Detection of haplotypic variation and natural hybridization in halepensis-complex pine species using chloroplast simple sequence repeat (SSR) markers

In this investigation, nine chloroplast, paternally inherited simple-sequence repeat (cpSSR) markers were used to describe genetic variation of three closely related species belonging to the halepensis complex ( Pinus halepensis Ait., P. brutia Mill. and P. eldarica Medwed.). Both the infinite allele model (IAM) and stepwise-mutation model (SMM) have been applied to the analysis of the genetic structure of natural populations and the geographical distribution of haplotypic variation. SMM-based estimators performed better than IAM-based estimators for large values of within-population diversity and divergence between population pairs. Overall, large haplotypic variation and high genetic divergence were detected for both P. halepensis and P. brutia . The genetic structures of the three species are discussed with consideration to the evolutionary and ecological characteristics of these species. Three highly informative markers showing size variants distinguishing P. halepensis from the other two species were used to provide more information on the occurrence of natural hybridization in a Turkish sympatric population of P. halepensis and P. brutia . Strong evidence of introgression of ' halepensis ' chloroplast haplotypes into P. brutia seeds (but not vice versa) was detected. According to previous evidence from controlled crossings, matings between the above species seem to be successful only when P. halepensis is the pollen donor and P. brutia is the female parent (but not reciprocally). The existence of unidirectional gene flow in sympatric populations confirms previous evidence about partial reproductive barriers between P. halepensis and P. brutia . Implications of the above evidence for the evolutionary history of these species are discussed.     

Morphological and Chemical Differentiation between Tunisian Populations of Pinus halepensis,Pinus brutia,and Pinus pinaster

The lipid fraction of seeds from different pine species and populations was studied regarding total lipid content, fatty acid profile and vitamin E composition. The investigated seeds contained a high percentage of lipid (13.6 to 31.5 %). Lipid fractions were found to be rich in vitamin E, which varied significantly among species and populations. P. halepensis (Ph−Hn) showed the highest content of vitamin E (256.3 mg/kg of seeds) and the uppermost content of α-tocopherol (44 mg/kg). However, P. halepensis (Ph−Kas) was the richest in γ-tocopherol (204.9 mg/kg). Lipid fractions had a low content of δ-tocopherol (1.2 to 3.6 mg/kg. The highest content of γ-tocotrienol (∼18 %) was determined for P. halepensis (Ph−Dc and Ph−Hn). Thirteen fatty acids were identified by GC-FID with significant variation between the investigated species. The linoleic acid was the major fatty acid followed by oleic acid and palmitic acid. The chemical differentiation among species for the composition of fatty acids and vitamin E was confirmed by PCA. Significant correlations were observed between the content of vitamin E and fatty acids and ecological parameters of P. halepensis populations.     

Relations between apical structure and growth patterns in Pinus halepensis Mill,seedlings

Abstract

The process of primary growth in 2-year-old seedlings of 11 populations of Pinus halepensis Mill. is described. At the end of the first growing season one type of apical structure was observed: type-1, a tuft of primary needles placed close together, surrounding and protecting a meristematic apex.

At the end of the 2nd growing season, three types of apical structure were observed: type-1; type-2, a terminal winter bud; and type-3; a «bud» with characteristics of both type-1 and type-2.

Morphological observation along with an anatomical examination of the winter bud led to the conclusion that the definitive growth pattern in juvenile P. halepensis is monocyclic with a variable number of summer shoots. This growth pattern is reached by some P. halepensis populations in 3–4 years, by contrast, in other pine species two years are usually needed.

The populations studied differed both in growth potential (differences in number of cycles, ratio of first cycle to total growth, growth rates) and in the developmental stages of the apical meristem.

Four groups could be identified: (i) Morocco and Spain, (a limited growth, few cycles, a high ratio of 1st cycle to total growth, and growth in 2nd season almost entirely due to free growth); (ii) Algeria and Greece, (moderate to low growth, a large number of cycles, a low ratio of 1st cycle to total growth, and very early formation of apical structure with preformed primordia); (iii) Israel and Central Italy (a high growth, a large number of cycles, a medium ratio of 1st cycle to total growth, and early formation of apical structure with preformed primordia); (iiii) Greece, France and Italy, which was intermediate between group (i) and the other 2 groups.     

Dr. Alfred Jeske 75 jahre

The chemical composition, phytotoxic and antifungal activities of the essential oils isolated by hydrodistillation from the needles of Tunisian Aleppo pine harvested from different provenances were evaluated. The chemical composition analysed by gas chromatography/mass spectrometry (GC/MS) revealed variability among provenances displaying interesting chemotypes, (Z)-caryophyllene (16.16–28.9%), β-myrcene (8.5–22.9%), α-pinene (11.7–13.14%), β-pinene (3.13–11.8%), bicyclogermacrene (5.2–12.37%), α-terpinolene (8.11–11.01%) and α-humulene (2.85–5.2%), which were the main components in the oil. Antifungal ability of Aleppo pine oils was tested by disc agar diffusion against 10 phytopathogenic fungi. Weak antifungal activity was observed for the essential oils isolated. Furthermore, in contrast, the herbicidal activity investigated for three common weeds in Tunisian cereal crops was very strong and seed germination was inhibited at a low concentration and their herbicidal effects were higher than those of a commercial herbicide.     

Can Cone Scales and Bark of Aleppo Pine Be Used in the Biomonitoring of Some Heavy Metals?

The highest levels of Pb in soils and needles were detected in the Lisbon Botanical Garden, reflecting the intense car circulation since the end of the late nineteenth century, and the presence of tetraethyl lead in petrol. Levels of Pb in Aleppo bark were at or above the threshold values for plant tissues as defined by the UN-ECE/EC (>4–30 μg/g). Nickel was easily washed by distilled water from the bark, indicating a predominant adsorption due to atmospheric deposition, while only Pb was water-extracted from cone scales, but no more than 0.88 μg/g (4.3% of the total). If we take into account the dry weight of each cone, we can assess the amount of extractable lead present in a given time (ranges between 3.0 and 15.5 μg), although its eventual transfer to the ground through rainfall will be hampered by the cone architecture. The average levels of total Pb in the cones (12.7–35.4 μg/g) fall within the range defined by the UN-ECE/EC, generally; only the reference value is below 4 μg/g. Due to the high capabilities of allocating Pb inside the structure and through adsorption, Aleppo bark and cones can be very interesting tools in the monitoring of Pb.     

The value of leaf cuticle characteristics in the identification and classification of Iberian Mediterranean members of the genus Pinus

This study reports the value of leaf cuticle characteristics in the identification and classification of Iberian Mediterranean species of the genus Pinus (P. nigra subsp. salzmannii, P. pinaster, P. pinea and P. halepensis), with the aim of using these characters to identify isolated cuticles and stomata in palynology slides. Preparations were made of the cuticles of pine needles belonging to one natural Iberian population of each of the above species. A number of epidermal morphological characteristics were then recorded with the aim of distinguishing these species from one another. The structure of the stomatal complex (the shape and arrangement of the subsidiary cells) was different in each species. The aperture of the epistomatal chamber was significantly smaller in P. pinea than in the other species examined, and the variables recorded for the thickening of the guard cells provided relationships that clearly distinguished all four taxa. The width and length of the stomata and the upper woody lamellae, the central distance between the external limits of the medial lamellae borders and the length of the stem were the most useful variables in this respect. The present results contribute to the ongoing discussion regarding the taxonomic classification of the members of Pinus, and provide valuable clues for the identification of Iberian Mediterranean pine species from small pine needle fragments or isolated stomata. After validation of the present results for multiple populations, these results could also be used to help identify fossil leaf macroremains and the scattered/isolated stomata commonly observed in palaeopalynological samples. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 161 , 436–448.     

Small‐scale spatial variability in the distribution of ectomycorrhizal fungi affects plant performance and fungal diversity

The effects of spatial heterogeneity in negative biological interactions on individual performance and species diversity have been studied extensively. However, little is known about the respective effects involving positive biological interactions, including the symbiosis between plants and ectomycorrhizal (EM) fungi. Using a greenhouse bioassay, we explored how spatial heterogeneity of natural soil inoculum influences the performance of pine seedlings and composition of their root‐associated EM fungi. When the inoculum was homogenously distributed, a single EM fungal taxon dominated the roots of most pine seedlings, reducing the diversity of EM fungi at the treatment level, while substantially improving pine seedling performance. In contrast, clumped inoculum allowed the proliferation of several different EM fungi, increasing the overall EM fungal diversity. The most dominant EM fungal taxon detected in the homogeneous treatment was also a highly beneficial mutualist, implying that the trade‐off between competitive ability and mutualistic capacity does not always exist.     

Analysis of spatial genetic structure in an expanding Pinus halepensis population reveals development of fine-scale genetic clustering over time

We analysed the change of spatial genetic structure (SGS) of reproductive individuals over time in an expanding Pinus halepensis population. To our knowledge, this is the first empirical study to analyse the temporal component of SGS by following the dynamics of successive cohorts of the same population over time, rather than analysing different age cohorts at a single time. SGS is influenced by various factors including restricted gene dispersal, microenvironmental selection, mating patterns and the spatial pattern of reproductive individuals. Several factors that affect SGS are expected to vary over time and as adult density increases. Using air photo analysis, tree-ring dating and molecular marker analysis we reconstructed the spread of reproductive individuals over 30 years beginning from five initial individuals. In the early stages, genotypes were distributed randomly in space. Over time and with increasing density, fine-scale (< 20 m) SGS developed and the magnitude of genetic clustering increased. The SGS was strongly affected by the initial spatial distribution and genetic variation of the founding individuals. The development of SGS may be explained by fine-scale environmental heterogeneity and possibly microenvironmental selection. Inbreeding and variation in reproductive success may have enhanced SGS magnitude over time.     

Effects of repeated burning on Mediterranean communities of the northeastern Iberian Peninsula

Question: In relation to a single fire, do repeated wildfires in Mediterranean type ecosystems (1) reduce plant species richness or diversity; (2) modify patterns of abundance or dominance of plant species or (3) alter plant composition? Location: Pinus halepensis dominated communities of Catalonia, northeastern Iberian Peninsula, western Mediterranean Basin. Methods: Regional, paired design with 14 study sites, each consisting of a once burnt area (1994) and a twice burnt area (1975–1993 and 1994). Ten years after the last fire, we recorded all vascular plant species present in nested plots and quantified their relative abundances on transects. We compared species richness, diversity, dominance and relative abundance and species‐area correlations between paired once and twice burnt areas and assessed their floristic composition similarity. Results: No statistically significant differences were found in species richness or diversity. Slopes of species‐area correlations were higher in once burnt areas. In twice burnt areas, dominance by one or two species was higher. P. halepensis showed lower relative abundance and nanophanerophytes showed higher relative abundance. No differences were found for resprouter, seeder or resprouter‐seeder species. Floristic composition similarity between paired areas tended to be higher in less productive sites. Conclusions: Fire recurrence had contrasting effects on species richness at different spatial scales. Repeated burning reduced the relative abundance of the dominant tree species, which resulted in a higher relative abundance of shrubs. It also promoted the dominance of herbs, particularly Brachypodium retusum. However, it did not change the relative abundance of regenerative groups. Paired areas were more similar as they were more Mediterranean in terms of climatic conditions.     

Implications of foliar terpene content and hydration on leaf flammability of Quercus ilex and Pinus halepensis

We investigated the implications of foliar hydration and terpene content on leaf flammability in two widely distributed forest species of the Mediterranean basin, Quercus ilex, which does not store terpenes, and Pinus halepensis, a terpene-storing species. The experiments were carried out in plants grown under different water regimes that generated a wide range of foliar hydration and terpene contents. We monitored the temperatures and time elapsed to reach the smoke, pyrolysis and flame phases. Smoke appeared much earlier (37 versus 101 s) and at lower temperatures (96 versus 139 degrees C) in Quercus ilex than in Pinus halepensis. Quercus ilex reached pyrolysis earlier than Pinus halepensis (278 versus 338 s) but at the same temperature (365-371 degrees C). There were no significant differences in time elapsed nor in temperature for flammability (386-422 s; 505-487 degrees C in both species). Quercus ilex had lower water hydration than Pinus halepensis (41 versus 100%) and the leaf content of terpenes in Quercus was three orders of magnitude lower. The results of this study show no differences in the flame phase between the two species and the absence of a significant relationship between temperature and elapsed time of the different flammability phases in relation to monoterpene content; thus indicating that the role of monoterpenes in flammability phases is smaller than that of the water content. This, however, does not exclude the effects of terpene content on plant combustibility and fire propagation once fires start.