Needles of 20 years old spruces from Prednji vrh, in the area affected by the Sostanj power station, and from Pokljuka (Alps region, with a relatively unpolluted environment), were analysed for total sulphur content (St). Measurements were taken in a period between April and August twice monthly and in the period from August to April every four weeks (May 1993 - June 1994). Sulphur content in one and two years old spruce needles from Prednji vrh were much higher (one year old needles 2.06-2.50 mg/g needle dry weight, 1993) than in needle samples from Pokljuka (one year old needles 1.05-1.25 mg/g needle dry weight, 1993).

Endophytic fungi species composition in healthy needles of Austrian pine (Pinus nigra Arn.) was investigated at eight locations in Slovenia. Results from October 1994 and January 1995 isolations were compared with analyses of macronutrients, sulphur and lead content of the needles. About 80 species of microfungi were revealed. From the observations and measurements described it was not possible to discriminate between the effect of environmental factors and effect of air pollutants. The pollutants load of all measured elements is low and the environmental factors are very diverse. From the dendogram of isolation frequencies it seems that the age of the tree affects the species composition and frequencies of endophytes to the greatest extent.

Three principal approaches are used in our laboratory to analyze plant genomes: (i) The construction of high density molecular maps: We have established a refined microdissection procedure to construct chromosomal and subchromosomal libraries. Synchronized meristematic root tip tissue from which metaphase spreads can be prepared with a novel dropspread technique in combination with nl-scale PCR allowed the cloning of DNA of 10 or less chromosomes or chromosome segments. The construction of high-resolution maps from discrete genome regions can greatly facilitate genetic fingerprinting, gene isolation and QTL studies. (ii) Synteny analysis: The Aegilops-based deletion mapping system in wheat with an array of deleted chromosome parts and chromosome-specific RFLP markers has lead to the construction of a high density physical consensus map of wheat. The integration of wheat, barley and oat RFLP markers proves the colinearity between the wheat A-, B- and D-genomes, the H- genome of barley and the E-genome of Agropyron. (iii) Gene mapping in situ and chromosomes at high resolution: For the sensitivity enhancement of fluorescence in situ hybridization (FISH), the efficient preparation of plant chromosomes for high resolution scanning electron microscopy, mapping of low-copy sequences, and for comparative in situ hybridization a modified drop technique for plant protoplasts was developed. A tandemly amplified repetitive sequence element from microdissected barley chromosomes has enabled the karyotyping of single Gramineae genomes in a single step. These sequences are also useful for simultaneous double or triple hybridization experiments in an attempt to localize new sequence on specific chromosomes or chromosome segments. The physical mapping of the Sec-1 locus has been refined on the satellite of chromosome 1R of rye, and the syntenic locus on barley chromosome 1H identified. A method was developed for in situ hybrization and signal detection at high resolution using a field emission scanning electron microscope and a backscattered electron detector. Colloidal gold particles were localized on chromosome structures resembling the 30 mm fibre. Comparatively, an rDNA probe was located in the secondary constriction and highly compact adjacent regions of barley chromosomes.

Annual research has proven that the level of the so called natural cytogenetic damage to Norway spruce in Slovenia has been correctly estimated at 8. The results of cytogenetic bioindication calculated in accordance with this level show that average cytogenetic damage in the period from 1985 to 1992 enable us to determine environmental influences on the expression of cytogenetic damage.

The ectomycorrhizal biodiversity of selected Norway spruce stands on an Alpine limestone mountain was studied by inventories of occurring fruitbodies and descriptions of new ectomycorrhizal types. Based on fruitbody surveys a total of 137 ectomycorrhizal and 48 saprophytic species was recorded. Dependent on elevations as well as microclimatic, soil, and stand conditions there were significant differences in the fungal composition and number of species between the investigated forest plots.

Preliminary results and methodology related to the use of the fluorescence in situ hybridization technique for detection of plants hybrids are presented. Total genomic DNA probe preparation and the non-isotopic biotin probe labelling technique is presented. The use of an excess of unlabelled competitor DNA from a related plant species gives more constant results in determining whole plant genomes with in situ experiments on plant hybrids. Preliminary results of flourescence in situ hybridization for detecting alien rye chromatin and localisation on metaphase chromosomes and interphase nuclei in the bread wheat cultivar Yugoslavia are described. The methodology for detection of interspecies hybrids of common and sessile oak with flourescence in situ hybridization is given. The advantege of the use of total genomic biotin labelled probes for in situ hybridization is that it requires less effort than preparing species specific probes and the ease of interpretation of the results. The technique of fluorescence in situ hybridization is less species than other cytological techniques for higher plants and almost the same methodology can be used for detection of oak hybrids. The extraction of a high quantity of species specific DNA of high quality and probe preparation and labelling are the most important step.

The effect of atmospheric NH3 deposition on freezing tolerance of needles of Scots pine was investigated. Needles from 3 month old seedlings and from 20-30 year old field-grown Scots pine (Pinus sylvestris L.) trees were studied. Seedlings were exposed to NH3 during the first stage of hardening in the laboratory. Field-grown trees were growing in a "clean" or an NH3 polluted area. In the 3 month old seedlings, exposure to 1000 ml 1** (-1) NH3 for 4 weeks resulted in a 40 increase of total nitrogen content. Freezing tolerance of the needles was significantly enhanced in the NH3 exposed seedlings. In the field-grown trees, current and one year old needles were studied. Freezing tolerance of the current year needles did not differ significantly between the two forest stands. However, one year old needles from the NH3 polluted stand were significantly less frost hardened from October until February. Total nitrogen content was higher in both current and one year old needles from the NH3 polluted stand. At the same time, potassium content of these needles was lower compared to the needles from the clean stand. The content of calcium, magnesium and phosphorus did not differ significantly. It was concluded that a decrease of freezing tolerance upon atmospheric NH3 was not directly related to an increased nitrogen content, but might probably be due to the nutrient imbalance of the needles.

Biotrophic fungal interaction with host plants involve alterations in host metabolism in favour of the fungus, characteristic of perturbations in the levels of plant growth regulators. Pyrenopeziza brassicae mycelia, culture filtrate and spore extracts contained zeatin riboside and iso-pentenyladenisine-type cytokinins as determined using HPLC-ELISA. The significance of cytokinin synthesis by P. brassicae in relation to infection of host plant tissue is discussed.

Types of ectomycorrhizae on Norway spruce were determined in soil cores from two differently polluted forest research plots from the emission zone of Sostanj Thermal Power Plant (TPP). The two plots are comparable regarding site characteristics, but are differently polluted by the emmissions from the TPP. During the vegetations season 1993, 21 soil cores were taken from each plot. In these all fine roots and ectomycorrhizal roots tips were counted and the percentages of different types were calculated. Additionally, soil mycorrhizae from underneath fungal fruit bodies were taken in order to characterize new types of ectomycorrhizae. Classical anatomical methods and molecular tools were applied for identification. 17 types of ectomycorrhizae were determined on 28443 root tips in soil cores from the heavily polluted plot (Zavodnje). The predominant types were Paxillus involutus, Xerocomus badius, Piceirhiza parallela and Piceirhiza inflata. On the less polluted plot (Mislinja) 24 different types were determined on 38502 root tips in equal volume and number of soils cores. The most frequent types were Hydnum rufescens, Amphinema byssoides, Piceirhiza oleiferans, Cenococcum geophilum and Lactarius lignyotus. Mycobioindication of forest site pollution through selective sensitive (here Hydnum rufescens), in comparison to unsensitive (here Paxillus involutus), fungal species in ectomycorrhizae is discussed.