Diversity, migration routes, and worldwide population genetic structure of Lecanosticta acicola, the causal agent of brown spot needle blight.

Lecanosticta acicola is a pine needle pathogen causing brown spot needle blight that results in premature needle shedding with considerable damage described in North America, Europe, and Asia. Microsatellite and mating type markers were used to study the population genetics, migration history, and reproduction mode of the pathogen, based on a collection of 650 isolates from 27 countries and 26 hosts across the range of L. acicola. The presence of L. acicola in Georgia was confirmed in this study. Migration analyses indicate there have been several introduction events from North America into Europe. However, some of the source populations still appear to remain unknown. The populations in Croatia and western Asia appear to originate from genetically similar populations in North America. Intercontinental movement of the pathogen was reflected in an identical haplotype occurring on two continents, in North America (Canada) and Europe (Germany). Several shared haplotypes between European populations further suggests more local pathogen movement between countries. Moreover, migration analyses indicate that the populations in northern Europe originate from more established populations in central Europe. Overall, the highest genetic diversity was observed in south-eastern USA. In Europe, the highest diversity was observed in France, where the presence of both known pathogen lineages was recorded. Less than half of the observed populations contained mating types in equal proportions. Although there is evidence of some sexual reproduction taking place, the pathogen spreads predominantly asexually and through anthropogenic activity.

Occurrence and pathogenicity of Corinectria spp. - an emerging canker disease of Abies sibirica in Central Siberia.

During recent years, a new disease of Siberian fir (A. sibirica) emerged in Central Siberia, exhibiting symptoms of stem/branch deformation, cambium necrosis, and dieback of branches and twigs, the causal agent remaining unknown. The aim was to identify agent of the disease and to investigate its pathogenicity to A. sibirica and Norway spruce (Picea abies). Symptomatic tissues of fir were subjected to pure culture isolation of anticipated pathogen(s). Obtained isolates were subjected to molecular identification, phylogenetic analyses, and pathogenicity tests with A. sibirica saplings, and seeds and seedlings of A. sibirica and P. abies. The study demonstrated that, (i) most commonly isolated fungus from canker wounds of A. sibirica exhibited Acremonium-like anamorphs; (ii) phylogeny demonstrated that investigated fungi belong to genus Corinectria, but are genetically well separated from other worldwide known Corinectria spp.; (iii) one species of isolated fungi has the capacity to cause the disease and kill A. sibirica saplings and seedlings, but also seedlings of P. abies. Guidelines for future research were defined in order to generate needed information on species description, its origin and ecology, and estimation of potential risks upon the eventual invasion of the pathogen to new geographic areas, in particular of Europe.

Genetic Analyses Suggest Separate Introductions of the Pine Pathogen Lecanosticta acicola Into Europe.

Lecanosticta acicola is a heterothallic ascomycete that causes brown spot needle blight on native and nonnative Pinus spp. in many regions of the world. In this study we investigated the origin of European L. acicola populations and estimated the level of random mating of the pathogen in affected areas. Part of the elongation factor 1-α gene was sequenced, 11 microsatellite regions were screened, and the mating type idiomorphs were determined for 201 isolates of L. acicola collected from three continents and 17 host species. The isolates from Mexico and Guatemala were unique, highly diverse and could represent cryptic species of Lecanosticta. The isolates from East Asia formed a uniform and discrete group. Two distinct populations were identified in both North America and Europe. Approximate Bayesian computation analyses strongly suggest independent introductions of two populations from North America into Europe. Microsatellite data and mating type distributions indicated random recombination in the populations of North America and Europe. Its intercontinental introduction can most likely be explained as a consequence of the movement of infected plant material. In contrast, the spread of L. acicola within Europe appears to be primarily due to conidial dispersion and probably also ascospore dissemination.

Description and prevalence of a putative novel mycovirus within the conifer pathogen Gremmeniella abietina.

The European race of Gremmeniella abietina (Lagerberg) Morelet is the causal agent of stem canker and shoot blight on numerous conifers in Europe and North America. It comprises different species and biotypes in which the presence of mycoviruses has been determined. In this report, we describe the full-length sequence of the RNA-dependent RNA polymerase (RdRp) of a putative novel virus, Gremmeniella abietina RNA virus 6 (GaRV6), with 2165 nt and a GC content of 54.7 %. A BLASTp search using the deduced RdRp amino acid sequence confirmed GaRV6 to be related to members of a still unassigned virus taxon, which includes, e.g., Fusarium graminearum dsRNA mycovirus 4 (FgV-4) and the mutualistic Curvularia thermal tolerance virus (CThTV). The prevalence and genetic diversity of GaRV6 was also studied within the European race of G. abietina. We examined 162 isolates originating from Canada, the Czech Republic, Finland, Italy, Montenegro, Serbia, Spain, Switzerland, Turkey and the United States. According to direct specific reverse transcription (RT) PCR screening based on the RdRp sequence, the virus appears to be present only in Spain, where it is relatively abundant but genetically highly uniform.

The European race of Gremmeniella abietina hosts a single species of Gammapartitivirus showing a global distribution and possible recombinant events in its history.

The population genetics of the family Partitiviridae was studied within the European race of the conifer pathogen Gremmeniella abietina. One hundred sixty-two isolates were collected from different countries, including Canada, the Czech Republic, Finland, Italy, Montenegro, Serbia, Spain, Switzerland, Turkey and the United States. A unique species of G. abietina RNA virus-MS1 (GaRV-MS1) appears to occur indistinctly in G. abietina biotypes A and B, without a particular geographical distribution pattern. Forty-six isolates were shown to host GaRV-MS1 according to direct specific RT-PCR screening, and the virus was more common in biotype A than B. Phylogenetic analysis based on 46 partial coat protein (CP) cDNA sequences divided the GaRV-MS1 population into two closely related clades, while RNA-dependent RNA polymerase (RdRp) sequences revealed only one clade. The evolution of the virus appears to mainly occur through purifying selection but also through recombination. Recombination events were detected within alignments of the three complete CP and RdRp sequences of GaRV-MS1. This is the first time that recombination events have been directly identified in fungal partitiviruses and in G. abietina in particular. The results suggest that the population dynamics of GaRV-MS1 do not have a direct impact on the genetic structure of its host, G. abietina, though they might have had an innocuous ancestral relationship.

Genetic variation and relationships in Laetiporus sulphureus s. lat., as determined by ITS rDNA sequences and in vitro growth rate.

The aim of this study was to characterise the genetic variation and molecular relationships of the brown rot polypore, Laetiporus sulphureus s. lat., from Europe, South America, Africa, and Asia, using ITS sequences of the nu-rDNA and by comparing the growth rate in vitro. In a NJ analysis of the sequences of 130 individuals of L. sulphureus s. lat., eight distinct clusters emerged, supported by BS values of 70-100%. Within each cluster, the ITS rDNA sequence variation was below 3%. The sequences were also analysed together with Laetiporus sequences available from GenBank. Results demonstrated the possible presence of L. huroniensis in Europe (invalidly named L. montanus) and L. gilbertsonii in South America, and provided more information on the Pan-American and European distribution of one of the clades, currently known in North America as L. sulphureus. L. conifericola formed a separate distinct clade. Moreover, the analysis revealed two unknown Laetiporus taxa in Korea, one in South Africa, and one in Europe. As L. sulphureus is described from Europe (France), and we show that more than one taxon exist here, it is presently not possible to define L. sulphureus s. str. Certain biological differences between some of the clades (in vitro growth rates, chemical composition, and pigmentation) were demonstrated and discussed.

Variations in rDNA ITS of Czech Armillaria species determined by PCR and HPLC.

We analysed 40 isolates of species Armillaria. borealis, A. cepistipes, A. gallica, A. mellea, A. ostoyae and A. tabescens, mostly collected in the Czech Republic, by PCR-RFLP of the ITS rRNA genes using the restriction endonucleases AluI, HinfI and MboI. Restriction fragments were analysed by ion-exchange high performance liquid chromatography which proved to be more useful informative, and less time-consuming than classical electrophoresis on agarose gel. The HPLC method enabled detection of some heterozygous strains. HinfI discriminated between all six species. Ten isolates were sequenced to confirm changes in restriction sites found by restriction analysis. Cluster analysis based on the restrictions patterns of restriction endonucleases AluI and HinfI divided the analysed species into three groups. The first and the most distant group contained all A. mellea isolates, the second group was formed by A. tabescens and the third group contained species A. borealis, A. cepistipes, A. gallica and A. ostoyae. The A. tabescens group was very homogenous regardless of the origin of isolates (Czech Republic, France and Finland).