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High genetic variation and extensive gene flow may help forest trees with adapting to ongoing climate change, yet the genetic bases underlying their adaptive potential remain largely unknown. We investigated range-wide patterns of potentially adaptive genetic variation in 64 populations of European beech (Fagus sylvatica L.) using 270 SNPs from 139 candidate genes involved either in phenology or in stress responses. We inferred neutral genetic structure and processes (drift and gene flow) and performed differentiation outlier analyses and gene-environment association (GEA) analyses to detect signatures of divergent selection.Beech range-wide genetic structure was consistent with the species’ previously identified postglacial expansion scenario and recolonization routes. Populations showed high diversity and low differentiation along the major expansion routes. A total of 52 loci were found to be putatively under selection and 15 of them turned up in multiple GEA analyses. Temperature and precipitation related variables were equally represented in significant genotype-climate associations. Signatures of divergent selection were detected in the same proportion for stress response and phenology-related genes. The range-wide adaptive genetic structure of beech appears highly integrated, suggesting a balanced contribution of phenology and stress-related genes to local adaptation, and of temperature and precipitation regimes to genetic clines. Our results imply a best-case scenario for the maintenance of high genetic diversity during range shifts in beech (and putatively other forest trees) with a combination of gene flow maintaining within-population neutral diversity and selection maintaining between-population adaptive differentiation.

The study aims to analyse the stability of the narrow crowned Norway spruce (pendula form) compared to the normal spruce form (pyramidalis form) and the hybrids of the two forms, in 5 field trials (Comandău, Lepşa 12, Ilva Mică and Voineasa) located in the Romanian Carpathians. Trees height (Th), breast height diameter (Dbh), height growth of the last year, crown diameter (Cd), number of branches per whorl (Nbw) and dominant branch diameter (Dbd) traits were measured and survival rate (Sr) was determined, at 20 years old. Also, branches finesse (Bf), trees volume (Tv) and trees slenderness (Ts) were calculated. In order to compare the wood density (Cwd) there were collected cores. In all trials ANOVA revealed significant (p 0.05) differences between the two forms of spruce and the hybrids (mainly between those that have a different crown form mother), especially for the stability and quality traits. Factorial ANOVA revealed a high influence (p 0.001) of the locality and also a significant influence (p 0.05) of the locality × spruce form interaction. The factor "form" was significant for some traits involved in Norway spruce stability (Ts, Cd, Nbw). The pendula trees present higher values for Sr, Dbh and Tv, and lower values for Ts, Cd, Nbw, Dbd and Bf, compared to pyramidalis spruce form, which showed a higher stability. Heritability was in generally low (h

In this study, different approaches were used to investigate the vulnerability of beech forests, located at the eastern limit of their natural range, to climate change. To accomplish this, six 2500 m2 plots were sampled in four European beech forest genetic resources, located in Romania at different altitudinal levels, varying from 230 to 580 m in the Bacău hills and between 650 and 1300 m in the Curvature Carpathian (Braşov region). The analysis of trees phenotypic traits, their radial growth, and the regeneration, did not indicate a vulnerability of the sampled stands to the fluctuations of the environmental factors from the 1950-2014 period. The growth indices of all three populations of Bacău hills are negatively correlated with both June air temperature of current year and September of the previous year. The precipitation amount of September previous year positively influenced the growth indices. The radial growth of plots in Braşov region is slightly linked to the climate. The temperature during the growing season represents a limiting factor for stands that are located outside of the optimal altitudinal species distribution (600-1200 m, in Romania), especially at low altitudes. Our results indicated that a rise of the temperature accompanied by a possible reduction of the precipitations (as is predicted for the coming years) could increase the sensibility of beech forests at lower altitude.

Forest genetic resources (FGR) of Romania harbour high genetic diversity although human impact, during the last two centuries, has strongly affected forest ecosystem composition and structure. Dynamic in situ conservation of FGR was the first choice approach to conserve the adaptation potential in natural populations of target tree species by selecting most representative and valuable populations. However, only 17 FGR growing in extreme environmental conditions have been selected and incorporated into the National Catalogue of Forest Genetic Resources. Establishment of new FGR in marginal tree populations that are close to the xeric limit of speciesʼ natural distribution should be extended. Almost half of tree populations selected for adaptation and included in the National Catalogue of Basic Materials for Production of Forest Reproductive Materials are represented by oak species, which is explained by lack of natural regeneration in the last decades. The shortage of native forest reproductive material of oak species (e.g. Quercus robur, Q. pedunculiflora, Q. pubescens) remains one of the biggest challenges in the conservation and management of rear edge oak populations in Romania. Molecular fingerprinting of FGR should constitute an important tool for genetic monitoring of evolutionary changes and to assist ecological reconstruction of fragmented tree populations. Collaboration with neighboring countries is an important aspect for dynamic conservation of FGR and their sustainable management.

In Romania, the forest genetic resources are included in two National Catalogues approved by ministerial orders. First is The National Catalogue of Forest Genetic Resources which includes 698 cores with main role of conservation, 577 of them being preserved in situ. Thus, the total area of in situ conservation is 10559.3 ha, being constituted especially for the main species from Romanian forests: European beech, Norway spruce, Silver fir and Oaks. At the same time, a large number of the conservation cores were established for some species with a low participation in the forest area (e.g. European larch, Scots pine) but located in natural populations where the environmental conditions are limited Most of the in situ conservation cores are also included in The National Catalogue of Forest Reproductive Materials. A total of 138 cores, accounting for 24% of the total number of the in situ conservation stands, with an area of 2140.7 ha, are the forest genetic resources located in different types of protected areas (national and natural parks, scientific and natural reservations, biosphere reservation, forest park or secular forests).

The Balkan Peninsula is one of the largest and most important European glacial refugia. However, the evolutionary history and phylogeographic pattern of temperate tree species that survived in the Balkans glacial refugia and their contribution to the genetic structure of the current population in the Carpathian Mountains remains poorly understood. Using polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP), and extensive population sampling, we explored the phylogeographic pattern of Carpinus betulus in both the Balkan Peninsula and the Carpathian region. We aimed to determine the locations of potential glacial refugia, in order to delineate post-glacial colonization routes, and also to test if northern cryptic refugia had persisted during the Last Glacial Maximum (LGM). Our results provide strong support for the existence of multiple refugia similar to the ‘refugia-within-refugia’ scenario, which would suggest that Carpinus betulus has experienced a complex evolutionary history. In agreement with pollen data, our findings corroborate with previous hypotheses suggesting that hornbeam has a distinct postglacial evolution in the southeast of Europe (Carpathians and Balkan Peninsula) compared with that in Western Europe. Three postglacial re-colonization routes, from three distinct effective glacial refugia: (1) the Dinaric Alps; (2) the Pirin and Rhodope Mountains and (3) the Strandzha Mountains, were detected within the Balkan Peninsula. The pattern of cpDNA haplotypes distribution across the Ukrainian Carpathians revealed a “suture zone”, which is a consequence of contact due to postglacial re-colonization between hornbeam populations originating from all three effective glacial refugia. The peculiar haplotype geographical structure found in C. betulus and the identified multiple glacial refugia in the Balkans may have direct consequences in the management and conservation of hornbeam forest genetic resources.

The monitoring studies carried out in the southern Romanian Carpathians (Retezat and Bucegi - Piatra Craiului Mts) provide a scientific support for long term ecosystem research (LTER). Their general objective is to characterize the air pollution and its potential effects upon forest ecosystems’ status and biodiversity in close connection with climatic changes. Two monitoring networks were placed in selected forest ecosystems for each LTER site (11 plots in Retezat and 22 plots in Bucegi - Piatra Craiului, respectively). Both in Retezat and Bucegi - Piatra Craiului Mts, the distribution of mean ozone (O3) concentrations are significantly correlated with altitude. The location (plot) has a significant influence on ozone concentration and in some cases the exposure period along the growing season has an influence. Ambient ozone (O3) and ammonia (NH3) concentrations were below toxicity levels for forest trees, their health status (crown condition) and volume growth being insignificantly affected. More than 45% of the rain events were acidic with pH≤5.5, thus contributing to an increased N deposition on forest stands. In Retezat Mts crown condition was relatively better than in Bucegi - Piatra Craiului Mts. As a consequence, in 2009 European beech (Fagus sylvatica) was the least affected species, with a percentage of damaged trees (crown defoliation higher than 25%) between 8.0% (Retezat) and 9.8% (Bucegi - Piatra Craiului), followed by Norway spruce (Picea abies) with 12.9% (Retezat) and 27.7% (Bucegi - Piatra Craiului). The most affected species was Silver fir (Abies alba) with 31.6% of damaged trees (Bucegi-Piatra Craiului). In the last period, both European beech and conifers (Norway spruce and Silver fir) with high defoliation (damaged trees) had lower annual relative increment than trees with low defoliation (healthy trees). A definite trend regarding rapid environmental modifications is not evident and no significant reduction of biodiversity occurred. In addition, all the vegetation biodiversity components of the forest ecosystems are specific to the stand type’s structures and local climate conditions, and no quantifiable consequences of specific air pollutants on ecosystems’ diversity occurred.

Background: Local adaptation is a key driver of phenotypic and genetic divergence at loci responsible for adaptive traits variations in forest tree populations. Its experimental assessment requires rigorous sampling strategies such as those involving population pairs replicated across broad spatial scales. Methods: A hierarchical Bayesian model of selection (HBM) that explicitly considers both the replication of the environmental contrast and the hierarchical genetic structure among replicated study sites is introduced. Its power was assessed through simulations and compared to classical 'within-site' approaches (FDIST, BAYESCAN) and a simplified, within-site, version of the model introduced here (SBM). Results: HBM demonstrates that hierarchical approaches are very powerful to detect replicated patterns of adaptive divergence with low false-discovery (FDR) and false-non-discovery (FNR) rates compared to the analysis of different sites separately through within-site approaches. The hypothesis of local adaptation to altitude was further addressed by analyzing replicated Abies alba population pairs (low and high elevations) across the species' southern distribution range, where the effects of climatic selection are expected to be the strongest. For comparison, a single population pair from the closely related species A. cephalonica was also analyzed. The hierarchical model did not detect any pattern of adaptive divergence to altitude replicated in the different study sites. Instead, idiosyncratic patterns of local adaptation among sites were detected by within-site approaches. Conclusion: Hierarchical approaches may miss idiosyncratic patterns of adaptation among sites, and we strongly recommend the use of both hierarchical (multi-site) and classical (within-site) approaches when addressing the question of adaptation across broad spatial scales.

Summary After the nursery testing, twelve Swiss stone pine (Pinus cembra L.) provenances from the Alps and Carpathian Mountains were planted out at two sites located at high elevation in the Southern and Northern Carpathians. Total height growth (H), annual height growth (h), root collar diameter (RCD), branches per whorl (BW) and survival (SV) were measured and analyzed. Analysis of variance showed highly significant (p < 0.01; p < 0.001) differences between provenances for all traits, except survival, suggesting that selection at the provenance level could be possible. Also, over locations analysis revealed significant genotype x environment interaction, demonstrating that some provenances react differently to environmental conditions and, selection should take this into account. The phenotypic coefficient of variation was moderate for growth and high for number of branches per whorl suggesting that selection within provenance can also be applied. Finding of significant and highly significant age-age and trait-trait phenotypic correlations indicated that early and indirect selection in Swiss stone pine species is possible. According to DUNCAN’s multiple range test the best performing provenances of the two mountain ranges were selected for operational planting and breeding programmes. The results of this study validate that a very slow growing species, such as Swiss stone pine may still possess very high genetic variation in growth rate; consequently, this trait can be improved. Finally, an attempt has been made to develop a seed transfer guidelines for the species by using the pattern of geographic variation as a basis.

The effects of air pollution on the genetic structure of Norway spruce, European silver fir and European beech were studied at four polluted sites in Slovakia, Romania and Czech Republic. In order to reduce potential effects of site heterogeneity on the health condition, pair-wise sampling of pollution-tolerant and sensitive trees was applied. Genotypes of sampled trees were determined at 21 isozyme gene loci of spruce, 18 loci of fir and 15 loci of beech. In comparison with Norway spruce, fewer genetic differences were revealed in beech and almost no differentiation between pollution-tolerant and sensitive trees was observed in fir. In adult stands of Norway spruce, sensitive trees exhibited higher genetic multiplicity and diversity. The decline of pollution-sensitive trees may result thus in a gradual genetic depletion of pollution-exposed populations of Norway spruce through the loss of less frequent alleles with potential adaptive significance to altered stressing regimes in the future. Comparison of the subsets of sensitive and tolerant Norway spruce individuals as determined by presence or absence of discolorations ("spruce yellowing") revealed different heterozygosity at 3 out of 11 polymorphic loci.

An international cooperative project on distribution of ozone in the Carpathian Mountains, Central Europe was conducted from 1997 to 1999. Results of that project indicated that in large parts of the Carpathian Mountains, concentrations of ozone were elevated and potentially phytotoxic to forest vegetation. That study led to the establishment of new long-term studies on ecological changes in forests and other ecosystems caused by air pollution in the Retezat Mountains, Southern Carpathians, Romania and in the Tatra Mountains, Western Carpathians on the Polish–Slovak border. Both of these important mountain ranges have the status of national parks and are Man & the Biosphere Reserves. In the Retezat Mountains, the primary research objective was to evaluate how air pollution may affect forest health and biodiversity. The main research objective in the Tatra Mountains was to evaluate responses of natural and managed Norway spruce forests to air pollution and other stresses. Ambient concentrations of ozone (O3), sulfur dioxide (SO2), nitrogen oxides (NOx) as well as forest health and biodiversity changes were monitored on densely distributed research sites. Initial monitoring of pollutants indicated low levels of O3, SO2, and NOx in the Retezat Mountains, while elevated levels of O3 and high deposition of atmospheric sulfur (S) and nitrogen (N) have characterized the Tatra Mountains. In the Retezat Mountains, air pollution seems to have little effect on forest health; however, there was concern that over a long time, even low levels of pollution may affect biodiversity of this important ecosystem. In contrast, severe decline of Norway spruce has been observed in the Tatra Mountains. Although bark beetle seems to be the immediate cause of that decline, long-term elevated levels of atmospheric N and S depositions and elevated O3 could predispose trees to insect attacks and other stresses. European and US scientists studied pollution deposition, soil and plant chemistry, O3-sensitive plant species, forest insects, and genetic changes in the Retezat and Tatra Mountains. Results of these investigations are presented in a GIS format to allow for a better understanding of the changes and the recommendations for effective management in these two areas. Keywords: Air quality, Ozone, Biodiversity, Ecology, Forests, Carpathian Mountains, Poland, Slovakia, Romania

The geographic distribution throughout Europe of each of 32 chloroplast DNA variants belonging to eight white oak species sampled from 2613 populations is presented. Clear-cut geographic patterns were revealed by the survey. These distributions, together with the available palynological information, were used to infer colonisation routes out of the glacial period refugia. In western Europe in particular, movements out of the Iberian and the Italian Peninsulas can be clearly identified. Separate refugia are also present in eastern Balkans, whereas further west in this peninsula similarities with Italy were evident. Movements resulting in the exchange of haplotypes between refugia both during the present interglacial and probably also during earlier glacial cycles were therefore inferred. The consequences of these past exchanges is that phylogenetically divergent haplotypes have sometimes followed very similar colonisation routes, limiting somewhat the phylogeographic structure. Cases of geographic disjunction in the present-day distribution of haplotypes are also apparent and could have been induced by the existence of rapid climatic changes at the end of the glacial period (specifically the Younger Dryas cold period), which resulted in range restriction following an early warm period during which oak first expanded from its primary refugia. This cold phase was followed by a new period of expansion at the outset of the Holocene, involving in some cases ‘secondary’ refugia. It is expected that these short climate oscillations would have led to a partial reshuffling of haplotype distribution. Early association between haplotypes and oak species are also suggested by the data, although extensive introgression among species has ultimately largely blurred the pattern. This implies that colonisation routes may have been initially constrained by the ecological characteristics of the species hosting each chloroplast variant. We suggest for instance that two oak species distributed in the north of the Iberian Peninsula ( Quercus petraea and Q. pubescens ) are recent post-glacial immigrants there. When considered together, conclusions on the location of glacial period refugia and the colonisation routes derived from molecular information and fossil pollen data appear to be both largely compatible and complementary.

A consortium of 16 laboratories have studied chloroplast DNA (cpDNA) variation in European white oaks. A common strategy for molecular screening, based on restriction analysis of four PCR-amplified cpDNA fragments, was used to allow comparison among the different laboratories. A total of 2613 oak populations (12,214 individual trees from eight species) were sampled from 37 countries, and analysed with the four fragments. They belong to eight related oak species: Quercus robur, Q. petraea, Q. pubescens, Q. frainetto, Q. faginea, Q. pyrenaica, Q. canariensis and Q. macranthera. During this survey, 45 chloroplast variants were detected and are described together with their phylogenetic relationships, but several of these haplotypes were pooled when there were some risks of confusion across laboratories during the survey, and finally 32 remained that were mapped and used in diversity analyses. A strong phylogeographic structure is apparent from the data, where related haplotypes have broadly similar geographic distributions. In total, six cpDNA lineages are identified, which have distinct geographic distributions, mainly along a longitudinal gradient. Most haplotypes found in northern Europe are also present in the south, whereas the converse is not true, suggesting that the majority of mutations observed were generated prior to postglacial recolonisation, corroborating the conclusions of earlier studies. The description of a new western European lineage constitutes a major finding, compared to earlier phylogenetic treatments. Although the eight oak species studied systematically share cpDNA variants when in sympatry, they partition cpDNA diversity differently, as a consequence of their different ecology and life history attributes. Regional differences in levels of differentiation also exist (either species-specific or general); these seem to be related to the intensity of past and present management of the forests across Europe but also to the level of fragmentation of the range within these regions.

A total of 1113 oak trees from 222 populations originating from eight countries (Austria, Bosnia-Herzegovina, Croatia, Hungary, Romania, Slovakia, Slovenia, Yugoslavia) were sampled in natural populations or in provenance tests. The sampled trees belong to four different species (Quercus robur, Quercus petraea, Quercus pubescens, Quercus frainetto) and to several putative subspecies. Variation at four chloroplast DNA (cpDNA) fragments was studied using restriction enzymes, resulting in the detection of 12 haplotypes. One haplotype was present in 36% of the trees, and six were found in 6–17% of the trees. The haplotypes are shared extensively between species and subspecies. They belong to three different lineages (A, C and E) and are phylogeographically structured in the region investigated. Haplotypes of lineage E dominate to the east of the Carpathian mountains in Romania, whereas the Carpathian Basin seems to have been colonised along several different colonisation routes, from the Balkan peninsula but also from Italy. The data support the possible role of climatic instability during the late glacial period in shaping this complex geographic structure. The presence of several secondary refugia could be inferred in the region, which have played a major role in the second step of recolonisation, at the onset of the Holocene period. # 2002 Elsevier Science B.V. All rights reserved.

In the Retezat Mountains concentrations of O3, NO2 and SO2 in summer season 2000-2002 were low and below toxicity levels for forest trees. While NH3 concentrations were low in 2000, the 2001 and 2002 concentrations were elevated indicating possibility for increased N deposition to forest stands. More than 90% of the rain events were acidic with pH values5.5, contributing to increased acidity of soils. Crown condition of Norway spruce (Picea abies) and European beech (Fagus sylvatica) was good, however, defoliation described as25% of foliage injured increased from 9.1% in 2000 to 16.1% in 2002. Drought that occurred in the southern Carpathians between fall 2000 and summer 2002 and frequent acidic rainfalls could cause the observed decline of forest condition. Both Norway spruce and European beech with higher defoliation had lower annual radial increments compared to the trees with low defoliation. Ambient O3 levels found in the Retezat did not affect crown condition of Norway spruce or European beech.