Aleksandra Wojnicka-Półtorak

Effects of heavy metal pollution on genetic variation and cytological disturbances in thePinus sylvestris L. population

This isoenzymatic and cytogenetic study has shown significant differences in genetic composition between two groups ofPinus sylvestris trees: tolerant and sensitive to heavy metal pollution. Total and mean numbers of alleles and genotypes per locus were higher in the pollution-sensitive group of trees, but heterozygosity (Ho) was lower in this group. Fixation index (F) indicates that trees tolerant for pollution were in the Hardy-Weinberg equilibrium, while the sensitive group had a significant excess of homozygosity. Cytological analyses demonstrated numerous aberrations of chromosomes in meristematic root tissue of seedlings developed from seeds collected from trees in the polluted area. The aberrations included chromosome bridges and stickiness, laggards, retarded and forward chromosomes, and their fragments. The mitotic index was markedly lower in this group of seedlings, as compared to the control. Both isoenzymatic and cytological analyses showed a significant influence of heavy metal ions on the genetic structure of thePinus sylvestris population.

Adaptation strategies and referencing trial of Scots and black pine populations subjected to heavy metal pollution

The impact of industrial heavy metal pollution on Scots pine (Pinus sylvestris L.) and black pine (Pinus nigra Arn.) populations was investigated. Sampled pine stands, which were located in Upper Silesia (southern Poland) in an area strongly polluted by heavy metals, consisted of resistant and sensitive trees. To evaluate the adaptation process, genetic structure and diversity was tested using isozyme analysis. Higher levels of Zn, Pb, Cd and Cu were detected in needles of sensitive trees compared with resistant ones. With respect to morphology, Scots pines were more distinctly impaired than black pines. Although black pines had lower heavy metal concentrations, levels in 1-year-old needles, other than Cu, significantly exceeded “reference plant” values (Markert 1994). In both species, resistant trees demonstrated a lower degree of genetic variation than metal-sensitive trees with respect to some enzyme loci (SHDH A, PGI, PGM, MDH C and DIA). This observation was corroborated in sensitive trees by the smaller number of identified alleles and alleles per locus, absence of private alleles and significant excess of homozygotes in relation to expected Hardy–Weinberg equilibrium values. Assuming that only resistant trees of both species survive under conditions of prolonged soil contamination, the observed genetic structure implies that remaining populations will be depleted of some alleles of unknown adaptive value to future selection pressures. Genetic changes induced by heavy metals suggest an important role for specific enzymes—FEST, SHDH A and B, GOT B and PGI—in the adaptation process. Our results may serve as a basis for selection and propagation of individuals appropriate for re-cultivation of areas chemically degraded by industrial activity.

Cross-species amplification and characterization of microsatellite loci in Pinus mugo Turra

Pinus mugo (dwarf mountain pine) is an important component of European mountain ecosystems. However, little is known about the present genetic structure and population differentiation of this species at the DNA level, possibly due to a lack of nuclear microsatellite markers (SSR) developed for Pinus mugo. Therefore in this study we transferred microsatellite markers originally developed for Pinus sylvestris and Pinus taeda to Pinus mugo. This cross-species amplification approach is much faster and less expensive than isolation and characterization of new microsatellite markers. The transfer rates from the source species to Pinus mugo were moderately low (26%). There were no differences in microsatellite repeat motifs between the source species and Pinus mugo. Nuclear microsatellite markers successfully transferred to Pinus mugo can be applied to various genetic studies on this species, due to the high level of their polymorphism and high value of polymorphic information content.

Volatiles as Chemosystematic Markers for Distinguishing Closely Related Species within the Pinus mugo Complex

Headspace solid‐phase microextraction (HS‐SPME) coupled to GC/MS analysis was used to identify the constituents of pine‐needle volatiles differentiating three closely‐related pine species within the Pinus mugo complex, i.e., P. uncinata Ramond ex DC., P. uliginosa G.E.Neumann ex Wimm., and P. mugo Turra. Moreover, chemosystematic markers were proposed for the three analyzed pine species. The major constituents of the pine‐needle volatiles were α‐pinene (28.4%) and bornyl acetate (10.8%) for P. uncinata, δ‐car‐3‐ene (21.5%) and α‐pinene (16.1%) for P. uliginosa, and α‐pinene (20%) and δ‐car‐3‐ene (18.1%) for P. mugo. This study is the first report on the application of the composition of pine‐needle volatiles for the reliable identification of closely‐related pine species within the Pinus mugo complex.

Biogeography and evolutionary factors determine genetic differentiation of Pinus mugo (Turra) in the Tatra Mountains (Central Europe)

Inter Simple Sequence Repeats (ISSR) markers were used to assess genetic diversity within and among populations of dwarf mountain pine (Pinus mugo Turra) growing in the Tatra National Park (UNESCO Biosphere Reserve) in Southern Poland (Central Europe). The analyzed population belongs to two different geobotanical sub-districts: the Western and High Tatras. The level of genetic diversity assessed in this study for P. mugo is generally comparable to that reported for the other pine species in the Pinaceae family assessed by ISSR markers, especially with respect to Nei’s genetic diversity and the percentage of polymorphic bands. Bayesian analysis clustered the analyzed populations into two groups, corresponding to their geobotanical locations in the Tatras. Significant divergence between the two genetical clusters was supported by the results of Analysis of Molecular Variance (AMOVA). According to the Mantel test, there was no correlation between the genetic distance and the geographical distance. The present study confirms the existence of two genetically distinct clusters of P. mugo populations in the Tatra Mountains. The observed high population-genetic differentiation of P. mugo in the Tatras could be attributed to several genetic, environmental and historical factors occurring in this mountain area.

Trace element contamination differentiates the natural population of Scots pine: evidence from DNA microsatellites and needle morphology.

The Scots pine is often used in the biomonitoring of forests. Studies on the chemical composition plus variability of its needles morphological structure allow for an assessment of the state of environmental pollution. However, in their natural populations, the response of individual trees to stress differs. This study reports on the influence of long-term soil contamination with trace elements on the morphology of the needles, its possible relation to the differentiation of the genetic pool, and their implications for biomonitoring. In the natural and self-renewable pine stand growing near the point polluter (zinc smelter, Upper Silesia, Poland), two categories of trees are observed with respect to their health status: pollution-tolerant (T) and pollution-sensitive (S). A detailed analysis of the trace element content of the needles reveals that the concentration of Cd, Zn, Pb, and Cu in the needles is significantly higher in S as compared to T individuals. The metal accumulation pattern decidedly follows the sequence Pb > Cd > Cu > Zn. An analysis of the fluctuating asymmetry (FA) of the needles reveals that sensitive trees showed an FA index ten times higher in comparison to tolerant ones. Moreover, the high differences between these S and T tree groups are also observed in the basic genetic diversity parameters investigated by an analysis of DNA simple sequence repeats (SSR). The concentration of trace elements in pine needles, distinct in sensitive and tolerant trees and in connection with their morphological and genetic characteristics, may reflect an adaptation process. The level of Mg and Fe content in the needles could be a physiological-toxicological index for evaluating trace element “lethality” expressed as Mg and Fe mineral-survival strategies. The example of differences described in this Scots pine population should be taken into consideration in ecotoxicological research to better interpret the obtained results.

Temporal dynamics in the genetic structure of a natural population of Picea abies

Abstract The temporal dynamics of the genetic diversity of the Norway spruce population provide valuable information on the conservation and management of its genetic resources. The relationships between genetic and demographic parameters are of fundamental importance for understanding the adaptability of forest tree populations. The study was aimed at determining the genetic differentiation of five age classes of a naturally regenerating Picea abies population from the Białowieża Primeval Forest (BPF) in Poland. Using mitochondrial DNA markers (nad1 intron b/c; mt15-D02) and nuclear DNA microsatellites (EAC2C08; EATC2B02; EATC2G05; SpAGD1) we determined the genetic structure between and within the age classes of the P. abies population. The significant subdivision of genetic variation (Fst) detected across the age classes is comparable to those found between different populations of this species. Two microsatellite loci behaved as “outlier loci,” exhibiting directional selection as revealed in the LOSITAN analysis. The significant deficit of heterozygosity may be a consequence of a temporal Wahlund effect and selective processes favoring homozygotes in the specific environment of the BPF. Population genetic structure can vary among life stages as a result of multiple factors, such as pollen and seed dispersal patterns, density of trees, past reproductive episodes, site conditions, and selective processes.

Adaptation Mechanisms of Pinus sylvestris L. in Industrial Areas

In spite of a positive direction of changes occurring in the contaminated environment, the local industry still appears to exert a negative influence on plant vegetation. Forests which grow in many highly industrialized zones enable research on the influence of anthropopression on the natural population and are one of the best models for the study of plant adaptation to heavy metals in soil. In some cases, it is possible to follow processes of re-naturalization occurring on post-industrial areas in situ. Research undertaken in heavily polluted regions pointed to an interesting phenomenon of differentiation among the Scots pine populations with respect to the health status. Adaptive genetic diversity reflects differences in the survival capabilities of individuals exposed to stress and shows the selective pressure against trees with specific genotypes. This chapter emphasizes on the Scots pine (Pinus sylvestris L.) as one of the most frequently used bioindicators in the European forests and their application in the study of microevolutionary processes in tree populations. It may enhance a better understanding of how the soil pollution can change the genetic structure of important forest species.

Genetic heterogeneity in age classes of naturally regenerated old growth forest of Picea abies (L.) Karst

Abstract The Białowiez·a Primeval Forest is located northeastern Poland. It is one of Europe’s most precious old growth forests in terms of abundance and richness of vegetation, retaining features of a primeval lowland forest which cannot be found anywhere else on the European continent. The aim of the study was to assess the genetic heterogeneity of the naturally regenerated Picea abies population using five chloroplast microsatellite markers. In total, 290 trees representing five age classes were studied. Clear patterns of genetic differentiation in relation to demographic substructuring were found within the population. The class of embryos exhibited the greatest genetic richness as evident from the highest number of alleles and haplotypes, the highest mean number of private alleles and haplotypes and the highest haplotype diversity. In the subsequent age classes, a significant decrease in the level of genetic variation was observed. Our data demonstrate that long-lived, highly outcrossing tree species growing in continuous stands can be genetically heterogeneous on a small geographic scale. The heterogeneity is related to age structure and it is likely due to the underlying mating system and selection processes.