Comparative phylogeography of four component species of deciduous broad-leaved forests in Japan based on chloroplast DNA variation

A phylogeographic study of four tree species (Padus grayana, Euonymus oxyphyllus, Magnolia hypoleuca, and Carpinus laxiflora) growing in Japanese deciduous broad-leaved forests was conducted based on chloroplast DNA (cpDNA) variations. Using nucleotide sequences of 702–1,059 bp of intergenic spacers of cpDNA, 20, 27, eight, and eight haplotypes were detected among 251, 251, 226, and 262 individuals sampled from 67, 79, 75, and 71 populations of the above species, respectively. The geographical pattern of the cpDNA variations was highly structured in each species, and the following three regional populations were genetically highly differentiated among all four species: (1) the Sea of Japan-side area, (2) the Kanto region, and (3) southwestern Japan. Based on some interspecific similarities among the phylogeographic patterns, the following migration scenario of Japanese deciduous broad-leaved forests was postulated. During the last glacial maximum (LGM), the forests were separately distributed in six regions. After LGM, as the climate warmed, the forests in eastern Japan separately expanded from each of the refugia along the Sea of Japan-side or along the Pacific Ocean-side. In contrast, those in southwestern Japan retreated and moved to high altitudes from each of the continuous forests.     

Pseudo-nitzschia pungens (Bacillariophyceae): A cosmopolitan diatom species?

Genetic, reproductive and morphological variation were studied in 193 global strains of the marine diatom species Pseudo-nitzschia pungens (Grunow ex Cleve) Hasle to assess potential intraspecific variation and biogeographic distribution patterns. Genetic differentiation between allo- and sympatric strains was investigated using the ITS1–5.8S–ITS2 rDNA region. Three ITS clades were found. Clones of opposite mating type were sexually compatible within clades I or II, and viable F1 hybrid offspring were produced in crosses between them. The molecular differences between these clades were correlated with slight but consistent morphological differences. At present, nothing can be said about morphology and mating behavior for clade III clones because only ITS data were available. The three ITS clades showed different geographic distributions. Clade II was restricted to the NE Pacific, whereas clones belonging to clade III originated from geographically widely separated areas (Vietnam, China and Mexico). ITS clade I was recovered in all locations studied: the North Sea (Belgium, The Netherlands, France), the eastern and western N Atlantic (Spain, Canada), the NW and S Pacific (Japan, New Zealand) and the NE Pacific (Washington State). Clade I thus appears to be globally distributed in temperate coastal areas and provides the first strong evidence to date for the global distribution of a biologically, genetically and morphologically defined diatom species.     

Chloroplast DNA phylogeography of <Emphasis Type="Italic">Pedicularis</Emphasis> ser. <Emphasis Type="Italic">Gloriosae</Emphasis> (Orobanchaceae) in Japan

Phylogeographic analyses using chloroplast DNA (cpDNA) variation were performed for Pedicularis ser. Gloriosae (Orobanchaceae). Eighty-one plants of 18 populations of 6 species (P. gloriosa, P. iwatensis, P. nipponica, P. ochiaiana, P. sceptrum-carolinum and P. grandiflora) were analyzed. Fifteen distinct haplotypes were identified based on six cpDNA regions: the intergenic spacer between the trnT and trnL 3′exon, trnL 3′exon-trnF, atpB-rbcL, accD–psaI, the rpl16 intron and the trnK region (including the matK gene). Via phylogenetic analyses of the haplotypes, two continental species, P. sceptrum-carolinum and P. grandiflora, were placed at the most ancestral position in the trees. The former species is widely distributed in the Eurasian continent, and the latter is distributed in Far East Asia. Two robust major cpDNA clades (clades I and II) were revealed in the Japanese archipelago, although the statistical values of monophyly of these clades were weak. Clade I included the haplotypes (A-1, A-2, B-1, B-2 and J) of three species (P. gloriosa, P. iwatensis and P. ochiaiana), and Clade II included seven haplotypes (C-D, E-1, E-2 and F-H) of P. nipponica. These results suggest that this series originated on the Eurasian continent and that subsequently populations at the eastern edge of the continent differentiated into the two Japanese lineages. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Internal transcribed spacer haplotype diversity and their geographical distribution in Dasyscyphella longistipitata (Hyaloscyphaceae, Helotiales) occurring on Fagus crenata cupules in Japan

Phylogeographic analysis of Dasyscyphella longistipitata (Hyaloscyphaceae, Helotiales), which occurs on decaying Fagus crenata cupules, was carried out. The ITS-5.8S regions of the 120 isolates from 12 sites in areas across Japan revealed 38 haplotypes, the majority of which comprised the haplotype designated H12 (42.5%). H12 was found in isolates from all sites, followed by H28 and H10, which were detected in isolates from 10 and 7 sites, respectively. Thirty-two haplotypes were obtained in single isolates. In the haplotype network, H12 was the root, and it formed interior clades with H28 and H10. Genetic diversity was higher in northern and southern Japan, but genetic distance was not correlated with geographical distance, nor with the phylogenetic clades of F. crenata. Therefore, it was concluded that D. longistipitata forms a genetic continuum that covers all of the areas in which it is distributed in Japan, with variations being generated in local populations from the major haplotypes.     

Differences in beech (Fagus crenata) regeneration between two types of Japanese beech forest and along a snow gradient

Differences in beech (Fagus crenata) regeneration were quantitatively investigated using power function analysis for the size–class (diameter at breast height, DBH) distribution and juvenile-to-canopy tree (J/C) ratio along a snow gradient throughout Japan. In snowy areas, all species combined, as well as F. crenata alone, showed constant regeneration, with parameter b≈−1.6 for the power function y=ax ^b (x=DBH, y=density), which is related to the DBH–class distribution. The good fit of the data to the function suggests that beech regenerates constantly with self-thinning patch dynamics. Parameter a, which indicates the abundance of small trunks, was high. Furthermore, the mean J/C ratio was ≈8, i.e., each parent beech tree produced eight juveniles. These results suggest that beech regenerates constantly with gap dynamics in snowy beech forests on the Japan Sea side of Japan (snowy). However, the fit of F. crenata was lower and nonsignificant in some forests in less snowy areas, despite the high fit of all species combined. In these areas, the mean of a was low, and b was often near zero for F. crenata regressions. These results suggest that the abundance of beech was low, and self-thinning was not evident because of the initial low abundance. Moreover, the mean J/C ratio was <1.0, suggesting that juvenile density was lower than that of canopy trees. Thus, the regeneration of F. crenata on the Pacific Ocean side of Japan (less snowy) is rather sporadic. Less snowy conditions may promote seed desiccation, predation of beechnuts and seedlings, and water stress. Lower F. crenata density may also reduce predator satiation and wind pollination.     

Intraspecific Genetic Diversity of Undaria pinnatifida in Japan, based on the Mitochondrial cox3 Gene and the ITS1 of nrDNA

The intraspecific genetic diversity of the kelp Undaria pinnatifida (Harvey) Suringar (Laminariales, Phaeophyceae) was investigated using DNA sequences of the mitochondrial cytochrome oxidase subunit 3 (cox3) gene and internal transcribed spacer 1 (ITS1) of nuclear ribosomal DNA in plants collected from 21 localities along the Japanese coast between 2001 and 2003. Morphological variation was also examined and compared with the genetic diversity. Cox3 analyses of 106 plants revealed 9 haplotypes (I–IX) that differed from each other by 1–7 bp (all synonymous substitutions). Haplotype I was distributed in Hokkaido and the northern Pacific coast of Honshu, while haplotype III was found along the Sea of Japan coast of Honshu. Other types were found along the central and southern coast of Honshu. ITS1 analyses of 42 plants revealed 0–1.7% nucleotide differences, but plants from the Sea of Japan coast and northern Japan had similar sequences. The lower genetic differentiation along the Sea of Japan and northern coasts might be due to the recent establishment (after the middle of the last glacial period) of the Sea of Japan flora. The cox3 haplotype of cultivated plants was found in natural populations occurring close to cultivation sites (Naruto, Tokushima Pref., and Hokutan, Hyogo Pref.). This suggests that cultivated plants possibly escaped and spread or crossed with plants of natural populations. Morphological analyses of variation in 10 characters were conducted using 66 plants. The results showed no significant local variation owing to the wide variation in each population and did not support any forma previously described. No correlations between the morphological characters and cox3 haplotypes were detected.     

Evolution of the Euphrasia transmorrisonensis complex (Orobanchaceae) in alpine areas of Taiwan

Aims To unravel isolation and differentiation of the genetic structure of the Euphrasia transmorrisonensis complex, a showy herb, among alpine regions of mountain peaks in subtropical Taiwan and to infer its evolutionary history. Location Alpine ecosystems of high‐montane regions of Taiwan. Methods Phylogenetic analyses of the trnL intron and the trnL–trnF intergenic spacer of chloroplast (cp) DNA, and the intertranscribed spacer (ITS) of nuclear ribosomal (nr) DNA between 18S and 26S were carried out on 18 populations of the E. transmorrisonensis complex in Taiwan. Results In total, 10 haplotypes for cpDNA and 14 haplotypes for nrDNA were detected. Three population groups located in the northern, north‐eastern, and south‐central regions of the Central Mountain Range (CMR) were revealed according to the frequencies of haplotypes and haplotype lineages of nrDNA. Balancing selection might have played a role in the evolution of Euphrasia in Taiwan. Main conclusions By integrating the spatial‐genetic patterns of cpDNA and nrDNA, two possible evolutionary histories of Euphrasia in Taiwan were inferred. The favourable hypotheses for interpreting the data suggest at least three origins of the E. transmorrisonensis complex in Taiwan, corresponding to each nuclear lineage in the northern (II), northern/north‐eastern (I), and central/southern regions (III) with subsequent hybridization between lineages I and II and lineages II and III. These lineage boundaries are strengthened by the finding that haplotypes of C derived from cpDNA were found in the geographical region of lineage II of nrDNA, while haplotypes of A derived from cpDNA were found in the region of lineage III of nrDNA. Thus, the origin of chloroplasts exclusive to lineages II and III supports their long‐term isolation from one another.     

Extensive Introgression of Mitochondrial DNA Found between two Genetically Divergent Forms of Threespine Stickleback, Gasterosteus Aculeatus, around Japan

The genetic structure and phylogenetic relationships of Gasterosteus aculeatus around Japan were investigated for 15 populations of the Japan Sea and Pacific Ocean forms previously detected by allozyme analyses. PCR-RFLP analysis of mtDNA ND5/6 gene indicated two major divergent clades: (1) Alaskan population and (2) Russian and Japanese populations. Japanese populations were clearly subdivided into G. aculeatus populations and hariyo (G. aculeatus leiurus). However, the relationships among Japanese G. aculeatus populations did not support the genetic divergence between Japan Sea and Pacific Ocean forms shown by allozyme analyses. This suggests that mtDNA genetype similarity between the two forms could have resulted from complete replacement of mtDNA from one form to the other. MtDNA introgression is possible in two direction: from Japan Sea form to Pacific Ocean form and the reverse. The haplotype composition of each population observed in the present study suggests that the second direction is more likely. The migrant number of females per generation (Nm) between populations of the two forms was remarkablely large (9.083–infinite), suggesting that gene flow is taking place between the populations at the present time or might have occurred until recent years.     

Regeneration dynamics, causal factors, and characteristics of pacific ocean-type beech (Fagus crenata) forests in Japan: A review

This paper reviews the differences in the distribution and regeneration ofFagus crenata between two types of Japanese beech forests, the Japan Sea (JS)-type and the Pacific Ocean (PO)-type, and discusses the causal factors and characteristics of these forests, particularly the PO-type.F. crenata in PO-type forests regenerates sporadically rather than constantly, whereas regeneration in the JS-type forests is relatively constant with gap dynamics.F. crenata has dominated in snowy areas both in the past, after the last glacial age, when there was less human disturbance, and in the present. Snow accumulation facilitates beech regeneration in snowy JS-type forests, but not in the less snowy PO-type. Snow protects beechnuts from damage caused by rodents, desiccation, and freezing. In addition, snow suppresses dwarf bamboo in the spring, thus increasing the amount of sunlight available for beech seedlings on the forest floor. Snow also supplies melt water during the growing season and limits the distribution of herbivores. Moreover, snow reduces the number of forest fires during the dry winter and early spring seasons. The low densities ofF. crenata impede its regeneration, because disturbed wind pollination lowers seed fertility and predators are less effectively satiated. In snowy JS-type beech forests,F. crenata dominates both at the adult and the juvenile stages because it regenerates well, while other species are eliminated by heavy snow pressure. On the less snowy PO-side, deciduous broad-leaved forests with various species are a primary feature, althoughF. crenata dominates because of its large size and long lifespan.     

Phylogeography of Pulsatilla vernalis (L.) Mill. (Ranunculaceae): chloroplast DNA reveals two evolutionary lineages across central Europe and Scandinavia

Aim The aim of this study was to test hypotheses regarding some of the main phylogeographical patterns proposed for European plants, in particular the locations of glacial refugia, the post‐glacial colonization routes, and genetic affinities between southern (alpine) and northern (boreal) populations. Location The mountains of Europe (Alps, Balkans, Carpathians, Central Massif, Pyrenees, Scandinavian chain, Sudetes), and central European/southern Scandinavian lowlands. Methods As our model system we used Pulsatilla vernalis, a widely distributed European herbaceous plant occurring both in the high‐mountain environments of the Alps and other European ranges and in lowlands north of these ranges up to Scandinavia. Based on a distribution‐wide sampling of 61 populations, we estimated chloroplast DNA (cpDNA) variation along six regions using polymerase chain reaction–restriction fragment‐length polymorphisms (PCR–RFLPs) (trnH–trnK, trnK–trnK, trnC–trnD, psbC–trnS, psaA–trnS, trnL–trnF) and further sequencing of trnL–trnF and trnH–psbA. In addition, 11 samples of other European species of Pulsatilla were sequenced to survey the genus‐scale cpDNA variation. Results Eleven PCR–RFLP polymorphisms were detected in P. vernalis, revealing seven haplotypes. They formed two distinct genetic groups. Three haplotypes representing both groups dominated and were widely distributed across Europe, whereas the others were restricted to localized regions (central Alps, Tatras/Sudetes mountains) or single populations. Sequencing analysis confirmed the reliability of PCR–RFLPs and homology of haplotypes across their distribution. The chloroplast DNA variation across the section Pulsatilla was low, but P. vernalis did not share haplotypes with other species. Main conclusions The genetic distinctiveness of P. vernalis populations from the south‐western Alps with respect to other Alpine populations, as well as the affinities between the former populations and those from the eastern Pyrenees, is demonstrated, thus providing support for the conclusions of previous studies. Glacial refugia in the Dolomites are also suggested. Isolation is inferred for the high‐mountain populations from the Tatras and Sudetes; this is in contrast to the case for the Balkans, which harboured the common haplotype. Specific microsatellite variation indicates the occurrence of periglacial lowland refugia north of the Alps, acting as a source for the post‐glacial colonization of Scandinavia. The presence of different fixed haplotypes in eastern and western Scandinavia, however, suggests independent post‐glacial colonization of these two areas, with possible founder effects.     

Genetic divergence in nuclear genomes between populations of <Emphasis Type="Italic">Fagus crenata</Emphasis> along the Japan Sea and Pacific sides of Japan

Genetic diversity and structure in Fagus crenata were studied by analyzing 14 nuclear microsatellite loci in 23 populations distributed throughout the species’ range. Although population differentiation was very low (F _ST = 0.027; R _ST = 0.041), both neighbor-joining tree and Bayesian clustering analyses provided clear evidence of genetic divergence between populations along the Japan Sea (Japan Sea lineage) and Pacific (Pacific lineage) sides of Japan, indicating that physical barriers to migration and gene flow, notably the mountain ranges separating the populations along the Japan Sea and Pacific sides, have promoted genetic divergence between these populations. The two lineages of the nuclear genome are generally consistent with those of the chloroplast genome detected in a previous study, with several discrepancies between the two genomes. Within-population genetic diversity was generally very high (average H _E = 0.839), but decreased in a clinal fashion from southwest to northeast, largely among populations of the Japan Sea lineage. This geographical gradient may have resulted from the late-glacial and postglacial recolonization to the northeast, which led to a loss of within-population genetic diversity due to cumulative founder effects.     

Phylogeographical structure revealed by chloroplast DNA variation in Japanese beech (Fagus crenata Blume)

Intraspecific genetic variation in three non-coding chloroplast DNA (cpDNA) regions (trnT-L and trnL-F spacers, and trnL intron) of Japanese beech (Fagus crenata Blume) was investigated. This species is a major constituent of the typical cool-temperate deciduous forests in Japan. Twenty-one F. crenata populations from throughout Japan, and four F. japonica populations, a close relative of F. crenata, were examined. Seven haplotypes were distinguishable in F. crenata based on nucleotide substitutions and indels. Pairwise nucleotide diversities among haplotypes ranged from 0.0000 to 0.0042 for F. crenata, including F. japonica. The geographical distribution of cpDNA haplotypes was found to be highly structured in F. crenata. Four haplotypes predominated: haplotypes FC1 and FC4 are prevalent on the Pacific Ocean coast, haplotype FC6 is prevalent on the Japan sea coast from the San-in district to Hokkaido, whilst haplotype FC3 is restricted to northern Kyushu and the western-most part of Honshu. Two haplotypes (FC5 and FC7) are restricted to single populations and one haplotype (FC2) is a derivative of FC1. Each of these haplotypes, except FC2, are thought to be derived from different glacial refugia. Phylogenetic analysis showed that neither F. crenata nor F. japonica was monophyletic for the haplotypes, suggesting either ancestral polymorphism or ancient introgression between the lineages of these two Fagus species.     

Probability distributions,vulnerability and sensitivity in Fagus crenata forests following predicted climate changes in Japan

Question: How much is the probability distribution of Fagus crenata forests predicted to change under a climate change scenario by the 2090s, and what are the potential impacts on these forests? What are the main factors inducing such changes? Location: The major islands of Japan. Methods: A predictive distribution model was developed with four climatic factors (summer precipitation, PRS; winter precipitation, PRW; minimum temperature of the coldest month, TMC; and warmth index, WI) and five non‐climatic factors (topography, surface geology, soil, slope aspect and inclination). A climate change scenario was applied to the model. Results: Areas with high probability (> 0.5) were predicted to decrease by 91%, retreating from the southwest, shrinking in central regions, and expanding northeastwards beyond their current northern limits. A vulnerability index (the reciprocal of the predicted probability) suggests that Kyushu, Shikoku, the Pacific Ocean side of Honshu and southwest Hokkaido will have high numbers of many vulnerable F. crenata forests. The forests with high negative sensitivity indices (the difference between simulated probabilities of occurrence under current and predicted climates) mainly occur in southwest Hokkaido and the Sea of Japan side of northern Honshu. Conclusion: F. crenata forest distributions may retreat from some islands due to a high WI. The predicted northeastward shift in northern Hokkaido is associated with increased TMC and PRS. High vulnerability and negative sensitivity of the forests in southern Hokkaido are due to increased WI.     

Extensive population expansion of Pedicularis longiflora (Orobanchaceae) on the Qinghai‐Tibetan Plateau and its correlation with the Quaternary climate change

The Qinghai‐Tibetan Plateau (QTP) is thought to be more strongly affected by the Quaternary glaciations than most other regions of the same latitude. It would be of great interest to investigate the population genetic structure of organisms distributed on the platform and its correlation with the Quaternary climatic oscillations. Here we used the chloroplast (cp)DNA trnT‐trnF sequence to study genetic variation and phylogeography of Pedicularis longiflora, an alpine herb with extensive distribution on the QTP. Based on a range‐wide sampling comprising 41 populations and 910 individuals, we detected 30 cpDNA haplotypes that were divided into five clades by phylogenetic and network analyses and a strong phylogeographical structure. All haplotypes but one in the three basal clades occur exclusively in the southeast QTP, whereas haplotypes in the young clade V occupy almost the whole species range. In particular, the young haplotype H18 occurs in 420 individuals, even at a frequency of 100% in some QTP platform populations and the Altai population. The haplotype distribution pattern, together with molecular clock estimation and mismatch distribution analysis, suggests that the southeast QTP was either a refuge for P. longiflora during the Quaternary climatic change or is the place of origin of the species. The present wide distribution of the species on the QTP platform has resulted from recent population expansions which could be dated back to 120 000–17 000 years ago, a period mostly before the last glacial maximum. The possible relationships among geographic genetic structure, climatic change and species diversification in Pedicularis are also discussed.     

Offshore Dolly Varden charr (<Emphasis Type="Italic">Salvelinus malma</Emphasis>) in the North Pacific

Many studies have investigated the ecology of charrs in freshwater, however, little is known about charrs in the ocean. This study examined the distribution, seasonal abundance, and some biological features of Dolly Varden (Salvelinus malma) in the Pacific Ocean. An analysis of by-catch data of Japanese offshore salmon monitoring showed that Dolly Varden were distributed across a wide range in the offshore waters of the Pacific Ocean, including the Japan Sea, Bering Sea, and Okhotsk Sea. The catch per unit effort showed a sharp increase from May to August, followed by a sharp decrease in September. Offshore areas served as an important summer habitat for anadromous Dolly Varden.     

Extensive length variation in the cpDNA trnT-trnF region of hemiparasitic Pedicularis and its phylogenetic implications

The cpDNA trnT-trnF region, a molecular marker widely used in the phylogenetic reconstruction at lower taxonomic levels, is relatively conserved in size and structure. In this region single length variation over 100 bp is much less common than small deletion for congeneric species of angiosperms. Here we examined evolutionary patterns of the trnT-trnF region in 43 species of Pedicularis, a species-rich genus with adaptive radiation. Four independent large deletions, varying from 203 to 297 bp in length, were detected from nine species of the genus, which might result from slipped-strand mispairing. These deletions occurred in different locations of the cpDNA region and in different clades of the phylogenetic tree, indicating that the deletion of large cpDNA fragments may be very frequent in the hemiparasitic lineage of the family Orobanchaceae. Parsimony analyses showed that section Cyathophora of Pedicularis, endemic to the Sino-Himalayan region, was a strongly supported monophyletic group. This section could have a recent origin followed by rapid radiation, considering that it is characterized by a large deletion in the trnT-trnF region and a relatively low interspecific sequence divergence.     

Intraspecific sequence variation of chloroplast DNA inPedicularis chamissonis Steven (Scrophulariaceae) and geographic structuring of the Japanese “Alpine” plants

In order to clarify evolutionary patterns and processes of intraspecific diversification ofPedicularis chamissonis Steven, we analyzed intraspecific variation of the nucleotide sequences of non-coding regions of chloroplast DNA: the intergenic spacers betweentrnT (UGU) andtrnL (UAA) 5′exon,trnL (UAA) 3′exon andtrnF (GAA), andatpB andrbcL. In 24 populations ofP. chamissonis, 33 nucleotide substitutions and 12 insertions/deletions were inferred, and their genetic distances ranged from 0.001 to 0.014. Seventeen distinct cpDNA haplotypes could be recognized and each haplotype was found to be geographically structured. Two major clades (the Northern and Southern clades) were revealed in phylogenetic analyses of cpDNA haplotypes. The haplotypes of the Northern clade had a wider distribution area in the populations of Mts. lide of central Honshu in Japan, northward to Unalaska Island in the Aleutians. Relationships among most haplotypes were unresolved polytomies. On the other hand, the haplotypes of the Southern clade occurred from the populations of Mt. Gassan southwards to Mt. Arakawa of central Honshu. Within this clade, three subclades were clearly recognized. From these results, we concluded that the haplotypes of the Northern and Southern clades inP. chamissonis might have traveled down to Japanese Archipelago from the north in not a single glacial period.     

Long-term occurrence of hybrids between Japan Sea and Pacific Ocean forms of threespine stickleback, <Emphasis Type="Italic">Gasterosteus aculeatus</Emphasis>, in Hokkaido Island,Japan

The distribution patterns of two genetically divergent forms (Japan Sea and Pacific Ocean forms) of Gasterosteus aculeatus and their hybrids in the Hokkaido Island, Japan were investigated. Hybrid frequencies and the mating pattern of natural F1-hybrids and backcrosses, were examined by using allozyme analysis. The distribution patterns matched those previously reported. All three sympatric localities in eastern Hokkaido Island included hybrids between the two forms in each year examined. A low frequency of hybrids of sticklebacks has been maintained in areas of sympatry of the two forms in Hokkaido Island for several years. The proportion of F1-hybrids was significantly lower than expected by random mating in six samples from four sympatric populations, suggesting form-assortative mating. Mating crosses of natural F1-hybrids existed in both patterns. Natural F1-hybrids backcrossed more frequently with Pacific Ocean form than Japan Sea form. However, the proportions of backcross individuals in two of three samples were not significantly different from the expected proportions by random mating between F1-hybrids and their parent forms. In other samples, despite the Pacific Ocean form predominant in a population, F1-hybrid predominantly backcrossed with the Japan Sea form. These results support the hypothesis of gene flow from Pacific Ocean to Japan Sea forms.     

Intraspecific diversity and distribution of the cosmopolitan species Pseudo‐nitzschia pungens (Bacillariophyceae): morphology,genetics, and ecophysiology of the three clades

Three clades of Pseudo‐nitzschia pungens, determined by the internal transcribed space (ITS) region, are distributed throughout the world. We studied 15 P. pungens clones from various geographical locations and confirmed the existence of the three clades within P. pungens, based on ITS sequencing and described the three subgroups (IIIaa, IIIab, and IIIb) of clade III. Clade III (clade IIIaa) populations were reported for the first time in Korean coastal waters and the East China Sea. In morphometric analysis, we found the ultrastructural differences in the number of fibulae, striae, and poroids that separate the three clades. We carried out physiological tests on nine clones belonging to the three clades growing under various culture conditions. In temperature tests, only clade III clones could not grow at lower temperatures (10°C and 15°C), although clade I and II clones grew well. The estimated optimal growth range of clade I clones was wider than that of clades II and III. Clade II clones were considered to be adapted to lower temperatures and clade III to higher temperatures. In salinity tests, clade II and III clones did not grow well at a salinity of 40. Clade I clones were regarded as euryhaline and clade II and III clones were stenohaline. This supports the hypothesis that P. pungens clades have different ecophysiological characteristics based on their habitats. Our data show that physiological and morphological features are correlated with genetic intraspecific differentiation in P. pungens.     

Distribution of chloroplast DNA haplotypes in the contact zone of Fagus crenata in the southwest of Kanto District, Japan

We have investigated cpDNA haplotype distribution in 24 populations of Fagus crenata in the southwest of Kanto District, Japan, and clarified the extent of intermixing of haplotypes in the contact zone by additional fine-scale analysis of two areas. Two cpDNA haplotypes belonging to different lineages were detected, and their distribution had geographical structure. Intermixed populations with the two haplotypes were limited to a narrow area. The geographical boundary between the haplotypes extended from Hakone to the west of the Kanto Mountains through the northern foot of Mt Fuji. No relationship was observed between the boundary location and the current topography of the southwest of Kanto District.