusting an infected leaf (KUS-F 31787) onto the upper leaf surface of five healthy plants. Five non-inoculated plants served as controls. After two weeks, all inoculated plants formed white patches on the surface of leaves and stems, whereas the control plants remained symptomless. The fungus on the inoculated plants was identical to that observed on the initially diseased plant, fulfilling Koch's postulates. As a result, the causal agent of the powdery mildew on L. vulgare was confirmed as G. ambrosiae (Schwein.) U. Braun & R.T.A. Cook, based on the current taxonomy and nomenclature of this species by Qiu et al. (2020).. Previously powdery mildew collections on L. vulgare have been reported as Golovinomyces cichoracearum (≡ Erysiphe cichoracearum) s. lat. in Estonia, Finland, Germany, and Switzerland, Golovinomyces biocellatus in Spain, and Podosphaera fusca (probably P. xanthii according to the current taxonomy) in the former Soviet Union (now Russia and adjacent countries) (Farr and Rossman 2022). This study is the first report of powdery mildew disease caused by G. ambrosiae on L. vulgare in Korea. Qiu et al. (2020) confirmed the occurrence of G. ambrosiae on L. maximum, another species of the genus Leucanthemum. As powdery mildew causes damage to the cultivation of L. vulgare by loss of ornamental value, appropriate control measures should be developed.

First Report of Coleosporium clematidis Causing Rust Disease on Clematis patens in Korea.

Clematis patens (Ranunculaceae), often called big-flower clematis, is a perennial plant native to Northeast Asia, including China, Japan, and Korea. This plant is one of the popular ornamental plants because of its large and colorful flower. In Korea, it is widely cultivated for public and private gardening and medicinal purposes. In September of 2021, symptoms of rust disease were found on C. patens at a public park (ca. 30 ha) in Jeonju (35°52'16"N, 127°03'16"E), Korea, where the disease occurred on 80% of C. patens plants (n = 50) surveyed, and disease severity in each affected plant ranged 60 to 90%. Symptoms appeared as light green, vein-limited chlorotic spots on the upper surface of infected leaves, and yellow or orange rust pustules were formed on the corresponding lower surface of leaves. A representative sample was deposited in the Kunsan National University Herbarium (KSNUH1522). Uredinia were yellow or orange, round to ellipsoidal, mostly scattered, and 0.5-1 mm in diameter. Urediniospores were pale yellow, ellipsoid or ovoid, 23.1 to 34.8 × 14.9 to 24.7 (average 29.3 ± 2.7 × 18.8 ± 2.2 µm [mean ± SD], n = 50) µm with a verrucose and hyaline wall of 1.0-2.0 µm thick. The morphological characteristics were similar to those reported for Coleosporium clematidis (Barclay 1889, Hiratsuka et al. 1992). To confirm morphological identification, genomic DNA was extracted from a representative specimen (KSNUH1522). The internal transcribed spacer (ITS) rDNA with primers ITS5-u and ITS4rust (Pfunder and Schürch 2001) and large subunit (LSU) rDNA with primers LRust1R and LRust3 (Beenken et al. 2012) were amplified for sequencing. Two resulting sequences (Acc. Nos. OM200310 for ITS, OM184262 for LSU) were blasted in GenBank. The ITS sequence of the Korean sample differs at a nucleotide with a sequence of C. clematidis from Clematis sp. (KX386005) but at eight nucleotides with other three sequences of C. clematidis (KX386007, KX386008 and KX386010). The LSU sequence differs at a nucleotide from the sequences of C. clematidis from Clematis sp. (KX386039, KX386040, KX386042). In phylogenetic trees of the ITS and LSU sequences, the Korean isolate formed a well-supported clade with the reference sequences of C. clematidis. For a pathogenicity test, urediniospores (1.25 ×106/ml) were harvested from the infected leaves and inoculated onto three healthy C. patens. Three non-inoculated plants served as controls. Inoculated and non-inoculated plants were kept in a plant growth chamber at 22°C, a 16/8 h of light cycle, 80% humidity. After three weeks, all inoculated plants formed yellow rust pustules on the lower surface of leaves, identical to what was previously observed in the field, whereas the control plants remained symptomless. The same pathogen was confirmed from the symptomatic plants, fulfilling Koch's postulates. Based on morphological characteristics, sequence data and pathogenicity test, the causal agent of rust on Clematis patens was identified as C. clematidis. To our knowledge, this is the first report of rust disease caused by C. clematidis on C. patens in Korea and previously recorded only in Japan (Hiratsuka et al. 1992). Coleosporium clematidis has been reported on about 60 species of Clematis in Asia and Africa but has not been reported in Europe and North America (Farr and Rossman 2022). In Korea, Clematis fusca var. violacea was previously reported as a host plant for the causal pathogen (Cho and Shin 2004). Given the high occurrence and severe damage, this disease could be a potential threat to the cultivation of C. patens.

Emergence of a New Rust Disease of Virginia Creeper (Parthenocissus quinquefolia) through a Host Range Expansion of Neophysopella vitis.

Virginia creeper (or five-leaved ivy; Parthenocissus quinquefolia) is one of the most popular and widely grown climbers worldwide. In September 2021, Virginia creeper leaves with typical rust symptom were found in an arboretum in Korea, with severe damage. Globally, there is no record of a rust disease on Virginia creeper. Using morphological investigation and molecular phylogenetic inferences, the rust agent was identified as Neophysopella vitis, which is a rust pathogen of other Parthenocissus spp. including Boston ivy (P. tricuspidata). Given that the two ivy plants, Virginia creeper and Boston ivy, have common habitats, especially on buildings and walls, throughout Korea, and that N. vitis is a ubiquitous rust species affecting Boston ivy in Korea, it is speculated that the host range of N. vitis may recently have expanded from Boston ivy to Virginia creeper. The present study reports a globally new rust disease on Virginia creeper, which could be a major threat to the ornamental creeper.

The hop downy mildew pathogen Pseudoperonospora humuli.

Pseudoperonospora humuli is an obligate biotrophic oomycete that causes downy mildew, one of the most devastating diseases of cultivated hop, Humulus lupulus. Downy mildew occurs in all production areas of the crop in the Northern Hemisphere and Argentina. The pathogen overwinters in hop crowns and roots, and causes considerable crop loss. Downy mildew is managed by sanitation practices, planting of resistant cultivars, and fungicide applications. However, the scarcity of sources of host resistance and fungicide resistance in pathogen populations complicates disease management. This review summarizes the current knowledge on the symptoms of the disease, life cycle, virulence factors, and management of hop downy mildew, including various forecasting systems available in the world. Additionally, recent developments in genomics and effector discovery, and the future prospects of using such resources in successful disease management are also discussed. TAXONOMY: Class: Oomycota; Order: Peronosporales; Family: Peronosporaceae; Genus: Pseudoperonospora; Species: Pseudoperonospora humuli. DISEASE SYMPTOMS: The disease is characterized by systemically infected chlorotic shoots called "spikes". Leaf symptoms and signs include angular chlorotic lesions and profuse sporulation on the abaxial side of the leaf. Under severe disease pressure, dark brown discolouration or lesions are observed on cones. Infected crowns have brown to black streaks when cut open. Cultivars highly susceptible to crown rot may die at this phase of the disease cycle without producing shoots. However, foliar symptoms may not be present on plants with systemically infected root systems. INFECTION PROCESS: Pathogen mycelium overwinters in buds and crowns, and emerges on infected shoots in spring. Profuse sporulation occurs on infected tissues and sporangia are released and dispersed by air currents. Under favourable conditions, sporangia germinate and produce biflagellate zoospores that infect healthy tissue, thus perpetuating the infection cycle. Though oospores are produced in infected tissues, their role in the infection cycle is not defined. CONTROL: Downy mildew on hop is managed by a combination of sanitation practices and timely fungicide applications. Forecasting systems are used to time fungicide applications for successful management of the disease. USEFUL WEBSITES: https://content.ces.ncsu.edu/hop-downy-mildew (North Carolina State University disease factsheet), https://www.canr.msu.edu/resources/michigan-hop-management-guide (Michigan Hop Management Guide), http://uspest.org/risk/models (Oregon State University Integrated Plant Protection Center degree-day model for hop downy mildew), https://www.usahops.org/cabinet/data/Field-Guide.pdf (Field Guide for Integrated Pest Management in Hops).

The PhyloCode applied to Cintractiellales, a new order of smut fungi with unresolved phylogenetic relationships in the Ustilaginomycotina.

The PhyloCode is used to classify taxa based on their relation to a most recent common ancestor as recovered from a phylogenetic analysis. We examined the first specimen of Cintractiella (Ustilaginomycotina) collected from Australia and determined its systematic relationship to other Fungi. Three ribosomal DNA loci were analysed both with and without constraint to a phylogenomic hypothesis of the Ustilaginomycotina. Cintractiella did not share a most recent common ancestor with other orders of smut fungi. We used the PhyloCode to define the Cintractiellales, a monogeneric order with four species of Cintractiella, including C. scirpodendri sp. nov. on Scirpodendron ghaeri. The Cintractiellales may have shared a most recent common ancestor with the Malasseziomycetes, but are otherwise unresolved at the rank of class.

Dual RNA-Seq analysis of Medicago truncatula and the pea powdery mildew Erysiphe pisi uncovers distinct host transcriptional signatures during incompatible and compatible interactions and pathogen effector candidates.

Powdery mildew (PM) is a serious fungal disease of legumes. To gain novel insights into PM pathogenesis and host resistance/susceptibility, we used dual RNA-Seq to simultaneously capture host and pathogen transcriptomes at 1 d post-inoculation of resistant and susceptible Medicago truncatula genotypes with the PM Erysiphe pisi (Ep). Differential expression analysis indicates that R-gene mediated resistance against Ep involves extensive transcriptional reprogramming. Functional enrichment of differentially expressed host genes and in silico analysis of co-regulated promoters suggests that amplification of PTI, activation of the JA/ET signaling network, and regulation of growth-defense balance correlate with resistance. In contrast, processes that favor biotrophy, including suppression of defense signaling and programmed cell death, and weaker cell wall defenses are important susceptibility factors. Lastly, Ep effector candidates and genes with known/putative virulence functions were identified, representing a valuable resource that can be leveraged to improve our understanding of legume-PM interactions.

Transcriptomic Profiling of Acute Cold Stress-Induced Disease Resistance (SIDR) Genes and Pathways in the Grapevine Powdery Mildew Pathosystem.

Temperatures from 2 to 8°C transiently induce quantitative resistance to powdery mildew in several host species (cold stress-induced disease resistance [SIDR]). Although cold SIDR events occur in vineyards worldwide an average of 14 to 21 times after budbreak of grapevine and can significantly delay grapevine powdery mildew (Erysiphe necator) epidemics, its molecular basis was poorly understood. We characterized the biology underlying the Vitis vinifera cold SIDR phenotype-which peaks at 24 h post-cold (hpc) treatment and results in a 22 to 28% reduction in spore penetration success-through highly replicated (n = 8 to 10) RNA sequencing experiments. This phenotype was accompanied by a sweeping transcriptional downregulation of photosynthesis-associated pathways whereas starch and sugar metabolism pathways remained largely unaffected, suggesting a transient imbalance in host metabolism and a suboptimal target for pathogen establishment. Twenty-six cold-responsive genes peaked in their differential expression at the 24-hpc time point. Finally, a subset of genes associated with nutrient and amino acid transport accounted for four of the eight most downregulated transcripts, including two nodulin 1A gene precursors, a nodulin MtN21 precursor, and a Dynein light chain 1 motor protein precursor. Reduced transport could exacerbate localized nutrient sinks that would again be transiently suboptimal for pathogen growth. This study links the transient cold SIDR phenotype to underlying transcriptional changes and provides an experimental framework and library of candidate genes to further explore cold SIDR in several systems, with an ultimate goal of identifying novel breeding or management targets for reduced disease.

Effector mining from the Erysiphe pisi haustorial transcriptome identifies novel candidates involved in pea powdery mildew pathogenesis.

Pea powdery mildew (PM) is an important fungal disease caused by an obligate biotroph, Erysiphe pisi (Ep), which significantly impacts pea production worldwide. The phytopathogen secretes a plethora of effectors, primarily through specialized infection structures termed haustoria, to establish a dynamic relationship with its host. To identify Ep effector candidates, a cDNA library of enriched haustoria from Ep-infected pea leaves was sequenced. The Ep transcriptome encodes 622 Ep candidate secreted proteins (CSPs), of which 167 were predicted to be candidate secreted effector proteins (CSEPs). Phylogenetic analysis indicates that Ep CSEPs are highly diverse, but, unlike cereal PM CSEPs, exhibit extensive sequence similarity with effectors from other PMs. Quantitative real-time PCR of a subset of EpCSEP/CSPs revealed that the majority are preferentially expressed in haustoria and exhibit infection stage-specific expression patterns. The functional roles of EpCSEP001, EpCSEP009 and EpCSP083 were probed by host-induced gene silencing (HIGS) via a double-stranded (ds) RNA-mediated RNAi approach. Foliar application of individual EpCSEP/CSP dsRNAs resulted in a marked reduction in PM disease symptoms. These findings were consistent with microscopic and molecular studies, suggesting that these Ep CSEP/CSPs play important roles in pea PM pathogenesis. Homology modelling revealed that EpCSEP001 and EpCSEP009 are analogous to fungal ribonucleases and belong to the RALPH family of effectors. This is the first study to identify and functionally validate candidate effectors from the agriculturally relevant pea PM, and highlights the utility of transcriptomics and HIGS to elucidate the key proteins associated with Ep pathogenesis.

UV-C irradiation compromises conidial germination, formation of appressoria, and induces transcription of three putative photolyase genes in the barley powdery mildew fungus, Blumeria graminis f. sp. hordei.

UV-C irradiation is known to compromise germination of Blumeria graminis conidia and to reduce powdery mildew infestation. However, only scarce information is available on the effects of UV-C irradiation on B. graminis appressorium formation. Applying a Formvar® resin-based in vitro system allowed for analyzing B. graminis germination and appressorium formation in absence of plant defense. UV-C irradiation more strongly affected the differentiation of appressoria than conidial germination. In vivo and in vitro, a single dose of 100 J m-2 UV-C was sufficient to reduce germination to less than 20 % and decrease appressorium formation to values below 5 %. UV-C irradiation negatively affected pustule size and conidiation. White light-mediated photoreactivation was most effective immediately after UV-C irradiation, indicating that a prolonged phase of darkness after UV-C treatment increases the efficacy of B. graminis control. UV-C irradiation increased transcript levels of three putative B. graminis photolyase genes, while mere white light or blue light irradiation did not contribute to the transcriptional up-regulation. Thus, UV-C irradiation effectively controls B. graminis infestation and proliferation by restricting prepenetration processes. Nevertheless, photoreactivation plays an important role in UV-C-based powdery mildew control in crops and hence has to be considered for planning specific irradiation schedules.

Infection assays in Arabidopsis reveal candidate effectors from the poplar rust fungus that promote susceptibility to bacteria and oomycete pathogens.

Fungi of the Pucciniales order cause rust diseases which, altogether, affect thousands of plant species worldwide and pose a major threat to several crops. How rust effectors-virulence proteins delivered into infected tissues to modulate host functions-contribute to pathogen virulence remains poorly understood. Melampsora larici-populina is a devastating and widespread rust pathogen of poplar, and its genome encodes 1184 identified small secreted proteins that could potentially act as effectors. Here, following specific criteria, we selected 16 candidate effector proteins and characterized their virulence activities and subcellular localizations in the leaf cells of Arabidopsis thaliana. Infection assays using bacterial (Pseudomonas syringae) and oomycete (Hyaloperonospora arabidopsidis) pathogens revealed subsets of candidate effectors that enhanced or decreased pathogen leaf colonization. Confocal imaging of green fluorescent protein-tagged candidate effectors constitutively expressed in stable transgenic plants revealed that some protein fusions specifically accumulate in nuclei, chloroplasts, plasmodesmata and punctate cytosolic structures. Altogether, our analysis suggests that rust fungal candidate effectors target distinct cellular components in host cells to promote parasitic growth.

Phylogenetics and Phylogenomics of Rust Fungi.

Rust fungi (Pucciniales) are the most speciose and the most complex group of plant pathogens. Historically, rust taxonomy was largely influenced by host and phenotypic characters, which are potentially plastic. Molecular systematic studies suggest that the extant diversity of this group was largely shaped by host jumps and subsequent shifts. However, it has been challenging to reconstruct the evolutionary history for the order, especially at deeper (family-level) nodes. Phylogenomics offer a potentially powerful tool to reconstruct the Pucciniales tree of life, although researchers working at this vanguard still face unprecedented challenges working with nonculturable organisms that possess some of the largest and most repetitive genomes now known in kingdom fungi. In this chapter, we provide an overview of the current status and special challenges of rust genomics, and we highlight how phylogenomics may provide new perspectives and answer long-standing questions regarding the biology of rust fungi.

A De Novo-Assembly Based Data Analysis Pipeline for Plant Obligate Parasite Metatranscriptomic Studies.

Current and emerging plant diseases caused by obligate parasitic microbes such as rusts, downy mildews, and powdery mildews threaten worldwide crop production and food safety. These obligate parasites are typically unculturable in the laboratory, posing technical challenges to characterize them at the genetic and genomic level. Here we have developed a data analysis pipeline integrating several bioinformatic software programs. This pipeline facilitates rapid gene discovery and expression analysis of a plant host and its obligate parasite simultaneously by next generation sequencing of mixed host and pathogen RNA (i.e., metatranscriptomics). We applied this pipeline to metatranscriptomic sequencing data of sweet basil (Ocimum basilicum) and its obligate downy mildew parasite Peronospora belbahrii, both lacking a sequenced genome. Even with a single data point, we were able to identify both candidate host defense genes and pathogen virulence genes that are highly expressed during infection. This demonstrates the power of this pipeline for identifying genes important in host-pathogen interactions without prior genomic information for either the plant host or the obligate biotrophic pathogen. The simplicity of this pipeline makes it accessible to researchers with limited computational skills and applicable to metatranscriptomic data analysis in a wide range of plant-obligate-parasite systems.

The Poplar Rust-Induced Secreted Protein (RISP) Inhibits the Growth of the Leaf Rust Pathogen Melampsora larici-populina and Triggers Cell Culture Alkalinisation.

Plant cells secrete a wide range of proteins in extracellular spaces in response to pathogen attack. The poplar rust-induced secreted protein (RISP) is a small cationic protein of unknown function that was identified as the most induced gene in poplar leaves during immune responses to the leaf rust pathogen Melampsora larici-populina, an obligate biotrophic parasite. Here, we combined in planta and in vitro molecular biology approaches to tackle the function of RISP. Using a RISP-mCherry fusion transiently expressed in Nicotiana benthamiana leaves, we demonstrated that RISP is secreted into the apoplast. A recombinant RISP specifically binds to M. larici-populina urediniospores and inhibits their germination. It also arrests the growth of the fungus in vitro and on poplar leaves. Interestingly, RISP also triggers poplar cell culture alkalinisation and is cleaved at the C-terminus by a plant-encoded mechanism. Altogether our results indicate that RISP is an antifungal protein that has the ability to trigger cellular responses.

The sunflower downy mildew pathogen Plasmopara halstedii.

Downy mildew of sunflower is caused by Plasmopara halstedii (Farlow) Berlese & de Toni. Plasmopara halstedii is an obligate biotrophic oomycete pathogen that attacks annual Helianthus species and cultivated sunflower, Helianthus annuus. Depending on the sunflower developmental stage at which infection occurs, the characteristic symptoms range from young seedling death, plant dwarfing, leaf bleaching and sporulation to the production of infertile flowers. Downy mildew attacks can have a great economic impact on sunflower crops, and several Pl resistance genes are present in cultivars to protect them against the disease. Nevertheless, some of these resistances have been overcome by the occurrence of novel isolates of the pathogen showing increased virulence. A better characterization of P. halstedii infection and dissemination mechanisms, and the identification of the molecular basis of the interaction with sunflower, is a prerequisite to efficiently fight this pathogen. This review summarizes what is currently known about P. halstedii, provides new insights into its infection cycle on resistant and susceptible sunflower lines using scanning electron and light microscopy imaging, and sheds light on the pathogenicity factors of P. halstedii obtained from recent molecular data. TAXONOMY: Kingdom Stramenopila; Phylum Oomycota; Class Oomycetes; Order Peronosporales; Family Peronosporaceae; Genus Plasmopara; Species Plasmopara halstedii. DISEASE SYMPTOMS: Sunflower seedling damping off, dwarfing of the plant, bleaching of leaves, starting from veins, and visible white sporulation, initially on the lower side of cotyledons and leaves. Plasmopara halstedii infection may severely impact sunflower seed yield. INFECTION PROCESS: In spring, germination of overwintered sexual oospores leads to sunflower root infection. Intercellular hyphae are responsible for systemic plant colonization and the induction of disease symptoms. Under humid and fresh conditions, dissemination structures are produced by the pathogen on all plant organs to release asexual zoosporangia. These zoosporangia play an important role in pathogen dissemination, as they release motile zoospores that are responsible for leaf infections on neighbouring plants. DISEASE CONTROL: Disease control is obtained by both chemical seed treatment (mefenoxam) and the deployment of dominant major resistance genes, denoted Pl. However, the pathogen has developed fungicide resistance and has overcome some plant resistance genes. Research for more sustainable strategies based on the identification of the molecular basis of the interaction are in progress. USEFUL WEBSITES: http://www.heliagene.org/HP, http://lipm-helianthus.toulouse.inra.fr/dokuwiki/doku.php?id=start, https://www.heliagene.org/PlasmoparaSpecies (soon available).

Patterns of genomic variation in the poplar rust fungus Melampsora larici-populina identify pathogenesis-related factors.

Melampsora larici-populina is a fungal pathogen responsible for foliar rust disease on poplar trees, which causes damage to forest plantations worldwide, particularly in Northern Europe. The reference genome of the isolate 98AG31 was previously sequenced using a whole genome shotgun strategy, revealing a large genome of 101 megabases containing 16,399 predicted genes, which included secreted protein genes representing poplar rust candidate effectors. In the present study, the genomes of 15 isolates collected over the past 20 years throughout the French territory, representing distinct virulence profiles, were characterized by massively parallel sequencing to assess genetic variation in the poplar rust fungus. Comparison to the reference genome revealed striking structural variations. Analysis of coverage and sequencing depth identified large missing regions between isolates related to the mating type loci. More than 611,824 single-nucleotide polymorphism (SNP) positions were uncovered overall, indicating a remarkable level of polymorphism. Based on the accumulation of non-synonymous substitutions in coding sequences and the relative frequencies of synonymous and non-synonymous polymorphisms (i.e., PN/PS ), we identify candidate genes that may be involved in fungal pathogenesis. Correlation between non-synonymous SNPs in genes encoding secreted proteins (SPs) and pathotypes of the studied isolates revealed candidate genes potentially related to virulences 1, 6, and 8 of the poplar rust fungus.