The evaluation of data on the forest tree condition is weak point in forest decline monitoring as it enables only a very rough estimation of the state of forest health. The paper analyses the interpretation of the forest tree defoliation data in order to estimate their relevance and applicability. Quality control results of defoliation assessment in Slovenia are presented and the state of defoliation with its changes in the year 1987, 1991, 1993, 1994 on the 16 km grid.

We have been isolating AC/GT and AG/CT SSRs from the Norway spruce (Picea abies K.) nuclear genome. We isolated several hundreds positive clones from a small-insert genomic library and following sequence analysis we designed primers for 36 of them, 24 containing AG and 12 AC SSRs. After testing them on a panel of spruce individuals 25 of the primer pairs producted a single-locus hypervariable pattern, with the remaining ones giving either a single monomorphic product (18) or very poor amplification (19) or amplification of multiple bands (38). Segregation in accordance with a simple Mendelian model of inheritance was demonstrated for all the loci amplified with the primer pairs giving a simple variable pattern. We screened a panel of 19 spruce trees at these loci. The average number of alleles per locus was 14 and expected heterozygosity 0.80, with up to 23 alleles per locus and heterozygosities exceeding 0.94. This shows that nuclear SSRs can be very useful markers in the population genetics of trees even though the overall efficiency of the marker identification process is quite low due to the high percentage of primer pairs producting complex or "dirty" patterns. We attribute this phenomenon to the high complexity of the spruce genome. Other methods, including the construction of libraries highly enriched for SSR sequences, that we developed in order to make SSR retrival and typing easier and faster will be discussed. We recently extended the use of PCR amplified SSR markers to the chloroplast genome. We demonstrated that mononucleotide poly(A/T) stretches are frequent in the chloroplast genomes of plants and show high levels of between and within population variation, making them ideal tools for cytoplasmic population genetic overcoming the difficulties in finding within species variation that are frequently encountered when analysing the cpDNA molecule by RFLPs or PCR-RFLPs. We will present results of the analysis of mediterranean pine species populations by using a set of cpSSRs that are distributed over the whole cpDNA molecule and discuss the possible applications of such markers for studying gene flow and for paternity analysis.

Giemsa C-banding method was applied on chromosomes in the female gametophyte tissue of Norway spruce (Picea abies (L.) Karesten). A karyogram of one single tree growing in the Botanical Garden of the University of Graz, Austria, was made by using an image analysis system. All chromosomes (n=12) showed pericentic C-bands. Chromosomes II, V and IX possed intercalary C-bands located at secondary constrictions. Telomeric C-bands appeared in chromosomes II and III on both arms.

Cytogenetical and biochemical investigations of common oak (Quercus robur L.) were applied in order to find suitable markers in bioindication of oak populations. In cytogenetical analyses the chromosome aberration and mitotic index were recorded. Chromosome banding, B-chromosomes, location and number of nuclear organizer regions were indicated using Giemsa C-banding method, fluorescence staining and silver impregnation. In biochemical analyses SDS-electrophoresis of soluble proteins and isoelectric focusing of isoperoxidases were used.

It was our aim to investigate whether the cytokinin status of Norway spruce seedlings (Picea abies L. Karst.) was changed when the plants were grown under nutritive stress. Cytokinins in shoots and roots from spruce seedlings, grown in various hydrocultural media with complete (control) or poor nutrient supply (stress), were determined by an indirect competitive enzyme-liked immunosorbent assay. The shoots of spruce seedlings grown in a poor acidic culture medium, to which aluminium ions (AlCl3, 0.8 mM) were added, showed up to 5-fold higher concentrations of zeatin riboside and isopentenyladenosine (iPa). When adding naphtylacetic acid to the nutrient medium also higher levels of cytokinin ribosides were measured in the shoots. In the roots however cytokinin riboside levels did not increase significantly under Al- or auxin treatment. Feeding experiments using tritiated iPA have shown that the metabolism of this cytokinin is strongly retarded in the roots of stressed seedlings. The experiments show that acid conditions and nutrient shortage together with aluminium ions in the rhizosphere increase the number of lateral roots in Norway spruce seedlings. It is assumed that the increased number of sites of cytokinin biosynthesis and the reduced metabolism of iPA result in the higher content of cytokinin ribosides in the shoots of seedlings.

In 1992, a monitoring survey on a national scale was carried out using Hypogymnia physodes (L.) Nyl. as a biomonitor for trace element air pollution. The primary aim was to analyse epiphytic lichens collected at 86 sampling locations of the 16 x 16 km bioindication grid using k(0)-based instrumental neutron activation analysis (k(0)-INAA), and X-ray fluorescence spectrometry (XRF) for sulphur and lead, to obtain information about the levels of elements in the atmosphere and to identify significant pollution sources. The geographical concentration patterns of the trace elements obtained from the lichen data were mapped and compared with the index of atmospheric purity (IAP) calculated on the basis of data from lichen thalli type mapping, obtained on a more dense bioindication grid in 1991. The results obtained showed good agreement between the mapping of sulphur and trace elements with the status of lichen vegetation. The most exposed regions with elevated trace element levels and lower values of IAP were in the north-western Alpine part of Slovenia which coincides with high precipitation, and in the east of Slovenia, where many local pollution sources are situated.

With the development of molecular marker technologies to construct detailed molecular maps of crop species it has become possible to study much more precisely than before the possible roles of ABA by analysing the distribution of quantitative trait lici (QTL). This has been done with mapping populations of maize and wheat to study the importance of endogenous ABA in determining stomatal function, water-use efficiency and root growth in plants growing under water-stressed conditions. An F2 population of maize was mapped with 100 RFLP markers and F2 plants and their F3 progenies sampled for leaf and xylem ABA contents and stomatal conductance. Correlations using genotype mean data at QTL significant for ABA content showed that stomatal conductance was more likely to be regulated by xylem ABA content than leaf ABA content. The same maize F3 population was used to identify QTL for root growth was positively associated with root ABA content. Water-use efficiency, measured as 13C discrimination, was measured in a mapping population of doubled haploid lines of wheat. Leaf ABA concentrations were also measured. Comparison of QTL for the two traits showed that leaf ABA concentrations were unlikely to have had an effect on plant water-use efficiency in those field conditions. QTL analysis is a very powerful technique for testing the likely roles of endogenous ABA.

This article deals with use of several bioindication methods in forest decline studies within an international TEMPUS project carried out in Slovenia. The aim of the project was to introduce and connect different bioindication methods in order to improve methods and qualify the research staff working in this research field in Slovenia. The following bioindication methods were introduced or adapted for forest decline research in Slovenia: biochemical and physiological markers of air pollution / stress impact on forest trees (Graz Univ.: photosynthetic pigments, thiols, ascorbic acid, protective enzyme systems; Cambridge Univ.: plant hormones; Munich Univ.: mineral nutrition of forest trees); epiphytic lichens as indicators of air quality on forest sites (Graz Univ., Ljubljana Univ); determination of types of ectomycorrhizae as indicators of forest soil condition (Munich Univ., Ljubljana Univ.). The significance and practical use of bioindication methods in environmental studies is also discussed.

Tissue specific localization of soluble secondary metabolites was demonstrated in ectomycorrhizae and uninfected roots of Picea abies, Larix decidua, Pinus sylvestris and Abies alba. Compounds were analysed by HPLC and by histochemical methods. Specific distribution and age-dependent changes were observed with several metabolites. For catechin corresponding patterns were found within the cortex of all investigated conifers. Levels of this metabolite were highest within the apical cortex cells and decreased on aging of the tissue. In spruce contents of catechin showed evident seasonal changes. Stilbene glucosides were detected within the pericycle of spruce roots and as well the phloem tissue of larch with levels increasing during root development. In larch two anthocyanins accumulated in the calyptra. In older tissues of larch roots larixin-glucosid and quercetin-rhamnosid were detected, the latter with highest levels in the phloem tissue. For cell wall bound ferulic acid increasing levels could be found concomitant with the development of the root.

Work was carried out to investigate the effect of pollution on the needle cytokinin content of mature Picea abies (L.) Karst individuals. Needles were collected from trees growing at two sites, one polluted and relatively non-polluted, in central Slovenia. After ethanolic extraction and purification, the needle cytokinin extracts were separed by reverse-phase HPLC. Cytokinin content was analysed using four direct ELISA techniques specific for Z-, DHZ- and iP-type cytokinins and the O-glucoside conjugate of Z. Results showed a trend towards the accumulation of cytokinins in needles from trees subject to high levels of atmospheric pollution, particulary with respect to ribotide derivatives. No difference was found in the (OG)Z content between polluted and non-polluted samples. The results are discussed with respect to the origins of the observed accumulation.