Spiral grain in bristlecone pines (Pinus longaeva) exhibits no correlation with environmental factors

Six hundred and eleven Great Basin bristlecone pines (Pinus longaeva) were surveyed in two separate groves in California’s White Mountains. The presence and direction of spiral grain were recorded for each tree as well as elevation above sea level, horizon angles, latitude and longitude, trunk diameter, whether the tree was dead, and whether the trunk was broken. The proportions of left-handed, right-handed and straight trees were similar in every part of both groves, although the groves lie at different elevations. No significant correlation was found between the direction of spiral grain and any environmental factor. The hypothesis that spiral grain is an adaptation to distribute sap evenly between the roots and the crown in Pinus longaeva is not strongly supported, since spiral grain is not correlated with asymmetric environments and most trees exhibit <90° rotation through the main stem. The data also do not support the idea that spiral grain makes the tree more resistant to breaking in strong winds. Right-handed spiral grain is predicted by this hypothesis, but most bristlecone pines are either left-handed or exhibit no spiral grain. Bristlecone pines are often uprooted from thin soils by strong winds, but rarely are the main stems broken by this mechanism. Spiral grain in Pinus longaeva growing in California’s White Mountains does not appear to be under environmental control.     

Recognizing site adversity and drought-sensitive trees in stands of bristlecone pine (Pinus longaeva)

Bristlecone pine has been called the world’s oldest known living thing. The tree grows in mountainous terrain near timberline in three southwestern states. Because of the species’ longevity, sensitivity to climatic fluctuations, and unique growth form, it has attracted considerable attention from both scientists and recreationists. This paper discusses bristlecone pine in relation to its environment in east-central Nevada and sets forth criteria for recognizing trees that may be old or sensitive to climatic changes.     

Suitability of pines and other conifers as hosts for the invasive Mediterranean pine engraver (Coleoptera: Scolytidae) in North America

The invasive Mediterranean pine engraver, Orthotomicus erosus (Wollaston) (Coleoptera: Scolytidae), was detected in North America in 2004, and it is currently distributed in the southern Central Valley of California. It originates from the Mediterranean region, the Middle East, and Asia, and it reproduces on pines (Pinus spp.). To identify potentially vulnerable native and adventive hosts in North America, no-choice host range tests were conducted in the laboratory on 22 conifer species. The beetle reproduced on four pines from its native Eurasian range--Aleppo, Canary Island, Italian stone, and Scots pines; 11 native North American pines--eastern white, grey, jack, Jeffrey, loblolly, Monterey, ponderosa, red, Sierra lodgepole, singleleaf pinyon, and sugar pines; and four native nonpines--Douglas-fir, black and white spruce, and tamarack. Among nonpines, fewer progeny developed and they were of smaller size on Douglas-fir and tamarack, but sex ratios of progeny were nearly 1:1 on all hosts. Last, beetles did not develop on white fir, incense cedar, and coast redwood. With loblolly pine, the first new adults emerged 42 d after parental females were introduced into host logs at temperatures of 20-33 degrees C and 523.5 or 334.7 accumulated degree-days based on lower development thresholds of 13.6 or 18 degrees C, respectively.     

Family 34 glycosyltransferase (GT34) genes and proteins in Pinus radiata (radiata pine) and Pinus taeda (loblolly pine)

Using a functional genomics approach, four candidate genes (PtGT34A, PtGT34B, PtGT34C and PtGT34D) were identified in Pinus taeda. These genes encode CAZy family GT34 glycosyltransferases that are involved in the synthesis of cell‐wall xyloglucans and heteromannans. The full‐length coding sequences of three orthologs (PrGT34A, B and C) were isolated from a xylem‐specific cDNA library from the closely related Pinus radiata. PrGT34B is the ortholog of XXT1 and XXT2, the two main xyloglucan (1→6)‐α‐xylosyltransferases in Arabidopsis thaliana. PrGT34C is the ortholog of XXT5 in A. thaliana, which is also involved in the xylosylation of xyloglucans. PrGT34A is an ortholog of a galactosyltransferase from fenugreek (Trigonella foenum‐graecum) that is involved in galactomannan synthesis. Truncated coding sequences of the genes were cloned into plasmid vectors and expressed in a Sf9 insect cell‐culture system. The heterologous proteins were purified, and in vitro assays showed that, when incubated with UDP‐xylose and cellotetraose, cellopentaose or cellohexaose, PrGT34B showed xylosyltransferase activity, and, when incubated with UDP‐galactose and the same cello‐oligosaccharides, PrGT34B showed some galactosyltransferase activity. The ratio of xylosyltransferase to galactosyltransferase activity was 434:1. Hydrolysis of the galactosyltransferase reaction products using galactosidases showed the linkages formed were α‐linkages. Analysis of the products of PrGT34B by MALDI‐TOF MS showed that up to three xylosyl residues were transferred from UDP‐xylose to cellohexaose. The heterologous proteins PrGT34A and PrGT34C showed no detectable enzymatic activity.     

RAPD markers linked to a gene for resistance to pine needle gall midge in Japanese black pine (Pinus thunbergii)

Linkage of RAPD markers to a single dominant gene for resistance to pine needle gall midge was investigated in Japanese black pine (Pinus thunbergii). Three primers that generated linked markers were found after 1160 primers were screened by bulked segregant analysis. The distances between the resistance gene, R, and the marker genes OPC06580, OPD01700, and OPAX192100 were 5.1 cM, 6.7 cM and 13.6 cM, respectively. OPC06580 was in coupling phase to R, whereas OPD01700 and OPAX192100 were in repulsion phase to R. A linkage map for a resistant tree was constructed using 96 macrogametophytes. In linkage analysis, 98 out of 127 polymorphic markers were assigned to 17 linkage groups and six linked pairs. The total length of this map was 1469.8 cM, with an average marker density of 15.6 cM. The genome length was estimated to be 2138.3 cM, and the derived linkage map covered 67.5% of the genome. Although the linked markers OPC06580, OPAX192100, and OPD01700, belonged to the same linkage group, no precise positions were found for OPC06580 or OPD01700. Received: 15 May 1999 / Accepted: 29 July 1999     

Removal of nitrogen during needle senescence in Scots pine (Pinus sylvestris L.)

The concentrations of arginine, protein and total nitrogen (N) and the abundance of¹⁵N were measured in 3-and 4-year-old needles of Scots pine trees fertilized with either 0 (C), 36 (N1) or 73 (N2) kg N ha^-1 year^-1 annually for 22 years (average doses of N). Remaining green needles and needles that were shed were compared and removal of N from total, protein and arginine pools was calculated. Earlier investigations had shown that high arginine concentrations are found in needles of trees that have an excessive N supply (Näsholm and Ericsson 1990). This study aimed to elucidate the fate of the accumulated arginine during needle senescence. It was speculated that a low removal of arginine during senescence would implicate that the primary function of arginine is in N detoxification and not in N storage. Moreover, litter quality would be altered if needles are shed with high concentrations of arginine and this might affect the turnover of N in forest ecosystems. In remaining green needles, the concentration of total N increased with increasing N supply. Protein N concentrations were higher in fertilized trees, but did not differ between the two N treatments. Arginine N was low in C and N1 trees but high in N2 trees. Senescent needles from C and N1 trees had about equal total N concentrations while in N2 trees this concentration was significantly higher. Protein N in senescent needles did not differ between treatments. Arginine N, however, was less than 0.1 mg g^–1 dw in C and N1 trees but was higher than 1.5 mg g^–1 dw in N2 trees. Removal of N was highest in N1 trees followed by C trees while N2 trees removed least N from senescing needles. The high concentration of total N in senescent needles from N2 trees was to a great extent explained by a high arginine concentration.The ¹⁵N value of remaining, green needles was higher (less negative) in N2 trees than in C and N1 trees. The same pattern was found for senescent needles. Comparisons of ¹⁵N values between remaining, green and senescent needles within each treatment showed a significant increase in ¹⁵N for all treatments during senescence possibly indicating losses of N as NH₃ (g) from needles during senescence. It is concluded that arginine, accumulated in response to high N supply, is retranslocated only to a small extent during needle senescence. The ecological and physiological implications of this finding are discussed.     

A mitochondrial DNA minisatellite reveals the postglacial history of jack pine (Pinus banksiana), a broad-range North American conifer

Jack pine (Pinus banksiana Lamb.) is a broadly distributed North American conifer and its current range was covered by the Laurentian ice sheet during the last glacial maximum. To infer about the history and postglacial colonization of this boreal species, range-wide genetic variation was assessed using a new and highly variable minisatellite-like marker of the mitochondrial genome. Among the 543 trees analysed, 14 distinct haplotypes were detected, which corresponded to different repeat numbers of the 32-nucleotide minisatellite-like motif. Several haplotypes were rare with limited distribution, suggesting recent mutation events during the Holocene. At the population level, an average of 2.6 haplotypes and a mean haplotype diversity (H) of 0.328 were estimated. Population subdivision of genetic diversity was quite high with G(ST) and R(ST) values of 0.569 and 0.472, respectively. Spatial analyses identified three relatively homogeneous groups of populations presumably representative of genetically distinct glacial populations, one west and one east of the Appalachian Mountains in the United States and a third one presumably on the unglaciated northeastern coastal area in Canada. These results indicate the significant role of the northern part of the US Appalachian Mountains as a factor of vicariance during the ice age. A fourth distinct group of populations was observed in central Québec where the continental glacier retreated last. It included populations harbouring haplotypes present into the three previous groups, and it had higher level of haplotype diversity per population (H = 0.548) and lower population differentiation (G(ST) = 0.265), which indicates a zone of suture or secondary contact between the migration fronts of the three glacial populations. Introgression from Pinus contorta Dougl. var. latifolia Engelm. was apparent in one western population from Alberta. Altogether, these results indicate that the mitochondrial DNA variation of jack pine is geographically highly structured and it correlates well with large-scale patterns emerging from recent phylogeographical studies of other tree boreal species in North America.     

Genetic variation at allozyme and RAPD markers in Pinus longaeva (Pinaceae) of the White Mountains, California

We compared genetic diversity estimated from allozymes and from random amplified polymorphic DNA (RAPDs) in a sample of 210 Great Basin bristlecone pines (Pinus longaeva Bailey) from three groves in the White Mountains, California, USA. The White Mountains are the most westerly extension of bristlecone pine and home to the oldest known living trees. We assayed two forks of each tree to determine whether they originated from multiple seed caches of the Clark's nutcracker. Despite the limited and fragmented distribution of bristlecone pine, its level of genetic diversity was comparable to that of other pines, but lower than that reported for eastern populations of Great Basin bristlecone pine. Twenty-six of 36 allozymes were polymorphic (p(95) = 38.9%; p = 63.0%), with observed heterozygosity (H(o)) of 0.122 and expected heterozygosity (H(e)) of 0.134. The proportion of the total variation among populations (G(ST)) was only 0.011. The high proportion of trees with multiple stems was not due to germination in seed caches; only six of 210 forked trees had multiple allozyme genotypes. Of the 42 RAPD loci scored, 27 were monomorphic. Genetic diversity for RAPDs was nearly the same as that for allozymes (p(95) = 34.1%, H(e) = 0.130). However, the estimates of diversity and differentiation were much higher (H(e) = 0.321, G(ST) = 0.039) after excluding monomorphic loci.     

Northernmost North American Pinus contorta var. latifolia (lodgepole pine) sociations and vegetation diversity relative to its central range east of the Rocky Mountains

Lodgepole pine (Pinus contorta var. latifolia) stands were sampled in central Yukon, Canada (61.5–64°N latitude), which represented the northernmost 9% of the tree's North American range. Within this area, lodgepole pine occupied only ? 2% of the landscape. This study determined: 1) what forest sociations occurred (i.e. structural dominance‐types); 2) how plant growth form composition and richness differed from the central portion of the species’ geographical range; and 3) if stands were biased towards occurring on more thermally favorable south‐facing slopes. Five lodgepole pine sociations were recognized among 100 relevés: Rhododendron groenlandicum (Labrador tea); Cladonia arbuscula (green reindeer lichen); Calamagrostis purpurascens (purple reedgrass); Hylocomium splendens (stairstep moss) and Alnus viridis (green alder, n = 4 relevés). Rhododendron stands were proportionally more common on low gradient sites and had more total plant cover than the other sociations. Cladonia and Calamagrostis stands were typically associated with dry coarse‐textured soils and warm dry sites, respectively; whereas the composition of the Hylocomium sociation reflected the detrimental influences of atypically dense forest canopies on understory vascular plants. Only the Calamagrostis sociation was unique to the study region. Species richness among common northern lodgepole pine sociations averaged 16–19 taxa per relevé (p > 0.05). Northern compared to central range (n = 1394) relevés were compositionally different based on little overlap of their datasets in the ordination space. Northern vegetation had less (p < 0.001) total plant (129% vs 184%), deciduous shrub (9% vs 26%), broad‐leaved herb (5% vs 25%), and bryophyte (27% vs 54%) cover; had greater macro‐lichen cover (13% vs 5%) and lower floristic richness (11 vs 24 taxa) and was less than half as phytosociological diverse. Lodgepole pine stands in the northernmost portion of their range were not biased towards occurring on south‐facing slopes, which suggested an ecological potential for range expansion.     

Comparative mapping in Pinus: sugar pine (Pinus lambertiana Dougl.) and loblolly pine (Pinus taeda L.)

The majority of genomic research in conifers has been conducted in the Pinus subgenus Pinus mostly due to the high economic importance of the species within this taxon. Genetic maps have been constructed for several of these pines and comparative mapping analyses have consistently revealed notable synteny. In contrast, little genomic research has been conducted on the Pinus subgenus Strobus, even though these pines have strong ecological relevance. We report a consensus genetic linkage map for sugar pine (Pinus lambertiana Dougl.) constructed with 399 single nucleotide polymorphisms markers derived from annotated genes. The map is 1,231 cM in length and organized into 19 linkage groups. Two of the mapping populations were derived from trees that were segregating for the major gene of resistance (Cr1) to Cronartium ribicola, the fungal pathogen responsible for white pine blister rust. The third mapping population was derived from a full-sib cross segregating for partial resistance to white pine blister rust. In addition, we report the first comparative mapping study between subgenera Strobus and Pinus. Sixty mapped markers were found in common between sugar pine and the loblolly pine reference map with 56 of them (93%) showing collinearity. All 19 linkage groups of the sugar pine consensus map coaligned to the 12 linkage groups of the loblolly pine reference map. The syntenic relationship observed between these two clades of pines provides a foundation for advancing genomic research and genetic resources in subgenus Strobus.     

Callus of Pinus roxburghii (Chir pine) and its cytology

Callus was initiated form different vegetative parts of 3 to 5-week-old seedlings of Pinus roxburghii Sargent. Best results were obtained on Murashige and Skoog's revised medium supplemented with 4 mg1⁻¹ NAA or 2,4-D, 1 mg1⁻¹ kinetin and 15% coconut milk. Callus was grown successfully for more than a year without deterioration in its growth. Growth rate studies on the calli form cytoledon explants were undertaken with different concentrations of auxins and cytokinins. Histogenic differentiation of tracheids ocurred in all the calli, with the formation of bordered pits preceding the reticulate thickenings of the tracherary walls. Cells in the 4 to 12-week-old calli were predominantly diploid, though a few polyploid and aneuploid cells were also noticed. Chromosome bridges and laggards were observed in a number of cells.     

Long-term effects of exogenous methyl jasmonate application on Scots pine (Pinus sylvestris) needle chemical defence and diprionid sawfly performance

Scots pine [ Pinus sylvestris L. (Pinaceae)] trees with four different seed origins were exposed to exogenous applications of the elicitor, methyl jasmonate (MeJA), for three consecutive years. We studied the effects of MeJA on needle chemistry (including monoterpenes, sesquiterpenes, and tricyclic resin acids), plant growth, and the performance of two diprionid sawflies, the European pine sawfly ( Neodiprion sertifer Geoffr.) and the common pine sawfly ( Diprion pini L.) (both Hymenoptera: Diprionidae). In general, foliar MeJA application affected the whole range of needle secondary chemistry with significantly higher concentrations of two monoterpenes, β-pinene and limonene, in particular. Furthermore, for some seed origins the growth rates of N. sertifer and D. pini larvae were lower on needles of MeJA-treated plants with either high total terpene or high resin acid concentrations. However, inconsistencies in diprionid sawfly performance within each studied Scots pine origin suggest genetic variance in needle secondary chemistry. The differences between selected seed origins and notably variable responses to MeJA application imply that adaptation of the seed to new conditions may have had an impact on secondary chemistry and, thus, on insect performance. Finally, our results suggest that modification of Scots pine defence by a low-concentration exogenous elicitor affects the production of terpenoids in the newly growing needles, leading to poorer pine sawfly performance in origins with high terpenoid content, while not harming the growth of Scots pine trees.     

Phylogenetic analysis of the hard pines (Pinus subgenus Pinus,Pinaceae) from chloroplast DNA restriction site analysis

Phylogenetic analysis of plastid DNA restriction site and rearrangement mutations suggests a number of major revisions to taxonomy and phylogenetic concepts in the hard pines. Total genomic DNA from 18 species that sampled all nine subsections was digested with 19 restriction enzymes, blotted, and probed with 70% of the Douglas-fir (Pseudotsuga menziesii) chloroplast genome, or, with clones encompassing the entire chloroplast genome of Pinus contorta. A total of 204 site mutations and five rearrangement mutations were generated, of which 126 were phylogenetically informative. Wagner parsimony analyses revealed 11 clades that were strongly supported by bootstrap and decay index analyses. All North American species except P. resinosa formed a distinct monophyletic group that was strongly separated from the Eurasian species. Within the Eurasian clade subsect. Sylvestres was polyphyletic; its Mediterranean species were closely allied with members of sect. Pinea. Sect. Pinea appeared polyphyletic as well; both species of its subsect. Leiophyllae showed a close affinity to Mesoamerican pines of subsect. Oocarpae in sect. Pinus. Within the North American pines subsects. Ponderosae and Oocarpae were polyphyletic. Despite its shallow fossil record, subsect. Contortae emerged as a sister group to all of the North American pines apart from P. resinosa, which was allied with Eurasian species of subsect. Sylvestres. The remaining North American subsections formed two groups: a poorly resolved clade with subsects. Ponderosae and Sabinianae, and sequentially nested clades represented by: P. radiata; P. taeda; representatives of subsects. Oocarpae and Ponderosae from Mesoamerica, and subsect. Leiophyllae. We present estimates of divergence times for each of these major clades based on molecular clocks calibrated using two hard pine fossil observations.     

Paternal cytoplasmic transmission in Chinese pine (Pinus tabulaeformis)

Summary. In this paper, the stages of normal sexual reproduction between pollen tube penetration of the archegonium and early embryo formation in Pinus tabulaeformis are described, emphasizing the transmission of parental cytoplasm, especially the DNA-containing organelles – plastids and mitochondria. The pollen tube growing in the nucellus contained an irregular tube nucleus followed by a pair of sperm cells. The tube cytoplasm contained abundant organelles, including starch-containing plastids and mitochondria. The two sperm cells differed in their volume of cytoplasm. The leading sperm, with more cytoplasm, contained abundant plastids and mitochondria, while the trailing one, with a thin layer of cytoplasm, had very few organelles. The mature egg cell contained a great number of mitochondria, whereas it lacked normal plastids. At fertilization, the pollen tube penetrated into the egg cell at the micropylar end and released all of its contents, including the two sperms. One of the sperm nuclei fused with the egg nucleus, whereas the other one was retained by the receptive vacuole. Very few plastids and mitochondria of male origin were observed around the fusing sperm and egg nuclei, while the retained sperm nucleus was surrounded by a large amount of male cytoplasm. The discharged tube cytoplasm occupied a large micropylar area in the egg cell. In the free nuclear proembryo, organelles of maternal and paternal origins intermingled in the neocytoplasm around the free nuclei. Most of the mitochondria had the same features as those of the egg cell, but some appeared to be from sperm cells and tube cytoplasm. Plastids were obviously of male origin, with an appearance similar to those of the sperm or tube cells. After cellularization of the proembryo, maternal mitochondria became more abundant than the paternal ones and the plastids enlarged and began to accumulate starch. The results reveal the cytological mechanism for paternal inheritance of plastids and biparental inheritance of mitochondria in Chinese pine. Correspondence and reprints: State Key Laboratory of Plant Physiology and Biochemistry, College of Biological Science, China Agricultural University, Beijing 100094, People’s Republic of China.     

Chloroplast microsatellites and mitochondrial nad1 intron 2 sequences indicate congruent phylogenetic relationships among Swiss stone pine (Pinus cembra), Siberian stone pine (Pinus sibirica), and Siberian dwarf pine (Pinus pumila)

We studied the phylogenetic relationships among the three stone pine species, Pinus cembra, P. sibirica, and P. pumila, using chloroplast microsatellites and mitochondrial nad1 intron 2 sequences. The three chloroplast microsatellite loci combined into a total of 18 haplotypes. Fourteen haplotypes were detected in 15 populations of P. cembra and one population of P. sibirica, five of which were shared between the two species, and the two populations of P. pumila comprised four species-specific haplotypes. Mitochondrial intron sequences confirmed this grouping of species. Sequences of P. cembra and P. sibirica were identical, but P. pumila differed by several nucleotide substitutions and insertions/deletions. A repeat region found in the former two species showed no intraspecific variation. These results indicate a relatively recent evolutionary separation of P. cembra and P. sibirica, despite their currently disjunct distributions. The species-specific chloroplast and mitochondrial markers of P. sibirica and P. pumila should help to trace the hybridization in their overlapping distribution area and to identify fossil remains with respect to the still unresolved postglacial re-colonization history of these two species.     

Delayed seed dispersal in Pinus torreyana (Torrey pine)

Summary Torrey pine exhibits a pattern of seed release intermediate between an open and closed-cone species. It was found that the cones open at maturity, but that seed fall from some cones continues for up to 13 years. Approximately 77% of the total seed crop was present in age classes one or more years past the time of seed maturity.Serotiny is thought to evolve in response to fire, and partial serotiny at the population level has been attributed to circumstances in which the strong selective pressure of fire is relaxed. We hypothesize that delayed seed dispersal in Torrey pine is a stable trait which is the result of a pattern of large relatively rare crown fires separated by intervals longer than one generation. The reduced serotiny observed in other conifers on headlands and islands in California seems consistent with our hypothesis.     

Evolution of genome size in pines (Pinus) and its life-history correlates: supertree analyses

Genome size has been suggested to be a fundamental biological attribute in determining life-history traits in many groups of organisms. We examined the relationships between pine genome sizes and pine phylogeny, environmental factors (latitude, elevation, annual rainfall), and biological traits (latitudinal and elevational ranges, seed mass, minimum generation time, interval between large seed crops, seed dispersal mode, relative growth rate, measures of potential and actual invasiveness, and level of rarity). Genome sizes were determined for 60 pine taxa and then combined with published values to make a dataset encompassing 85 species, or 70% of species in the genus. Supertrees were constructed using 20 published source phylogenies. Ancestral genome size was estimated as 32 pg. Genome size has apparently remained stable or increased over evolutionary time in subgenus Strobus, while it has decreased in most subsections in subgenus Pinus. We analyzed relationships between genome size and life-history variables using cross-species correlations and phylogenetically independent contrasts derived from supertree constructions. The generally assumed positive relation between genome size and minimum generation time could not be confirmed in phylogenetically controlled analyses. We found that the strongest correlation was between genome size and seed mass. Because the growth quantities specific leaf area and leaf area ratio (and to a lesser extent relative growth rate) are strongly negatively related to seed mass, they were also negatively correlated with genome size. Northern latitudinal limit was negatively correlated with genome size. Invasiveness, particularly of wind-dispersed species, was negatively associated with both genome size and seed mass. Seed mass and its relationships with seed number, dispersal mode, and growth rate contribute greatly to the differences in life-history strategies of pines. Many life-history patterns are therefore indirectly, but consistently, associated with genome size.     

Phylogeography of the white-breasted nuthatch (Sitta carolinensis): diversification in North American pine and oak woodlands

Pine and oak woodlands are common North American floral communities with distinct regional species composition. The white-breasted nuthatch (Aves: Sitta carolinensis) is a common resident bird of North American pine and oak woodlands, and is distributed continentally across the highly disjunct distribution of these woodlands. We propose three historical hypotheses to explain the evolution of the white-breasted nuthatch in its principal habitat. (i) The species evolved in situ in the regional pine-oak communities and the isolation of populations in these regions is captured in cryptic genetic variation. (ii) Migration of individuals between regions is frequent enough to maintain the widespread distributions and prevent regional divergence. (iii) The species have recently expanded to occupy their current distributions and an insufficient amount of time has passed for divergence to occur. Phylogenetic analysis of mitochondrial DNA (ND2 gene) variation (N = 216) in the white-breasted nuthatch reveals four reciprocally monophyletic clades concordant with the distribution of the regional North American pine and oak woodlands, and supports hypothesis 1 of in situ evolution of populations in the regional pine and oak communities. Within-clade population structure and demographic history are also discussed.     

Endobacteria in some ectomycorrhiza of Scots pine (Pinus sylvestris)

The diversity of cultivable endobacteria associated with four different ectomycorrhizal morphotypes (Suillus flavidus, Suillus variegatus, Russula paludosa and Russula sp.) of Scots pine (Pinus sylvestris) was analysed by restriction fragment length polymorphism profiling of PCR-amplified rDNA intergenic spacer regions and by sequence analyses of 16S rRNA genes. Ectomycorrhizal root tip surface-sterilization methods were developed and assessed for their efficiencies. Bacterial communities from surface-sterilized ectomycorrhizal root tips were different from those of ectomycorrhizal root tips without surface-sterilization for all the morphotypes studied. Endobacteria belonging to the genera Pseudomonas, Burkholderia and Bacillus were isolated from more than one ectomycorrhizal morphotype, whereas species of Rahnella, Janthinobacterium and Rhodococcus were only isolated from the single morphotypes of S. variegatus, R. paludosa and Russula sp., respectively. Some of the isolated endobacteria utilized fungal sugars more readily than typical plant sugars in carbon utilization assays.