A study was carried out to clarify the role of the fungus Chalara fraxinea in decline of Fraxinus excelsior , which is observed on a large scale in central and northern Europe with high incidence of tree mortality. The aims of this work were: (i) to check for the presence of C. fraxinea in various tissues of declining F. excelsior by agar culture isolations and by direct analysis of plant tissues using molecular techniques (DNA extraction, ITS-PCR, cloning, ITS sequencing and T-RFLP); (ii) to study fungal communities inhabiting tissues with symptoms; and (iii) to test the pathogenicity of C. fraxinea to F. excelsior . Chalara fraxinea was isolated from 93% of stem cankers, 91% of necrotic leaf stalks, 27-28% of bark wounds and 30% of visually healthy leaf stalks. Molecular analyses of necrotic leaves, leaf stalks and bark revealed the presence of 25 different fungal taxa, 14 of which were detected in all three types of tissue sample. Chalara fraxinea was the second most common species (61% of samples), and only Cryptococcus foliicola occurred more often (70%). All eight of the tested C. fraxinea isolates induced necroses in bark and cambium on each of 86 inoculated trees, and all controls remained healthy. Average length of necroses caused by different C. fraxinea strains varied from 4·2 to 8·9 cm, but the differences were statistically insignificant. Instead, differences in resistance of individual trees to C. fraxinea were observed.

Currently, massive dieback of Fraxinus excelsior is observed in countries of eastern, northern and central Europe, and the reasons for it are unclear. The aims of the present work were (a) to study fungal communities in declining F. excelsior crowns; (b) to clarify role of fungi in the decline. Shoots from symptomatic crowns were collected in four localities in central Sweden, and distributed into the following categories: (a) visually healthy; (b) initial necroses; (c) advanced necroses; (c) dead tops. The most frequently isolated fungi were Gibberella avenacea, Alternaria alternata, Epicoccum nigrum, Botryosphaeria stevensii, Valsa sp., Lewia sp., Aureobasidium pullulans and Phomopsis sp., and these taxa were consistently found in shoots of all four symptomatic categories. Forty-eight taxa of other fungi were isolated, and fungal diversity was not exhausted by the sampling eVort. The same taxa of fungi were dominant in F. excelsior shoots of diVerent symptomatic categories, and moderate to high similarity of fungal communities was observed in shoots despite the symptoms. Forty-four isolates from 24 fungal taxa were used for artiWcial inoculations of 277 1-year-old F. excelsior seedlings in bare root nursery. After 2 years, only four fungi caused symptomatic necroses of bark and cambium: A. alternata, E. nigrum, Chalara fraxinea and Phomopsis sp. The most pathogenic was C. fraxinea, inducing symptoms on 50% of inoculated trees, while three other fungi caused necroses on 3-17% of inoculated trees. Infection biology of C. fraxinea and environmental factors determining susceptibility of F. excelsior to decline deserve future investigations.

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Ash dieback is a recent widespread disease on ash (Fraxinus sp.) that is causing important economic and ecological losses throughout Europe. The disease is initiated by the ascomycetous fungus Hymenoscyphus pseudoalbidus (anamorph Chalara fraxinea). The main aim of this study was to investigate seasonal pattern of lesion development associated with ash dieback. We present data on the spread of 324 natural lesions in ash shoots, branches and stems surveyed over a 32 month period. Most lesions were active and showed the greatest rate of growth during the summer; however, lesions were active throughout the year. Tree mortality was high, with more than a third of the surveyed trees dying during the study. Although many lesions permanently ceased to develop, the rate at which new lesions emerged was greater than the rate at which lesions entered a resting phase. The most common cause for a lesion going into a permanent state of rest was that it had encountered a branch-base. Genotype analysis showed that multiple infections can occur in a single tree given that different genotypes were identified in different lesions as well as in single lesions. A weak positive correlation was noted between tree health and tree size and a weak negative correlation was noted between tree overall health and lesion activity. The lower limit for H. pseudoalbidus growth in culture was between 4.0°C and 0.5°C.

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