Phylogeography of Bivalve Cyclina sinensis: Testing the Historical Glaciations and Changjiang River Outflow Hypotheses in Northwestern Pacific

Background

The marginal seas of northwestern Pacific are characterized by unique topography and intricate hydrology. Two hypotheses have been proposed to explain genetic patterns of marine species inhabiting the region: the historical glaciations hypothesis suggests population genetic divergence between sea basins, whereas the Changjiang River outflow hypothesis suggests genetic break in line with the Changjiang Estuary. Here the phylogeography of bivalve Cyclina sinensis was investigated to test the validity of these two hypotheses for intertidal species in three marginal seas—the East China Sea (ECS), the South China Sea (SCS), and the Japan Sea (JPS).

Methodology/Principal Findings

Fragments of two markers (mitochondrial COI and nuclear ITS-1) were sequenced for 335 individuals collected from 21 populations. Significant pairwise Φ_ST were observed between different marginal sea populations. Network analyses illustrated restricted distribution of haplogroups/sub-haplogroups to sea basins, with a narrow secondary contact zone between the ECS and SCS. Demographic expansion was inferred for ECS and SCS lineages using mismatch distributions, neutral tests, and extended Bayesian Skyline Plots. Based on a molecular clock method, the divergence times among COI lineages were estimated dating from the Pleistocene.

Conclusions

The phylogeographical break revealed for C. sinensis populations is congruent with the historical isolation of sea basins rather than the putative Changjiang River outflow barrier. The large land bridges extending between seas during glaciation allowed accumulation of mutations and subsequently gave rise to deep divergent lineages. The low-dispersal capacity of the clam and coastal oceanography may facilitate the maintenance of the historical patterns as barriers shift. Our study supports the historical glaciations hypothesis for interpreting present-day phylogeographical patterns of C. sinensis, and highlights the importance of historical glaciations for generating genetic structure of marine coastal species especially those with low-dispersal abilities in northwestern Pacific.     

Genetic structure and historical demography of Collichthys lucidus inferred from mtDNA sequence analysis

Fluctuations of global climate changes during the Pleistocene had a huge impact on the marginal seas of the Northwestern Pacific. To examine historical demography and the population genetic structure of Collichthys lucidus, a total of 151 individuals from 7 locations were sequenced at the 5′ end of mitochondrial DNA control region and 380-bp fragments were obtained. The results of AMOVA showed that the genetic variation among the two groups was 78.39 % (P?=?0.78) and among populations within groups was 0.43 % (P?=?0.02). Both mismatch distribution analysis and neutrality tests showed C. lucidus has experienced a recent population expansion. Three distinct clades suggested that C. lucidus might be isolated in the three marginal seas during the Pleistocene periods. No genetic divergence within groups should be explained by insufficient time to attain migration-drift equilibrium. The changes in sea level during the Pleistocene had a major effect on the C. lucidus phylogeographical pattern because it limited its dispersal and at times created isolated populations.     

Comparative phylogeography in marginal seas of the northwestern Pacific

The maturation of marine phylogeography depends on integration of comparative information across different regions globally. The northwestern Pacific, characterized by unique tectonic setting, however, is still underrepresented. This study seeks to highlight its phylogeographical history based on the available population data, focusing on three seas: the East China Sea (ECS), the South China Sea (SCS) and the Sea of Japan (SOJ). We first conducted a literature survey to evaluate current research efforts and then reanalysed the population structure, historical demography and genealogy for two selections of studies (namely ‘the ECS category’ and ‘the multiple‐sea category’) to elucidate the evolutionary processes within and across the seas, respectively. For the ECS category, the meta‐analyses revealed most studies displayed a shallow phylogeny, indicating a single origin from the sea. Significant population structure was commonplace, particularly in molluck and crustacean studies, with proportions of 89% and 80%, respectively. Nearly all studies selected showed signals of population expansion: the times estimated were closely linked to a period of ~120–140 Kya rather than the last glacial maximum. For the latter category, divergent intraspecific lineages appeared among seas and overlapped in the adjacent regions, a pattern implying each sea had served as an independent refugium during glaciations. The genetic splits, however, were estimated to arise from separate events dating from late Miocene to middle Pleistocene. As phylogeography is still in its infancy in the region, more effort is needed to test and complement the general rules abstracted here. Finally, challenges and prospects were discussed to accelerate further research.     

Differentiation of golden-ringed dragonfly Anotogaster sieboldii (Selys, 1854) (Cordulegastridae: Odonata) in the insular East Asia revealed by the mitochondrial gene genealogy with taxonomic implications

Molecular phylogeographical analyses of Anotogaster sieboldii (Selys, 1854) were conducted to reveal the differentiation process of insular populations. The gene genealogy based on 845 bp of the mitochondrial genes (cytochrome oxidase subunit I and subunit II) indicated that A. sieboldii includes two deeply separated lineages. These two major lineages seem to have differentiated in Miocene before the formation of the insular East Asia. One lineage includes three inner clades that correspond to the populations of northern area (the Japanese main islands, Korean Peninsula, Yakushima), Amamioshima and Okinawajima. Populations of Central Ryukyu, including Amamioshima and Okinawajima, might have been divided from the northern populations in early Pleistocene. The other major lineage includes populations of the Yaeyama Group, Taiwan and East China. The former two populations were reconstructed as a reciprocal monophyletic group. Populations of Taiwan and Yaeyama Groups would have been separated from the continental ones in Pleistocene. These two highly divergent lineages should be recognized as distinct species. Furthermore, the mitochondrial lineages revealed six genetically distinct and geographically isolated assemblages: (1) northern populations, (2) Amamioshima, (3) Okinawajima, (4) Yaeyama Group, (5) Taiwan and (6) East China.     

Genome scan of the mitten crab Eriocheir sensu stricto in East Asia: population differentiation, hybridization and adaptive speciation

We examine the genetic structure and evolutionary history of the mitten crab Eriocheir sensu stricto in East Asia by employing a genome scan - amplified fragment length polymorphism. Population analysis reveals three divergent clades in Eriocheir s. s., which dominate the East China Sea-Yellow Sea, the Sea of Japan (plus Okinawa) and the South China Sea, respectively, mostly in agreement with our previous mtDNA analysis. With the tropical South China Sea inferred as the origin, the East China Sea-Yellow Sea and the Sea of Japan clades in the north diverged successively from the ancestral clade during the mid-Pleistocene. The divergence of the three clades likely resulted from isolation of the three marginal seas caused by sea level change in the Pleistocene. Two sympatric zones, one of the East China Sea-Yellow Sea and the South China Sea clades in southeast China and the other of the East China Sea-Yellow Sea and the Sea of Japan clades in Vladivostok, are demonstrated to be hybrid zones, with hybridization occurring currently in the former but historically in the latter. Adaptive speciation is observed in the divergence process of the three clades, possibly because of selection from accumulated temperature. Our study indicates that the genetic structure and evolutionary history of Eriocheir s. s. have been primarily affected by Pleistocene glacial cycles, secondarily by divergent selection and drainage isolation, but only minimally by human activities.     

Phylogeography and historical demography of the anadromous fish Leucopsarion petersii in relation to geological history and oceanography around the Japanese Archipelago

Phylogeographical patterns of marine and diadromous organisms are often influenced by dynamic ocean histories. For example, the marine realm around the Japanese Archipelago is an interesting area for phylogeographical research because of the wide variation in the environments driven by repeated shifts in sea level in the Quaternary. We analysed mitochondrial cyt b gene and nuclear myh6 gene sequences for individuals collected from throughout the range of the anadromous fish Leucopsarion petersii to assess the lineage divergence, phylogeographical pattern and historical demography in relation to geological history and oceanographic features around the archipelago. Leucopsarion petersii has two major lineages (the Japan Sea and Pacific Ocean lineages), which diverged during the late-early to middle Pleistocene. Geographical distributions of the two lineages were closely related to the pathways of the two warm currents, the Tsushima Current and the Kuroshio Current, that flow past the archipelago. Evidence of introgressive hybridization between these lineages was found at two secondary contact zones. Demographic tests suggested that the Japan Sea and Pacific Ocean lineages carried the genetic signal of different historical demographic processes, and these signals are probably associated with differences in habitat stability during recent glacial periods. The Japan Sea lineage has a larger body-size and more vertebrae, probably in relation to severe habitat conditions through Pleistocene climatic oscillations. Thus, the two lineages have long independent evolutionary histories, and the phylogeographical structure and demography of this species have been influenced both by historical events and the present-day oceanography around the Japanese Archipelago.     

Phylogeography of the mottled spinefoot Siganus fuscescens: Pleistocene divergence and limited genetic connectivity across the Philippine archipelago

Historical isolation during Pleistocene low sea level periods is thought to have contributed to divergence among marine basin populations across the Coral Triangle. In the Philippine archipelago, populations in the South China Sea, Sulu Sea–inland seas, and Philippine Sea‐Celebes Sea basins might have been partially isolated. Meanwhile, present‐day broadscale oceanographic circulation patterns suggest connectivity between these basins. To evaluate hypotheses regarding the influence of historical and contemporary factors on genetic structure, phylogeographic patterns based on mitochondrial control region sequences for a reef‐associated fish, Siganus fuscescens, were analysed. Three distinct lineages were recovered. One lineage was identified as the morphologically similar species Siganus canaliculatus, while two lineages are monophyletic with S. fuscescens. Clade divergence and demographic expansion in S. fuscescens occurred during the Pleistocene. A strong signal of latitudinal structure was detected (Φ_CT = 0.188), driven by marked differences in clade distribution: one clade is widely distributed (clade A), while a second clade (clade B) has a restricted northern distribution. Regional structure of clade A is consistent with the basin isolation hypothesis (Φ_CT = 0.040) and suggests isolation of the South China Sea (Φ_CT = 0.091). Fine‐scale structure was observed in the South China Sea and south Philippine Sea, while Sulu Sea and inland seas were unstructured. Genetic structure across multiple spatial scales (archipelagic, regional, and fine‐scale within basins) suggests the influence of vicariant barriers and contemporary limits to gene flow in S. fuscescens that may be influenced by oceanographic circulation, geographical distance between available habitats, and latitudinal temperature differences.     

Multiple ice‐age refugia in Pacific cod,Gadus macrocephalus

Pleistocene ice‐ages greatly influenced the historical abundances of Pacific cod, Gadus macrocephalus, in the North Pacific and its marginal seas. We surveyed genetic variation at 11 microsatellite loci and mitochondrial (mt) DNA in samples from twelve locations from the Sea of Japan to Washington State. Both microsatellite (mean H = 0.868) and mtDNA haplotype (mean h = 0.958) diversities were large and did not show any geographical trends. Genetic differentiation between samples was significantly correlated with geographical distance between samples for both microsatellites (F_ST = 0.028, r² = 0.33) and mtDNA (F_ST = 0.027, r² = 0.18). Both marker classes showed a strong genetic discontinuity between northwestern and northeastern Pacific populations that likely represents groups previously isolated during glaciations that are now in secondary contact. Significant differences appeared between samples from the Sea of Japan and Okhotsk Sea that may reflect ice‐age isolations in the northwest Pacific. In the northeast Pacific, a microsatellite and mtDNA partition was detected between coastal and Georgia Basin populations. The presence of two major coastal mtDNA lineages on either side of the Pacific Ocean basin implies at least two ice‐age refugia and separate postglacial population expansions facilitated by different glacial histories. Northward expansions into the Gulf of Alaska were possible 14–15 kyr ago, but deglaciation and colonization of the Georgia Basin probably occurred somewhat later. Population expansions were evident in mtDNA mismatch distributions and in Bayesian skyline plots of the three major lineages, but the start of expansions appeared to pre‐date the last glacial maximum.     

Land masses and oceanic currents drive population structure of Heritiera littoralis,a widespread mangrove in the Indo‐West Pacific

Phylogeographic forces driving evolution of sea‐dispersed plants are often influenced by regional and species characteristics, although not yet deciphered at a large spatial scale for many taxa like the mangrove species Heritiera littoralis. This study aimed to assess geographic distribution of genetic variation of this widespread mangrove in the Indo‐West Pacific region and identify the phylogeographic factors influencing its present‐day distribution. Analysis of five chloroplast DNA fragments’ sequences from 37 populations revealed low genetic diversity at the population level and strong genetic structure of H. littoralis in this region. The estimated divergence times between the major genetic lineages indicated that glacial level changes during the Pleistocene epoch induced strong genetic differentiation across the Indian and Pacific Oceans. In comparison to the strong genetic break imposed by the Sunda Shelf toward splitting the lineages of the Indian and Pacific Oceans, the genetic differentiation between Indo‐Malesia and Australasia was not so prominent. Long‐distance dispersal ability of H. littoralis propagules helped the species to attain transoceanic distribution not only across South East Asia and Australia, but also across the Indian Ocean to East Africa. However, oceanic circulation pattern in the South China Sea was found to act as a barrier creating further intraoceanic genetic differentiation. Overall, phylogeographic analysis in this study revealed that glacial vicariance had profound influence on population differentiation in H. littoralis and caused low genetic diversity except for the refugia populations near the equator which might have persisted through glacial maxima. With increasing loss of suitable habitats due to anthropogenic activities, these findings therefore emphasize the urgent need for conservation actions for all populations throughout the distribution range of H. littoralis.     

Phylogeographic heterogeneity of the brown macroalga Sargassum horneri (Fucaceae) in the northwestern Pacific in relation to late Pleistocene glaciation and tectonic configurations

Pleistocene glacial oscillations and associated tectonic processes are believed to have influenced the historical abundances and distribution of organisms in the Asia Northwest Pacific (ANP). Accumulating evidence indicates that factors shaping tempospatial population dynamics and distribution patterns of marine taxa vary with biogeographical latitude, pelagic behaviour and oceanographic regimes. To detect what kinds of historical and contemporary factors affected genetic connectivity, phylogeographic profiles of littoral macroalga Sargassum horneri in the ANP were analysed based on mitochondrial (Cox3) and chloroplast (rbcL) data sets. Five distinct clades were recovered. A strong signature of biogeographical structure was revealed (Φ(CT) = 0.487, P < 0.0001) derived from remarkable differentiation in clade distribution, as clade I is restricted to Chinese marginal seas (Yellow-Bohai Sea, East China Sea and South China Sea), whereas clades II-V are discontinuously scattered around the main Islands of Japan. Furthermore, two secondary contact regions were identified along the south Japan-Pacific coastline. This significant differentiation between the two basins may reflect historical glacial isolation in the northwestern Pacific, which is congruent with the estimates of clade divergence and demographic expansion during the late Quaternary low sea levels. Analysis of molecular variance and the population-pair statistic F(ST) also revealed significant genetic structural differences between Chinese marginal seas and the Japanese basin. This exceptional phylogeographic architecture in S. horneri, initially shaped by historical geographic isolation during the late Pleistocene ice age and physical biogeographical barriers, can be complicated by oceanographic regimes (ocean surface currents) and relocating behaviour such as oceanic drifting.     

Geographical distribution,dispersal and genetic divergence of the mantis shrimp Oratosquilla oratoria (Stomatopoda: Squillidae) in China Sea

The period of Cenozoic Himalayan orogeny was the developmental stage of the marginal seas and islands adjacent to the western Pacific. In the study, the molecular divergence dating between the northern and southern groups by fossil calibration was approximately 10.9 Myr, which was earlier than the time of the formation of Taiwan Island. Statistical-Dispersal Vicariance Analysis (S-DIVA) suggested that mantis shrimp of the China seas originated from two different ancestral regions (i.e., the Yellow Sea and South China Sea). However, the fluctuation of the Taiwan Strait in the late glacial period further influenced the phylogeographic structure of the mantis shrimp. In addition, the opposite ocean currents of the northern and southern seas of China, driving the planktonic larvae of marine species in opposite directions in the reproductive season, further accelerated their genetic divergence. In contrast, there were weak differences among the three northern seas due to their sharing one common ancestral region and then facing the dispersal event in the Cenozoic era and later sudden population expansion in the late Quaternary interglacial (54–75 ka). In conclusion, geological and climatic changes in past geological periods have greatly influenced the geographical distribution, dispersal and genetic divergence of the mantis shrimp.     

Phylogeography study of Ammodytes personatus in Northwestern Pacific: Pleistocene isolation, temperature and current conducted secondary contact

To assess the role of historical process and contemporary factors in shaping population structures in Northwestern Pacific, mitochondrial control region sequences were analyzed to characterize the phylogeography and population structure of the Japanese sand lance Ammodytes personatus. A total of 429 individuals sampled from 17 populations through the species' range are sequenced. Two distinct lineages are detected, which might have been divergent in the Sea of Japan and Pacific costal waters of Japanese Island, during the low sea level. Significant genetic structure is revealed between the Kuroshio and Oyashio Currents. However, significant genetic structure is also detected in the Sea of Japan, contracting expected homogenization hypothesis in Tsushima Current. The haplotype frequency of lineages in both sides of Japanese Island and significant genetic structure between north and south groups revealed that the distribution of lineage B and north group were highly limited by the annual sea temperature. The lack of lineage B in Qingdao population with low sea temperature reflects the sea temperature barrier. Lack of genetic structure in the south group and north group populations indicated that ocean currents within groups facilitated the dispersal of A. personatus.     

Migration and isolation during the turbulent Ponto‐Caspian Pleistocene create high diversity in the crustacean Paramysis lacustris

The Ponto‐Caspian brackish‐water fauna inhabits estuaries and rivers of the Black, Azov and Caspian seas and is fragmented by higher salinity waters and a major interbasin watershed. The fauna is known for the high levels of endemism, complex zoogeographic histories, and as a recent source of successful invasive species. It remains debated whether the Black and Azov Sea brackish‐water populations survived unfavourable Pleistocene conditions in multiple separate refugia or whether the two seas were (repeatedly) recolonized from the Caspian. Using microsatellite and mtDNA markers, we demonstrate deep among‐ and within‐basin subdivisions in a widespread Ponto‐Caspian mysid crustacean Paramysis lacustris. Five genetic clusters were identified, but their relationships did not reflect the geography of the region. The Azov cluster was the most distinct (4–5% COI divergence), despite its geographic position in the corridor between Black and Caspian seas, and may represent a new species. In the northern Black Sea area, the Dnieper cluster was closer to the Caspian cluster than to the neighbouring Danube–Dniester–Bug populations, suggesting separate colonizations of the Black Sea. Overall, the data implied a predominant gene flow from the east to the Black Sea and highlight the importance of Caspian Sea transgressions in facilitating dispersal. Yet, the presence of distinct lineages in the Black Sea points to the persistence of isolated refugial populations that have gained diagnostic differences under presumably high mutation rates and large population sizes. The unfavourable Pleistocene periods in the Black Sea therefore appear to have promoted diversification of the brackish‐water lineages, rather than extirpated them.     

Phylogeography of the flathead mullet Mugil cephalus in the north-west Pacific as inferred from the mtDNA control region

The population genetic structure and historical demography of the flathead mullet Mugil cephalus were investigated using the mtDNA control region (CR) sequences (909–1015 bp) of 126 individuals collected from seven locations in the north-west Pacific between 2005 and 2007. Haplotype diversity ( h = 0·9333–1·000) and nucleotide diversity (π = 0·0046–0·1467) varied greatly among the sampling locations. Phylogenetic analysis of the CR sequences indicated that M. cephalus in the north-west Pacific belongs to two highly divergent lineages (lineages 1 and 2), with the inferred population structure being closely associated with the distribution of both lineages. Two populations were identified, one from the East China Sea and the other from the South China Sea. The former samples were obtained from Taiwan and Qingdao of north China and associated with lineage 1 haplotypes. The latter samples were collected from the Philippines, Pearl River of South China and two samples from Japan, all of which were associated with lineage 2. Japanese samples from Okinawa and Yokosuka had different degrees of mixing between lineages 1 and 2. Historical demographic variables in both populations indicated that Pleistocene glaciations had a strong impact on M. cephalus in the north-west Pacific, resulting in a recent demographic decline of the East China Sea population but in demographic equilibrium for the South China Sea population. Japan appears to be a contact zone between lineages 1 and 2, but it may also be indicative of coexistence between resident and migratory populations. Further global studies are required to clarify the taxonomic status of this cosmopolitan species.     

Phylogeography of Quercus variabilis Based on Chloroplast DNA Sequence in East Asia: Multiple Glacial Refugia and Mainland-Migrated Island Populations

The biogeographical relationships between far-separated populations, in particular, those in the mainland and islands, remain unclear for widespread species in eastern Asia where the current distribution of plants was greatly influenced by the Quaternary climate. Deciduous Oriental oak (Quercus variabilis) is one of the most widely distributed species in eastern Asia. In this study, leaf material of 528 Q. variabilis trees from 50 populations across the whole distribution (Mainland China, Korea Peninsular as well as Japan, Zhoushan and Taiwan Islands) was collected, and three cpDNA intergenic spacer fragments were sequenced using universal primers. A total of 26 haplotypes were detected, and it showed a weak phylogeographical structure in eastern Asia populations at species level, however, in the central-eastern region of Mainland China, the populations had more haplotypes than those in other regions, with a significant phylogeographical structure (N _ST =0.751> G _ST =0.690, P<0.05). Q. variabilis displayed high interpopulation and low intrapopulation genetic diversity across the distribution range. Both unimodal mismatch distribution and significant negative Fu’s F_S indicated a demographic expansion of Q. variabilis populations in East Asia. A fossil calibrated phylogenetic tree showed a rapid speciation during Pleistocene, with a population augment occurred in Middle Pleistocene. Both diversity patterns and ecological niche modelling indicated there could be multiple glacial refugia and possible bottleneck or founder effects occurred in the southern Japan. We dated major spatial expansion of Q. variabilis population in eastern Asia to the last glacial cycle(s), a period with sea-level fluctuations and land bridges in East China Sea as possible dispersal corridors. This study showed that geographical heterogeneity combined with climate and sea-level changes have shaped the genetic structure of this wide-ranging tree species in East Asia.     

Phylogeography of Ophiorrhiza japonica (Rubiaceae) in continental islands,the Ryukyu Archipelago,Japan

Aim Phylogeographical patterns in the Ryukyu Archipelago have been explained primarily by landbridge formation and the opening of two straits in the Pliocene, namely the Tokara and Kerama gaps. These old straits have been considered to be the barriers most likely to determine genetic boundaries. To test this, we conducted a molecular analysis of the herb Ophiorrhiza japonica. We discuss the causes of and processes involved in its phylogeographical structure and explore aspects of island separation other than the duration of the straits to explain genetic boundaries at the gaps. Location Ryukyu Archipelago, Japan. Methods Plants were collected from 40 localities in the archipelago and vicinity. Non‐coding regions of chloroplast DNA were sequenced. The genealogical relationships among haplotypes were estimated using a statistical parsimony network. To examine the phylogeographical structure, we compared two parameters of population differentiation, namely G_ST and N_ST, and conducted correlation analysis of genetic and geographical distances. Genetic boundaries were identified using Monmonier’s maximum difference algorithm. To test vicariance–dispersal hypotheses, that is, vicariance after migration via the Pliocene landbridge or over‐sea dispersal in the Pleistocene, molecular dating analysis was conducted. Results A statistical parsimony network revealed that the haplotypes from the Ryukyu Archipelago and northwards coalesce to one ancestral haplotype in Taiwan. A clear phylogeographical structure was observed: plants within the same population and populations in geographical proximity were phylogenetically close. A genetic boundary was recognized across the Kerama Gap, but not across the Tokara Gap. Dating analysis suggested that population divergence across the Kerama Gap occurred in the early to late Pleistocene. Main conclusions The statistical parsimony network suggests migration from Taiwan and northward range expansion in the archipelago. Based on the divergence time, over‐sea dispersal in the Pleistocene is likely, although migration via a Pliocene landbridge is not totally rejected. Negligible genetic differentiation across the Tokara Gap suggests recent over‐sea dispersal, possibly facilitated by the small geographical width of the gap. Conversely, the large genetic differentiation across the Kerama Gap is probably explained by the large geographical distance across it. The past splitting of a landbridge would have had a significant influence on population differentiation after a certain geographical distance was reached.     

Nested clade and phylogeographic analyses of the chub, Leuciscus cephalus (Teleostei, cyprinidae), in Greece: implications for Balkan Peninsula biogeography

Phylogenetic relationships among Greek populations of the chub, Leuciscus cephalus, were investigated using 600 bp of the cytochrome b gene. The aim of this study was to test the assumption that the main difference in ichthyological composition between both sides of the Balkan Peninsula is directly linked to differences in the dispersion mechanisms used by fish in order to extend their distribution range. Phylogenetic and nested clade analyses clearly showed that populations in Greece are significantly differentiated. Greek populations were found to descend from three lineages in three geographical provinces: Western, Central, and Eastern Greece. The chub reached Western Greece at the beginning of the Pleistocene and Eastern Greece during the mid-Pleistocene. Chub dispersion occurred mainly by river confluence due to sea level lowering and river capture in Western Greece and sea dispersal with low-salinity conditions within the Aegean Sea in Eastern Greece. However, in Central Greece, the original mtDNA lineage has presumably been lost owing to a genetic introgression following a second invasion from the Danube during the final stage of the last glaciation. This study provides new elements for a better understanding of the composition of the contemporary ichthyofauna in Greece and highlights possible evolutionary mechanisms responsible for the high endemism rate in the Western Greek biogeographic province.     

Pleistocene isolation and recent gene flow in Haliotis asinina, an Indo-Pacific vetigastropod with limited dispersal capacity

Haliotis asinina is a broadcast-spawning mollusc that inhabits Indo-Pacific coral reefs. This tropical abalone develops through a nonfeeding larval stage that is competent to settle on specific species of coralline algae after 3-4 days in the plankton. Failure to contact an inductive algae within 10 days of hatching usually results in death. These life cycle characteristics suggest a limited capacity for dispersal and thus gene flow. This makes H. asinina particularly suitable for elucidating phylogeographical structure throughout the Indo-Malay Archipelagoes, and eastern Indian and western Pacific Oceans, all regions of biogeographical complexity and high conservation value. We assayed 482 bp of the mitochondrial cytochrome oxidase II gene in 206 abalone collected from 16 geographically discrete sites across the Indian and Pacific Oceans and Indo-Malay Archipelagoes. DNA sequence variation was analysed via population genetics and phylogenetics, and by nested clade analyses (NCA). Our data resolved clear phylogeographical breaks among major biogeographical regions, with sequence divergences ranging from a high of 3.7% and 3.0% between Indian and Pacific sites and Pacific and Indo-Malay sites, respectively, to a low of 1.1% between Indian and Indo-Malay sites. Despite the apparent limited dispersal capacity of H. asinina, no finer scale phylogeographical structure was resolved within the respective biogeographical regions. However, amova and NCA identified several significant associations between haplotypes and geographical distribution, most notably higher gene flow among geographical populations associated with major ocean currents. Our study provides further evidence that larval dispersal capacity alone is not a good predictor of population genetic structure in marine invertebrates. We infer instead that a combination of historical events (long-term barriers followed by range expansion associated with Pleistocene sea level changes) and contemporary processes (gene flow restricted by life history and oceanography) have shaped observed patterns of H. asinina phylogeography.     

Comparative phylogeography of the two pink salmon broodlines: an analysis based on a mitochondrial DNA genealogy

Over most of their natural northern Pacific Ocean range, pink salmon (Oncorhynchus gorbuscha) spawn in a habitat that was repeatedly and profoundly affected by Pleistocene glacial advances. A strictly two-year life cycle of pink salmon has resulted in two reproductively isolated broodlines, which spawn in alternating years and evolved as temporal replicates of the same species. To study the influence of historical events on phylogeographical and population genetic structure of the two broodlines, we first reconstructed a fine-scale mtDNA haplotype genealogy from a sample of 80 individuals and then determined the geographical distribution of the major genealogical assemblages for 718 individuals sampled from nine Alaskan and eastern Asian even- and nine odd-year pink salmon populations. Analysis of restriction site states in seven polymerase chain reaction (PCR)-amplified mtDNA regions (comprising 97% of the mitochondrial genome) using 13 endonucleases resolved 38 haplotypes, which clustered into five genealogical lineages that differed from 0.065 to 0.225% in net sequence divergence. The lineage sorting between broodlines was incomplete, which suggests a recent common ancestry. Within each lineage, haplotypes exhibited star-like genealogies indicating recent population growth. The depth of the haplotype genealogy is shallow ( approximately 0.5% of nucleotide sequence divergence) and probably reflects repeated decreases in population size due to Pleistocene glacial advances. Nested clade analysis (NCA) of geographical distances showed that the geographical distribution observed for mitochondrial DNA (mtDNA) haplotypes resulted from alternating influences of historical range expansions and episodes of restricted dispersal. Analyses of molecular variance showed weak geographical structuring of mtDNA variation, except for the strong subdivision between Asian and Alaskan populations within the even-year broodline. The genetic similarities observed among and within geographical regions probably originated from postglacial recolonizations from common sources rather than extensive gene flow. The phylogeographical and population genetic structures differ substantally between broodlines. This can be explained by stochastic lineage sorting in glacial refugia and perhaps different recolonization routes in even- and odd-year broodlines.     

Diversity and trans-arctic invasion history of mitochondrial lineages in the North Atlantic Macoma balthica complex (Bivalvia: Tellinidae)

The history of repeated inter- or transoceanic invasions in bivalve mollusks of the circumpolar Macoma balthica complex was assessed from mtDNA COIII sequences. The data suggest that four independent trans-Arctic invasions, from the Pacific, gave rise to the current lineage diversity in the North Atlantic. Unlike in many other prominent North Atlantic littoral taxa, no evidence for (postinvasion) trans-Atlantic connections was found in the M. balthica complex. The earliest branch of the mtDNA tree is represented by the temperate-boreal North American populations (=Macoma petalum), separated from the M. balthica complex proper in the Early Pliocene at latest. The ensuing trans-Arctic invasions established the North European M. b. rubra, which now prevails on the North Sea and northeast Atlantic coasts, about two million years ago, and the currently northwest Atlantic M. balthica lineage in the Canadian Maritimes, in the Middle Pleistocene. The final reinvasion(s) introduced a lineage that now prevails in a number of North European marginal seas and is still hardly distinguishable from North Pacific mtDNA (M. b. balthica). We used coalescence simulation analyses to assess the age of the latest invasion from the Pacific to the northeast Atlantic. The results refute the hypothesis of recent, human-mediated reintroductions between northeast Pacific and the North European marginal seas in historical times. Yet they also poorly fit the alternative hypotheses of an early postglacial trans-Arctic invasion (< 11 thousand years ago), or an invasion during the previous Eemian interglacial (120 thousand years ago). Divergence time estimates rather fall in the Middle Weichselian before the Last Glacial Maximum, in conflict with the conventional thinking of trans-Arctic biogeographical connections; an early Holocene reinvasion may still be regarded as the most plausible scenario. Today, the most recently invaded Pacific mtDNA lineage is found admixed with the earlier established European Atlantic "rubra" lineage in the Baltic Sea and in Barents Sea populations east of the Varanger peninsula, and it is practically exclusive in the White and Pechora seas. Yet mtDNA does not always constitute an unequivocal taxonomic marker at individual level; the marginal populations represent hybrid swarms of the Atlantic and Pacific lineages in their nuclear genes.