Inferring colonization history from analyses of spatial genetic structure within populations of Pinus strobus and Quercus rubra

Many factors interact to determine genetic structure within populations including adult density, the mating system, colonization history, natural selection, and the mechanism and spatial patterns of gene dispersal. We examined spatial genetic structure within colonizing populations of Quercus rubra seedlings and Pinus strobus juveniles and adults in an aspen-white pine forest in northern Michigan, USA. A 20-year spatially explicit demographic study of the forest enables us to interpret the results in light of recent colonization of the site for both species. We assayed 217 Q. rubra seedlings and 171 P. strobus individuals at 11 polymorphic loci using nine allozyme systems. Plant genotypes and locations were used in an analysis of spatial genetic structure. Q. rubra and P. strobus showed similar observed levels of heterozygosity, but Q. rubra seedlings have less heterozygosity than expected. Q. rubra seedlings show spatial genetic clumping of individuals on a scale to 25 m and levels of genetic relatedness expected from the clumped dispersion of half-siblings. In contrast, P. strobus has low levels of genetic relatedness at the smallest distance class and positive spatial genetic structure at scales < 10 m within the plot. The low density of adult Q. rubra outside the study plot and limited, spatially clumped rodent dispersal of acorns is likely responsible for the observed pattern of spatial genetic structure and the observed heterozygote deficit (i.e. a Wahlund effect). We attribute weaker patterns observed in P. strobus to the longer dispersal distance of seeds and the historical overlap of seed shadows from adults outside of the plot coupled with the overlap of seed shadows from younger, more recently established reproductive adults. The study demonstrates the utility of long-term demographic data in interpreting mechanisms responsible for generating contemporary patterns of genetic structure within populations.     

Microsatellite DNA analysis of genetic effects of harvesting in old-growth eastern white pine (Pinus strobus) in Ontario, Canada

Microsatellite DNA markers from 13 simple sequence repeat (SSR) loci were used to compare genetic diversity between preharvest pristine and postharvest residual gene pools of two adjacent virgin, old-growth ( approximately 250 years) stands of eastern white pine (Pinus strobus L.) in Ontario. There was concurrence in genetic diversity changes in the postharvest gene pools of the two stands. The total and mean numbers of alleles detected in each stand were reduced by approximately 26% after tree density reductions of approximately 75%. Approximately 18 and 21% of the low-frequency (0. 25 > P > or = 0.01) alleles and 76 and 92% of the rare (P < 0.01) alleles were lost from residual stands A and B, respectively, after harvesting. Multilocus gametic diversity was reduced by 38 and 85% and genotype additivity by approximately 50% in the residual stands after harvesting. Latent genetic potential of each stand was reduced by approximately 40%. Although heterozygosity was reduced (1-5%) in the postharvest residual stands, the reductions were not substantial and not comparable to those using other genetic diversity measures. The reductions in genetic diversity measures were slightly higher than those theoretically expected in postbottleneck populations according to drift theory. In the absence of substantial gene migration that could ameliorate the genetic losses, the ability of the postharvest white pine gene pools to adapt to changing environmental and disease conditions may have been compromised. The microsatellite DNA results for genetic effects of harvesting in old-growth eastern white pine stands were similar to those that we reported earlier from allozyme analysis (Buchert et al. 1997). The results indicate that silvicultural practices should ensure that the gene pools of remaining pristine old-growth stands are reconstituted in the regenerating stands.     

Microsatellite analysis reveals genetically distinct populations of red pine (Pinus resinosa, Pinaceae)

Red pine (Pinus resinosa Ait.) is an ecologically and economically important forest tree species of northeastern North America and is considered one of the most genetically depauperate conifer species in the region. We have isolated and characterized 13 nuclear microsatellite loci by screening a partial genomic library with di-, tri-, and tetranucleotide repeat oligonucleotide probes. In an analysis of over 500 individuals representing 17 red pine populations from Manitoba through Newfoundland, five polymorphic microsatellite loci with an average of nine alleles per locus were identified. The mean expected and observed heterozygosity values were 0.508 and 0.185, respectively. Significant departures from Hardy-Weinberg equilibrium with excess homozygosity indicating high levels of inbreeding were evident in all populations studied. The population differentiation was high with 28-35% of genetic variation partitioned among populations. The genetic distance analysis showed that three northeastern (two Newfoundland and one New Brunswick) populations are genetically distinct from the remaining populations. The coalescence-based analysis suggests that "northeastern" and "main" populations likely became isolated during the most recent Pleistocene glacial period, and severe population bottlenecks may have led to the evolution of a highly selfing mating system in red pine.     

Adventitious root initiation in hypocotyl and epicotyl cuttings of eastern white pine (Pinus strobus) seedlings

The present paper reports results of experiments to develop a system for studying adventitious root initiation in cuttings derived from seedlings. Hypocotyl cuttings of 2-week-old eastern white pine (Pinus strobus L.) seedlings were treated for 5 min with 0, 100, 200, 300, 400, 500 or 600 mg l^?1 (0, 0.54, 1.07, 1.61, 2.15, 2.69 or 3.22 mM) 1-naphthaleneacetic acid (NAA) to determine the effect on root initiation. The number of root primordia per cutting was correlated with NAA concentration and the square of NAA concentration. Thus, the number increased from less than one per cutting in the 0 NAA treatment to approximately 40 per cutting at 300 mg l-1 NAA, above which no substantial further increase was observed. The larger number of root primordia formed in response to increasing concentrations of NAA was due to the formation of primordia over a larger proportion of the hypocotyls. Histological analysis of the timing of root primordium formation in hypocotyl cuttings revealed three discernible stages. Progression through these stages was relatively synchronous among NAA-treated hypocotyl cuttings and within a given cutting, but variation was observed in the portion of different cuttings undergoing root formation. Control-treated hypocotyl cuttings formed root primordia at lower frequencies and more slowly than NAA-treated cuttings, with fewer primordia per cutting. Epicotyl cuttings from 11-week-old seedlings also formed adventitious roots, but more slowly than hypocotyl cuttings. NAA treatment of epicotyl cuttings caused more rapid root initiation and also affected the origin of adventitious roots in comparison with nontreated cuttings. NAA-treated epicotyl cuttings formed roots in a manner analogous to that of the hypocotyl cuttings, directly from preformed vascular tissue, while control-treated epicotyl cuttings first formed a wound or callus tissue and subsequently differentiated root primordia within that tissue. This system of inducing adventitious roots in pine stem cuttings lends itself to studying the molecular and biochemical steps that occur during root initiation and development.     

Hydrogen fluoride effects on plasma membrane composition,ATPase activity and cell structure in needles of eastern white pine (Pinus strobus) seedlings

Eastern white pine (Pinus strobus L.) seedlings were grown in controlled environment growth cabinets and fumigated with 0.4 and 1.6 g m^–3 hydrogen fluoride for 2–28 days. Plasma membranes were isolated from needles of treated and control seedlings and their chemical composition and ATPase activity examined to determine early effects of hydrogen fluoride action. In plants treated for 2 days with both fluoride levels, ratios of plasma membrane free sterols:phospholipids and sterols:proteins were drastically higher than ratios in control plants. Seedlings treated with hydrogen fluoride for 8 days contained plasma membranes with elevated phospholipid:protein and sterol:protein ratios and their plasma membrane ATPase activity was higher than that of control plants. Prolonged, 28-day hydrogen fluoride treatment with 1.6 g m^–3 level was the only treatment which produced a drastic inhibition of plasma membrane ATPase activity. During the initial stages of hydrogen fluoride treatment, treated cells did not show alterations of ultrastructure which were previously shown in cells of plants treated with soil applied sodium fluoride. The results of the present study indicate that the plasma membranes may be among the initial sites of hydrogen fluoride injury to plants as well as initial sites of defense reaction.     

Genetic diversity and spatial genetic structure of Pinus strobus (Pinaceae) across an island landscape inferred from allozyme and cpDNA markers

The Beaver Island Archipelago (BIA) provides a model system to address the impact of long-term isolation on genetic diversity and gene flow. Low lake levels are assumed to have caused the BIA to be attached to mainland Michigan for at least 4000 years (10000 yr B.P.- 6000 yr B.P.), eventually, rising lake levels would have kept the islands isolated since 6000 yr B.P. If the island populations of a plant species in the BIA were indeed once continuous with the mainland of Michigan, then we would expect similar levels of genetic diversity in populations of such a species on the islands vs. the mainland. We compared levels of allozyme genetic diversity of 20 plots of Pinus strobus in the BIA with two mainland populations in northern Michigan. In addition, if pollen is a primary agent of gene flow across islands, a low degree of allozyme differentiation among the island populations of P. strobus in the BIA would be evident. Furthermore, since seed dispersal is more limited than pollen dispersal in P. strobus, a more pronounced spatial genetic structure (SGS) is expected in allozymes than in cpDNA markers. To gain insights on the pattern of seed and pollen dispersal among the 20 plots, we further analyzed spatial autocorrelation using Moran's I-statistics for both data sets [biparentally inherited, allozymes and paternally inherited, cpDNA microsatellites (cpDNA SSR)]. We found a similar level of allozyme variability in both the BIA (mean H _e = 0.080) and the two mainland populations (mean H _e = 0.078). As predicted, we observed a low but significant degree of genetic divergence among populations for allozymes (mean F _ST = 0.033 across 20 plots). Our allozyme-based SGS analysis revealed significant evidence of SGS (i.e. isolation-by-distance; slope β = ?0.194 from regression analysis of observed averaged Moran's I values against the logarithm of the upper bound of six distance classes). In contrast, little evidence of SGS was found in cpDNA SSR data across the BIA (β = 0.013). These results suggest that although gene flow via seed dispersal is somewhat limited, pollen flow has been sufficient to maintain genetic diversity and prevent differentiation across the island landscape over several thousand years of isolation.     

Assessing genetic diversity and structure of fragmented populations of eastern white pine (Pinus strobus) and western white pine (P. monticola) for conservation management

Aims Many pine populations in Canada have fragmented distributions resulting from the effects of glaciations, overharvesting and white pine blister rust infections. Forest fragmentation can modify gene flow and reduce genetic diversity. Selective logging can reduce the density of trees, thereby altering mating patterns and increasing inbreeding. The hypothesis of the present study is that forest fragmentation will not increase inbreeding and will have no effect on genetic diversity parameters in the Canadian Pinus moniticola and P. strobus populations targeted because of (i) the long life span of the pine species, (ii) outbreeding and self-incompatibility of P. monticola and P. strobus and (iii) wind pollination resulting in high gene flow among populations. We studied the genetic diversity of P. strobus across its range in Canada, and we completed a detailed analysis of the genetic structure of P. monticola populations from western Canada using microsatellites genetic markers.Methods Seed samples from 10 P. monticola populations and 10 P. strobus populations were collected from western and eastern Canada, respectively. The mother trees included in seed lots were representative of each stand. Genomic DNA extracted from each sample was amplified with microsatellite primers. The intra- and interpopulation genetic diversity parameters were assessed using Popgene and Genepop softwares and the genetic distances among populations within each species using the PowerMarker software.Important findings Pinus monticola and P. strobus exhibited moderate to high genetic diversity. Also, both species showed low levels of inbreeding despite the geographic isolation and small stand size. Gene flow estimates were high and population differentiation values were relatively low for these fragmented forest sites.     

Microsatellite analysis of North American wapiti (Cervus elaphus) populations

Eleven populations of wapiti (Cervus elaphus) were analysed for genetic diversity using 12 microsatellite loci. Samples were taken from Vancouver Island, British Columbia; Burwash and French River herds in Ontario; Ya Ha Tinda Ranch, Alberta; and Banff, Elk Island, Jasper, Kootenay, Riding Mountain, Yellowstone and Yoho National Parks. Overall, wapiti populations have on average three to four alleles per locus and an average expected heterozygosity that ranged from 25.75 to 52.85%. The greatest genetic distances were observed between the Vancouver population and all other populations. Using the assignment test, Roosevelt wapiti (C. e. roosevelti Merriam 1897) assigned only to the Vancouver Island population. The distance and assignment values suggest a divergence of the Roosevelt wapiti from other populations and support the subspecific status for the Vancouver Island population. No evidence was found for the existence of unique Eastern wapiti (C. e. canadensis Erxleben 1777) in the Burwash or French River herds in Ontario. The overlapping distribution of genotypes from indigenous populations from Riding Mountain, Elk Island and Yellowstone National Parks suggests that wapiti were once a continuous population before settlers decimated their numbers. The lack of differentiation between these populations raises questions about the status of Manitoban (C. e.manitobensis Millais 1915) and Rocky Mountain (C. e.nelsoni Bailey 1935) subspecies.     

Genetic analysis of Pinus strobus and Pinus monticola populations from Canada using ISSR and RAPD markers: development of genome-specific SCAR markers

Pinus is the largest genus of conifers, containing over 100 species and is also the most widespread genus in the Northern Hemisphere. Pinus monticola and P. strobus are two closely related and economically important species in Canada. Morphological and allometric characteristics have been used to assess genetic variation within these two species but these markers are not reliable due to ecological variations. The purpose of the present study was to determine the level of genetic diversity within and among Canadian populations from the two species using molecular markers and to identify and characterize genome-specific inter-simple sequence repeats (ISSR) and random amplified polymorphic DNA (RAPD) markers. The level of genetic variation among populations was much lower for P. monticola than P. strobus. For both species, the among population variation values were smaller than within population variation. The populations from P. monticola were more closely genetically related than populations from P. strobus based on ISSR and RAPD analyses. Six ISSR and four RAPD markers specific to either P. monticola or P. strobus were cloned and sequenced. Primer pairs flanking these specific sequences were designed and genome specific SCAR markers for P. monticola and P. strobus were developed and characterized.     

Fine-scale spatial genetic structure and dispersal among spotted salamander (Ambystoma maculatum) breeding populations

We examined fine-scale genetic variation among breeding aggregations of the spotted salamander (Ambystoma maculatum) to quantify dispersal, interpopulation connectivity and population genetic structure. Spotted salamanders rely on temporary ponds or wetlands for aggregate breeding. Adequate breeding sites are relatively isolated from one another and field studies suggest considerable adult site fidelity; therefore, we expected to find population structure and differentiation at small spatial scales. We used microsatellites to estimate population structure and dispersal among 29 breeding aggregations in Tompkins County, New York, USA, an area encompassing 1272 km(2). Bayesian and frequency-based analyses revealed fine-scale genetic structure with two genetically defined demes: the North deme included seven breeding ponds, and the South deme included 13 ponds. Nine ponds showed evidence of admixture between these two genetic pools. Bayesian assignment tests for detection of interpopulation dispersal indicate that immigration among ponds is common within demes, and that certain populations serve as sources of immigrants to neighbouring ponds. Likewise, spatial genetic correlation analyses showed that populations < or = 4.8 km distant from each other show significant genetic correlation that is not evident at higher scales. Within-population levels of relatedness are consistently larger than expected if mating were completely random across ponds, and in the case of a few ponds, within-population processes such as inbreeding or reproductive skew contribute significantly to differentiation from neighbouring ponds. Our data underscore the importance of these within-population processes as a source of genetic diversity across the landscape, despite considerable population connectivity. Our data further suggest that spotted salamander breeding groups behave as metapopulations, with population clusters as functional units, but sufficient migration among demes to allow for potential rescue and recolonization. Amphibian habitats are becoming increasingly fragmented and a clear understanding of dispersal and patterns of population connectivity for taxa with different ecologies and life histories is crucial for their conservation.     

Microsatellite markers reveal spatial genetic structure of Tetranychus urticae (Acari: Tetranychidae) populations along a latitudinal gradient in Europe

The genetic structure of populations of the two-spotted spider mite Tetranychus urticae was investigated along a south–north European transect spanning from southern France to The Netherlands. Mites were collected on Urtica dioica in 6 sampling zones. Microsatellite variation at 5 loci revealed considerable genetic variation with an average heterozygozity of 0.49. Significant heterozygote deficiency was found in 7 populations out of the 18 samples analyzed and one of them was completely monomorphic. Tetranychus urticae populations show some level of genetic structuring. First, genetic differentiation between localities (F _ST estimates) was significant for all comparisons. Second, the analysis of molecular variance, AMOVA, indicates that there is an effect, albeit low (9%), of the locality in accounting for allele frequency variance. Geographic distance emerges as a factor responsible for this genetic structure. The results are discussed in relation to the biological features of the species and the known patterns of migration. Related agronomical issues are addressed.     

Genetic diversity in populations of Cronartium ribicola in plantations and natural stands of Pinus strobus

Genetic diversity was studied in 22 populations of the white pine blister rust fungus Cronartium ribicola from natural stands and plantations of eastern white pine, Pinus strobus. Pseudo-allelic frequencies were estimated at each of 7 putative RAPD loci by scoring for presence or absence of amplified fragments in dikaryotic aecidiospores. Analysis of genetic distance between all pairs of populations did not reveal any trend with regard to geographic origin or type of white pine stand. In addition, when hierarchical population structure was analysed, total genetic diversity (H _s =0.214) was mostly attributable to diversity within populations (H _s =0.199; AMOVA _st =0.121, P<0.01). Genetic diversity of populations relative to region of origin (east, centre, and west) or type of stand (natural stands vs plantations) was not significantly different from zero (P>0.10) Nevertheless, a significant proportion of genetic differentiation was found between populations within region or stand type (F _st =0.114; _sc =0.132, P<0.001). This result indicates that some population structure exists but that it appears to be independent of region of origin or type of stand. At least for 2 populations from white pine plantations, it appears possible that a recent introduction of a limited number of propagules was responsible for low levels of genetic diversity. We interpret these results as meaning that either long-distance dispersal is taking place between populations more than 1000 km apart or that these populations share a common recent ancestor. In addition, we suggest that C. ribicola may still be expanding its distribution by colonizing new plantations.     

Hierarchical genetic and spatial structure among varieties and populations of Hymenaea stigonocarpa (Fabaceae) in Brazilian savannah

Tree Genetics & Genomes - Tree breeders must almost always address multiple traits. That entails choosing selection methods and weighing up known or assumed economic-worth functions for traits....     

Parenchyma cells of secondary phloem in Pinus strobus

Summary Parenchyma cells of the secondary phloem in Pinus strobus have all the cellular organelles common in other plant cells. They have mitochondria, endoplasmic reticulum, ribosomes, dictyosomes, and plastids. Parenchyma cells are very conspicuous because of their organic inclusions, starch and lipids. Plasmodesmata in transverse and tangential walls of axial parenchyma cells and in end walls of ray parenchyma cells are regularly distributed and of uniform size, about 500 Å in diameter. In radial walls of axial parenchyma cells and horizontal walls of ray parenchyma cells plasmodesmata are located in primary pit-fields; there they are of variable size and often divided into several branches. The branches are confluent into a median nodule. Perforation of the transverse wall between two axial parenchyma cells and the resultant union of the cellular material of the two connected cells is reported.This research has been supported by NSF Grant GB 3193.     

Characterization of pollen tube development in Pinus strobus (Eastern white pine) through proteomic analysis of differentially expressed proteins

The differentially expressed proteins in pollen tubes indicate their specific roles in this stage of male gametophyte development. To isolate these proteins, 2-DE was done using ungerminated pollen and 2-day-old pollen tubes of Pinus strobus. Results show that 645 and 647 protein spots were clearly resolved from pollen grains and pollen tubes, respectively. Thirty-eight protein spots were expressed only in pollen tubes, while 19 increased in intensity. MALDI-TOF MS was used to generate tryptic peptide masses that were submitted to Mascot for identification. Of the differentially expressed proteins, 12% matched with hypothetical proteins, 33% did not hit any protein, and for the 55%, a putative function was assigned based on similarity of sequences with previously characterized proteins. Therefore, pollen tube development can be characterized by the cellular activities that involve metabolism, stress/defense response, gene regulation, signal transduction, and cell wall formation. This study expands our understanding of the changes in protein expression associated with pollen tube development and provides insights into the molecular programs that separate the development of the pollen tubes from pollen grains. This is the first report that describes a global analysis of differentially expressed proteins from the pollen tube of any seed plant.     

Hydrogen fluoride effects on plasma membrane composition and ATPase activity in needles of white pine (Pinus strobus) seedlings pretreated with 12 h photoperiod

 Eastern white pine (Pinus strobus L.) seedlings were pretreated with 12 h photoperiod to induce dormancy. Dormant plants were fumigated with 0.5 ppb (0.4 μg m^–3) or 2.0 ppb (1.6 μg m^–3) hydrogen fluoride (HF) for 2 – 28 days. Plasma membranes were isolated from needles of treated and control seedlings to determine their chemical composition and ATPase activity. For all analyses, only those plants which did not show needle necrosis were selected. The amount of plasma membrane phospholipid expressed on a plasma membrane protein basis was higher after 2 days in the 0.5 ppb HF treatment as compared to controls. After 2 days of 2.0 ppb HF treatment as well as after 8 and 28 days of both HF treatments phospholipid to protein ratios in fluoride treated seedlings were lower as compared to control levels. A decrease in sterol levels could be observed after 2 days in both HF treatments. A large increase in the ratio of sterols to proteins was observed in plasma membranes of eastern white pine seedlings treated with 0.5 ppb HF for 28 days. Increased sterol to phospholipid ratios were observed after 8 and 28 days in 0.5 ppb and after 2 and 8 days of 2.0 ppb HF treatment. A decrease in ATPase activity was observed after 8 days with both fluoride treatments. Drastic increase of ATPase activity was observed after 28 days of HF treated plants. Observed changes of sterol and phospholipid levels after only 2 days of fumigation suggest early fluoride effects on plasma membrane composition during plant dormancy. Received: 25 October 1995 / Accepted: 24 May 1996     

Radiation and soil temperature interactions on the growth and physiology of eastern white pine ( Pinus strobus L.) seedlings

A greenhouse experiment was set up during one growing season to test the hypothesis that soil temperature controls a significant part of the light response of eastern white pine (Pinus strobus L.) seedlings that is observed in the field. The experimental design was a three by three factorial split-plot design, with three levels of light availability: 10%, 40% and 80% of full light; and three levels of soil temperature: 16 °C, 21 °C, and 26 °C in the soil at midday. The results show significant interactions between light and soil temperature factors on several variables (gas exchange, root growth, leaf-mass ratio and leaf–mass per unit area), but not on shoot dry mass. These interactions indicate that, in the field, a significant proportion of the light response of young eastern white pine could result from changes in soil temperature, especially under conditions of limiting water availability. Our results suggest that soil temperature must be taken explicitly into account as a driving variable when relating the growth of young eastern white pine to photosynthetic radiation.