We report an annotated draft genome of Heterobasidion occidentale, a fungus (Basidiomycota, Agaricomycetes) that has pathogenic and saprophytic lifestyles. This fungus belongs to the H. annosum (Fr.) Bref. sensu lato species complex that comprises several root rot pathogens. Heterobasidion occidentale causes annosus root and butt rot primarily in true fir (Abies spp.) and spruce (Picea spp.) species throughout western North America.

Heterobasidion annosum sensu lato is a species complex of pathogenic white-rot wood decay fungi which cause root and butt rot in conifer and hardwood species across the Northern hemisphere. Annual losses to forest managers are valued in the billions of dollars, due to tree mortality, reduction in timber yield, and wood decay. In North America, H. irregulare and H. occidentale have a partially overlapping host and geographic range, cause similar disease symptoms and produce similar fruiting bodies, making discrimination between the two of them often difficult. We developed two sets of primers that bind specifically to conserved, but species-specific portions of glyceraldehyde 3-phosphate dehydrogenase and elongation factor 1α alleles. The method is sensitive enough to detect either species from infected wood. Analysis of North American isolates has further clarified the distribution of both species on this continent, including the detection of H. irregulare for the first time on ponderosa pine (Pinus ponderosa) and eastern white pine (Pinus strobus) in British Columbia. This method has the potential to be a valuable tool for the detection of the pathogen in exported/imported wood products, as well as for the further identification and assessment of the distribution of North American Heterobasidion species.

SummaryCharacterization of genes involved in differentiation of pathogen species and isolates with variations of virulence traits provides valuable information to control tree diseases for meeting the challenges of sustainable forest health and phytosanitary trade issues. Lack of genetic knowledge and genomic resources hinders novel gene discovery, molecular mechanism studies and development of diagnostic tools in the management of forest pathogens. Here, we report on transcriptome profiling of Heterobasidion occidentale isolates with contrasting virulence levels. Comparative transcriptomic analysis identified orthologous groups exclusive to H. occidentale and its isolates, revealing biological processes involved in the differentiation of isolates. Further bioinformatics analyses identified an H. occidentale secretome, CYPome and other candidate effectors, from which genes with species‐ and isolate‐specific expression were characterized. A large proportion of differentially expressed genes were revealed to have putative activities as cell wall modification enzymes and transcription factors, suggesting their potential roles in virulence and fungal pathogenesis. Next, large numbers of simple sequence repeats (SSRs) and single nucleotide polymorphisms (SNPs) were detected, including more than 14 000 interisolate non‐synonymous SNPs. These polymorphic loci and species/isolate‐specific genes may contribute to virulence variations and provide ideal DNA markers for development of diagnostic tools and investigation of genetic diversity.

Nine isolates of Phytophthora ramorum Werres, de Cock & Man in't Veld were screened using a variety of systemic and contact fungicides in vitro for mycelial growth inhibition and zoospore germination inhibition, and in planta for suppression of lesion expansion on rhododendron foliage. Three isolates from each of the major clonal lineages, NA1, NA2, and EU1 were used. Systemic fungicides were the most effective at preventing mycelial growth and zoospore germination of P. ramorum, and the results from testing on host plants at the labeled rate supported the in vitro results. Development of resistance to some chemicals used for routine control of P. ramorum in the nursery should be monitored, especially in the EU1 and NA2 populations. Metalaxyl-M had the lowest EC50 for both mycelial growth inhibition and zoospore germination inhibition for all isolates. EC50 was higher for zoospore germination inhibition of the EU1 isolates by two strobilurin fungicides, indicating possible cross-resistance in this group. Crown Copyright (C) 2014 Published by Elsevier Ltd. All rights reserved.

Introduction In southwest Oregon, an aggressive program of cutting and burning host plants in an effort to eradicate Phytophthora ramorum was initiated. It was soon apparent that tanoak (Notholithocarpus densiflorus (Hook. & Arn.) Manos, Cannon & S.H. Oh) resprouts were highly susceptible to P. ramorum and that infected sprouts hamper eradication efforts by maintaining inoculum on site (Hansen and Sutton 2006). The basidiomycete fungus, Chondrostereum purpureum causes a white rot of mostly broadleaf trees and has a wide host range. It invades through fresh wounds in the xylem or cut stumps and is a weak pathogen that can survive as a saprophyte. After the host tree is weakened or killed, C. purpureum is quickly replaced by other, more competitive decay fungi that are naturally occurring in the environment. This fungus is used as a biological control agent for woody vegetation all over the world (Becker et al. 2005, Bourdot et al. 2006). A preparation of mycelium of the fungus C. purpureum is registered under the trade name “ChontrolTM Paste” in the United States and Canada for use as a biological control agent and has been tested as a stump treatment on many hardwood species (EPA Registration No. 74200-1, 2004; and PMRA Registration No. REG. 2004-09, 2004). ChontrolTM is not registered for use in California, so indigenous isolates of C. purpureum are being obtained and formulated for use on California bay laurel (Umbellularia californica (Hook. & Arn.) Nutt.), the host responsible for P. ramorum inoculum buildup in California. In fall 2009, our research team established field trials near Brookings, Oregon to assess the bioherbicidal efficacy of the fungus C. purpureum on tanoak to inhibit resprouts. Chondrostereum purpureum was found occurring naturally on tanoak logs and stumps at other sites in the Brookings area. Laboratory testing of three California isolates of C. purpureum indicate that the fungus can colonize bay laurel stems, and further testing under natural conditions is planned in California. If a formulated product of C. purpureum and/or its mixture with other stem and wood decay fungi applied to tanoak and bay laurel does inhibit the trees from growing new sprouts, this P. ramorum inoculum reservoir would be reduced or eliminated in the ecosystem. In areas where the application of herbicides is not prudent or permitted, this biocontrol treatment would be an indispensable alternative to chemical herbicides.

Phytophthora ramorum is the causal agent of sudden oak death, a very serious disease that causes dieback and death of several important forest species in the USA. It was recently associated with a destructive disease of larch in the United Kingdom. The pathogen has a wide host range including ornamentals such as Rhododendron and Viburnum, and has been recovered from nursery soil samples. Diligent use of effective disinfectants in nurseries can contribute to integrated management strategies for P. ramorum. Nine disinfectants, some at different concentrations and duration, were evaluated in vitro and in vivo for their efficacy in controlling the growth of P. ramorum mycelia and the germination of sporangia. Chemprocide (R) (1.35%), PerCept (TM) (diluted 1:16) and 15% bleach treatments were the most consistent in their effectiveness for inhibiting both mycelia growth and sporangia germination. Chemprocide (R) (1.35%) and 10% bleach treatments were effective also for preventing P. ramorum recovery from contaminated plastic plant saucers and metal surfaces. Isolates representing the three clonal lineages (EU1, NA1, and NA2) of P. ramorum were included in all studies. Interestingly, 10 min treatments of ethanol (70% or 95%) were effective in preventing colony forming unit (CFU) growth, but were not as effective for preventing mycelia growth. Ethanol treatments for 5 min were among the least effective, and in every case was not significantly different from the water-treated controls. Concentration and treatment time were critical factors influencing efficacy in these studies. Differences in CFU production were observed among the clonal lineages with a high of 18 +/- 1.19 (n = 66) for EU1 isolates, and a low of 9.0 +/- 0.89 (n = 74) for NA2 isolates. Crown Copyright (C) 2012 Published by Elsevier Ltd. All rights reserved.

Foliar treatments were applied to potted Rhododendron sp. cv 'Cunningham's white' plants over a 3-week period and evaluated for their ability to protect the leaves from Phytophthora ramorum infection. Treatments included several chemical and biological control products and systemic acquired resistance (SAR) elicitors. Excised leaves from treated plants were challenged by application of P. ramorum sporangia. Our preliminary results indicated that the most effective treatments were the two products registered for prevention of P. ramorum in Canada, Aliette (R) and Subdue Maxx (R). Biocontrol treatments Rhapsody (R) and Actinovate (R) and SAR elicitors 3-aminobutyric acid (BABA) and Actigard (R) conferred a lesser degree of protection to the leaves and lowered the incidence of P. ramorum infection. Treatment with Sonata (R) was not significantly different than that with water. The chemical BABA did not inhibit P. ramorum growth in vitro. Plant trials are ongoing.

A survey was conducted on Vancouver Island and in coastal mainland British Columbia of naturally occurring fungal pathogens of salmonberry (Rubus spectabilis), an endemic plant species considered to be undesirable in conifer regeneration sites. Fungi were recovered from necrotic lesions on foliage and stems and identified to genus level. An in vitro pathogenicity test was performed on 281 fungal isolates using detached salmonberry leaves. Nineteen isolates were found to be moderately to highly aggressive, causing >50% leaf area necrosis within 14 days. These included Botrytis cinerea (17 isolates), Phoma sp. (3 isolates), and Septoria rubi (1 isolate). The three isolates of Phoma, identified as P. argillacea, were selected for further testing on salmonberry plants under greenhouse conditions. Mycelial or conidial inoculum were applied to the foliage of plants, which were incubated for 48 It at 20-22 degrees C and 100% relative humidity (RH) and then grown at 20-22 degrees C and 55-65% RH, with a 16 h photoperiod, under greenhouse conditions. Extensive necrosis of leaves and stems developed within 14 days for all isolates using mycelia and conidia as inoculum. A minimum of 18 h of continuous leaf wetness and mycelial inoculum of 2.6 x 10(3) colony forming units/mL was required for infection to occur. In a small host range test, three economically important conifer species, namely Douglas-fir (Pseudotsuga menziesii), western hemlock (Tsuga heterophylla), and western red cedar (Thuja plicata), showed tolerance at the seedling stage with <5% disease severity. On red raspberry (Rubus idaeus), P argillacea caused some initial damage (leaf spots and chlorosis) but failed to cause cane blight. These results demonstrate the pathogenicity of P. argillacea on salmonberry and the conditions required for establishing disease.

Phytophthora ramorum is an oomycete pathogen and causal agent of a disease commonly referred to as sudden oak death (SOD). The pathogen also causes foliar blight and shoot dieback of nursery plants, including Rhododendron and Viburnum. It is responsible for the widespread mortality of tanoak (Notholithocarpus densiflorus) and coast live oak (Quercus agrifolia) in coastal California and southwestern Oregon, as well as Japanese larch (Larix kaempferi) in the U.K. Thirty-three plant host species commonly found in eastern (8) and western (25) Canadian landscapes and forest sites were selected for this study. Detached leaves/needles were inoculated with P. ramorum EU2 lineage mycelia which was isolated from a stream bait near an infected larch plantation in Scotland, U.K. There was a large variation in aggressiveness and sporulation potential among the evaluated hosts. Among the non-conifer species, the EU2 isolate produced the largest lesions on Pacific dogwood (Cornus nuttallii), Camellia japonica (western species); red oak (Quercus rubra), yellow birch (Betula alleghaniensis), and white ash (Fraxinus americana) (eastern species). For conifer hosts, we found that the EU2 isolate was most aggressive on both balsam fir (Abies balsamea) in the east and grand fir (Abies grandis), western hemlock (Tsuga heterophylla), and western larch (Larix occidentalis) in the west. As for sporulation potential, red alder (Alnus rubra) and bigleaf maple (Acer macrophyllum) in the west produced significantly more sporangia than California bay laurel (Umbellularia californica). Sugar maple (Acer saccharum) in the east was a potential spore producer but not significantly different from California bay laurel. Among the conifer species, western hemlock needles in the west were asymptomatic but produced a small amount of sporangia. The conifer host that produced the most sporangia/mm2 lesion area was white spruce (Picea glauca) in the east and Sitka spruce (Picea sitchensis) in the west, which produced significantly more sporangia than California bay laurel in trial 1 but not in trial 2. Lesion area was biggest on grand fir (Abies grandis), balsam fir, and western larch. These results confirm the potential threat of EU2 lineage of P. ramorum to Canadian flora.