Molecular phylogeography and population structure of a mid-elevation montane frog Leptobrachium ailaonicum in a fragmented habitat of southwest China

Leptobrachium ailaonicum is a vulnerable anuran restricted to a patchy distribution associated with small mountain streams surrounded by forested slopes at mid-elevations (approximately 2000-2600 m) in the subtropical Mount Wuliang and Mount Ailao ranges in southwest China (Yunnan Province) and northern Vietnam. Given high habitat specificity and lack of suitable habitat in lower elevations between these ranges, we hypothesized limited gene flow between populations throughout its range. We used two mitochondrial genes to construct a phylogeographic pattern within this species in order to test our hypothesis. We also examined whether this phylogeographic pattern is a response to past geological events and/or climatic oscillations. A total of 1989 base pairs were obtained from 81 individuals of nine populations yielding 51 unique haplotypes. Both Bayesian and maximum parsimony phylogenetic analyses revealed four deeply divergent and reciprocally monophyletic mtDNA lineages that approximately correspond to four geographical regions separated by deep river valleys. These results suggest a long history of allopatric separation by vicariance. The distinct geographic distributions of four major clades and the estimated divergence time suggest spatial and temporal separations that coincide with climatic and paleogeographic changes following the orogeny and uplift of Mount Ailao during the late Miocene to mid Pliocene in southwest China. At the southern distribution, the presence of two sympatric yet differentiated clades in two areas are interpreted as a result of secondary contact between previously allopatric populations during cooler Pleistocene glacial cycles. Analysis of molecular variance indicates that most of the observed genetic variation occurs among the four regions implying long-term interruption of maternal gene flow, suggesting that L ailaonicum may represent more than one distinct species and should at least be separated into four management units corresponding to these four geographic lineages for conservation. (C) 2009 Elsevier Inc. All rights reserved.

Molecular phylogeography of endangered sharp-snouted pitviper (Deinagkistrodon acutus; Reptilia, Viperidae) in Mainland China

Using phylogenetic and population genetic approaches, the present study reports the phylogeographic structure of the sharp-snouted pitviper (Deinagkistrodon acutus), a threatened snake species with commercial and medicinal importance in China. The entire mitochondrial ND2 gene (NADH dehydrogenase subunit 2) sequences of 86 individuals of D. acutus from 14 localities across its range in China were determined. Based on the results of phylogenetic analyses, distribution of diagnostic sites, haplotype network, and AMOVA hierarchical analysis, an cast-west division of the whole D. acutus population could be observed. Geographically, a line formed by a lake, river, and mountain chain (the Poyang Lake, Gan River to the southern end of the Wuyi Mountains), results in vicariance and approximately vertically splits the range into two and the whole population into two main lineages (western and eastern). The bifurcating tree suggested generally west to east dispersal trend. The data fit the isolation by distance (IBD) model well. Star-like clusters in haplotype network, significantly negative values of Fs statistics, and unimodal mismatch distributions all suggest recent demographic expansions in four areas. The results show that isolation, dispersal, bottleneck, and expansion jointly constitute the history of D. acutus. In a haplotype network, the excessive predominance of central haplotypes, few medium-frequency haplotypes, predominance (73.1 %) of the singletons among the derived haplotypes, most of which are connected to the central haplotype by only one mutational step, unsymmetrical campanulate unimodal curve of mismatch distributions and leftwards shift of the peaks, all suggest that the whole D. acutus population is a young population with low genetic diversity. Based on the data, the first priority for conservation action should be given to the Huangshan unit. (c) 2007 Elsevier Inc. All rights reserved.

Comparative Cytogenetics and Molecular Phylogeography in the Group Astyanax altiparanae - Astyanax aff. bimaculatus (Teleostei, Characidae)

The genus Astyanax comprises small characin fish of the neotropical region. The so-called `yellow-tailed characins` compose one of the most widely distributed Astyanax groups. A. altiparanae and A. aff. bimaculatus, are evolutionarily closely related and commonly found in several Brazilian hydrographic basins. In the present work, chromosomal data of specimens of A. altiparanae and A. aff. bimaculatus from 4 hydrographic basins in the states of Sao Paulo (Upper Tiete, Paranapanema, Ribeira de Iguape) and Rio de Janeiro (Guapimirim) are shown. All the populations showed 50 chromosomes, with different karyotypic formula. Although only a single Ag-NOR bearing chromosome pair was observed, all populations possess multiple cistrons of 18S rDNA. FISH with the 5S rDNA probe showed single signals at the interstitial position of one metacentric chromosome pair. C-bands are distributed in the terminal and interstitial regions of several chromosomes. However, the As-51 satDNA are frugally located in a few chromosomes of fishes from Upper Tiete, Paranapanema and Guapimirim Rivers, being absent in individuals of A. aff. bimaculatus from Ribeira de Iguape River basin. Beside these 4 populations, molecular phylogeography studies were also performed in individuals from Middle and Lower Tiete River basin and from 2 additional collection sites in the Paranapanema and Ribeira de Iguape River basins. The phylogeographic analysis using 2 mtDNA regions (totalizing 1.314 bp of ND2 and ATPase6/8 genes) of 8 populations of the group of `yellow-tailed characins` from 3 major hydrographic basins showed structuring of populations, suggesting a correlation between chromosomal (nuclear) and molecular (mitochondrial) data. Copyright (C) 2011 S. Karger AG, Basel

Paleodistributions and Comparative Molecular Phylogeography of Leafcutter Ants (Atta spp.) Provide New Insight into the Origins of Amazonian Diversity

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Molecular phylogeography of tick-borne encephalitis virus in central Europe

In order to obtain a better understanding of tick-borne encephalitis virus (TBEV) strain movements in central Europe the E gene sequences of 102 TBEV strains collected from 1953 to 2011 at 38 sites in the Czech Republic, Slovakia, Austria and Germany were determined. Bayesian analysis suggests a 350-year history of evolution and spread in central Europe of two main lineages, A and B. In contrast to the east to west spread at the Eurasian continent level, local central European spreading patterns suggest historic west to east spread followed by more recent east to west spread. The phylogenetic and network analyses indicate TBEV ingressions from the Czech Republic and Slovakia into Germany via landscape features (Danube river system), biogenic factors (birds, red deer) and anthropogenic factors. The identification of endemic foci showing local genetic diversity is of paramount importance to the field as these will be a prerequisite for in-depth analysis of focal TBEV maintenance and long-distance TBEV spread.

Molecular phylogeography of the European coastal plants Crithmum maritimum L., Halimione portulacoides (L.) Aellen, Salsola kali L. and Calystegia soldanella (L.) R. Br.

The intraspecific phylogeography of four European coastal plants, Crithmum maritimum, Halimione portulacoides, Salsola kali and Calystegia soldanella, was inferred from AFLP and ITS data. Only in C. maritimum, H. portulacoides and S. kali, a spatial genetic structure was revealed. The phylogeographic similarities and dissimilarities of these species include: (1) All three have distinct Black/Aegean and Adriatic Sea clusters. (2) Salsola kali and H. portulacoides show a distinct Atlantic/North Sea/Baltic Sea cluster, while Atlantic and eastern Spanish material of C. maritimum clustered together. (3) In the west Mediterranean, only S. kali forms a single cluster, while both H. portulacoides and C. maritimum display a phylogeographic break in the vicinity of the southern French coast. For S. kali, AFLP and ITS data concur in identifying separate Atlantic, east and west Mediterranean clades. All these patterns are postulated to result from both temperature changes during the last glacial and contemporary sea currents. No geographic AFLP structure was revealed in C. soldanella, both at the range-wide and population level. This was attributed to the remarkable seed dispersal ability of this species and possibly its longevity and clonal growth, preserving a random pattern of genetic variation generated by long-distance seed dispersal over long time periods.

Molecular phylogeography of the Woodland Ringlet (Erebia medusa [Denis and Schiffermüller] 1775) in Europe

Phylogeography is a recent field of biological research that links phylogenetics to biogeography through deciphering the imprint that evolutionary history has left on the genetic structure of extant populations. During the cold phases of the successive ice ages, which drastically shaped species’ distributions since the Pliocene, populations of numerous species were isolated in refugia where many of them evolved into different genetic lineages. My dissertation deals with the phylogeography of the Woodland Ringlet (Erebia medusa [Denis and Schiffermüller] 1775) in Central and Eastern Europe. This Palaearctic butterfly species is currently distributed from central France and south eastern Belgium over large parts of Central Europe and southern Siberia to the Pacific. It is absent from those parts of Europe with mediterranean, oceanic and boreal climates. It was supposed to be a Siberian faunal element with a rather homogeneous population structure in Central Europe due to its postglacial expansion out of a single eastern refugium. An already existing evolutionary scenario for the Woodland Ringlet in Central and Eastern Europe is based on nuclear data (allozymes). To know if this is corroborated by organelle evolutionary history, I sequenced two mitochondrial markers (part of the cytochrome oxydase subunit one and the control region) for populations sampled over the same area. Phylogeography largely relies on the construction of networks of uniparentally inherited haplotypes that are compared to geographic haplotype distribution thanks to recent developed methods such as nested clade phylogeographic analysis (NCPA). Several ring-shaped ambiguities (loops) emerged from both haplotype networks in E. medusa. They can be attributed to recombination and homoplasy. Such loops usually avert the straightforward extraction of the phylogeographic signal contained in a gene tree. I developed several new approaches to extract phylogeographic information in the presence of loops, considering either homoplasy or recombination. This allowed me to deduce a consistent evolutionary history for the species from the mitochondrial data and also adds plausibility for the occurrence of recombination in E. medusa mitochondria. Despite the fact that the control region is assumed to have a lack of resolving power in other species, I found a considerable genetic variation of this marker in E. medusa which makes it a useful tool for phylogeographic studies. In combination with the allozyme data, the mitochondrial genome supports the following phylogeographic scenario for E. medusa in Europe: (i) a first vicariance, due to the onset of the Würm glaciation, led to the formation of several major lineages, and is mirrored in the NCPA by restricted gene flow, (ii) later on further vicariances led to the formation of two sub-lineages in the Western lineage and two sub-lineages in the Eastern lineage during the Last Glacial Maximum or Older Dryas; additionally the NCPA supports a restriction of gene flow with isolation by distance, (iii) finally, vicariance resulted in two secondary sub-lineages in the area of Germany and, maybe, to two other secondary sub-lineages in the Czech Republic. The last postglacial warming was accompanied by strong range expansions in most of the genetic lineages. The scenario expected for a presumably Siberian faunal element such as E. medusa is a continuous loss of genetic diversity during postglacial westward expansion. Hence, the pattern found in this thesis contradicts a typical Siberian origin of E. medusa. In contrast, it corroboratess the importance of multiple extra-Mediterranean refugia for European fauna as it was recently assumed for other continental species.

Phylogeography of an Atlantic forest passerine reveals demographic stability through the last glacial maximum

In this study we analyzed the phylogeographic pattern and historical demography of an endemic Atlantic forest (AF) bird, Basileuterus leucoblepharus, and test the influence of the last glacial maximum (LGM) on its population effective size using coalescent simulations. We address two main questions: (i) Does B. leucoblepharus present population genetic structure congruent with the patterns observed for other AF organisms? (ii) How did the LGM affect the effective population size of B. leucoblepharus? We sequenced 914 bp of the mitochondrial gene cytochrome b and 512 bp of the nuclear intron 5 of beta-fibrinogen of 62 individuals from 15 localities along the AF. Both molecular markers revealed no genetic structure in B. leucoblepharus. Neutrality tests based on both loci showed significant demographic expansion. The extended Bayesian skyline plot showed that the species seems to have experienced demographic expansion starting around 300,000 years ago, during the late Pleistocene. This date does not coincide with the LGM and the dynamics of population size showed stability during the LGM. To further test the effect of the LGM on this species, we simulated seven demographic scenarios to explore whether populations suffered specific bottlenecks. The scenarios most congruent with our data were population stability during the LGM with bottlenecks older than this period. This is the first example of an AF organism that does not show phylogeographic breaks caused by vicariant events associated to climate change and geotectonic activities in the Quaternary. Differential ecological, environmental tolerances and habitat requirements are possibly influencing the different evolutionary histories of these organisms. Our results show that the history of organism diversification in this megadiverse Neotropical forest is complex. Crown Copyright (c) 2012 Published by Elsevier Inc. All rights reserved.

菰的分子谱系地理研究及茭白的起源

近几十年来，栽培作物及其野生近缘（祖先）种的群体遗传结构和进化历史的研究日益受到关注，因为此类研究不仅能加深人们对作物起源和驯化的认识，而且有助于作物野生资源的开发以及作物的遗传改良。分子谱系地理分析（Molecular phylogeography）是此类研究中的重要手段之一，能够有效地了解近缘群体的进化历程以及重建作物的驯化历史。菰（Zizania latifolia Turcz.）是东亚广泛分布、具有重要价值的水生草本植物，其栽培型—茭白是中国第二大水生蔬菜。尽管已有不少涉及此类群的研究，但迄今我们对菰的群体遗传结构和进化历史以及茭白的起源几乎一无所知。本研究从29个基因片段中筛选了2个在菰种内水平上具有显著变异量的核基因片段，进行了分子谱系地理研究，揭示了菰的遗传多样性水平及其地理分布格局，探讨了菰的群体演化历史，并初步探讨了茭白的起源和驯化历史。主要结果如下： 1）基因片段筛选 开展分子谱系地理学研究，需要采用具有显著种内变异的分子标记，为此我们通过预实验对一批备选基因片段进行了筛选。在18个叶绿体和2个线粒体片段中，测序的总长度达17100 bp，但没有发现任何变异位点。在9个核基因片段中，7个片段具有变异位点，但只有2个片段的变异量在1%以上，分别是Adh1a（2.70%）和GPATb（1.25%）基因。所以，本研究最后确定了采用这2个核基因片段来进行菰的分子谱系地理的研究。 2）Adh1a基因谱系地理分析 对来自21个天然群体126个个体，通过Adh1a基因的直接PCR和克隆测序共得到了252条序列，中性检测表明，Adh1a基因符合中性进化模型、没有重组并且有足够的变异量，适合用于进行菰的谱系地理研究和进化历史的推断。在物种水平上，菰的核苷酸多态性水平为θsil = 0.0089，跟一些作物近缘类群相比处于中等偏低的水平;不同群体的核苷酸多态性水平相差明显，多态性最高的群体出现在东北地区，并有从北向南递减的趋势。群体间存在着显著的遗传分化（FST = 0.481）表明群体之间的基因流有限。群体间的遗传距离与地理距离没有相关性，地理邻近的群体并没有共享关系相近的单倍型，整个群体缺乏明显的谱系地理结构。目前的遗传多样性分布格局可能是由于近几十年来的生境退化和丧失造成过去群体的片段化所致。根据群体遗传多样性水平的差异和基因谱系的地理分布格局以及分子钟估算的与北美近缘种的分化时间，我们推断菰的群体经历了从东北向华南逐步扩散的过程。 3）GPATb基因谱系地理分析 为更可靠地推断菰的谱系地理格局，我们选择了GPATb基因作为标记开展了进一步的分析。对相同的采样群体和个体共252条序列的分析表明，在物种水平上，GPATb的核苷酸多态性只相当于Adh1a基因的1/4。综合2个基因的结果，可以认为菰的核苷酸多态性水平是维持在较低的水平上。该位点体现的群体遗传分化也没有Adh1a基因明显。群体之间相近的遗传多样性水平和较低分化程度可能意味着该基因位点受到平衡选择作用的影响，这一推论也得到了中性检测结果的支持。因此，GPATb基因可能受到选择作用的影响，不能用来支持或推翻Adh1a基因的结果。 4）茭白的起源和驯化 对来自13个省份的65个茭白品种的Adh1a和GPATb基因片段进行测序，结果发现，所有的品种在这2个基因上都具有相同的基因型，Adh1a位点上表现为杂合体，而GPATb位点上则是纯合体。这说明在茭白的起源过程中发生了严重的驯化瓶颈效应，也即茭白是单次起源的，是从菰的少数几个甚至单个个体驯化而来。虽然文献考据表明，茭白最早的栽培地区为太湖流域，但我们的分子证据却并不支持其为茭白的驯化起源地点，而可能是在太湖流域以北地区起源的。鉴于采样的局限性，这一结论还有待于进一步的验证。同时2个基因的数据表明来自陕西、四川、云南、湖南和福建的6个菰群体可能是从人为引种的茭白逃逸成为野生的。因此，在以后野生菰群体的采集中，特别是在茭白种植地区，要在详细调查的基础上设计采集方案。

Small mammals of the mid-Araguaia River in central Brazil, with the description of a new species of climbing rat

The mid-Araguaia River basin in central Brazil is considered a priority area for biodiversity conservation, and Parque Estadual do Cantao (PEC) is one of the most important protected areas in this ecotone between Cerrado and Amazonia. This area suffers an intensive human pressure with high rates of deforestation, and still remains poorly studied in terms of biodiversity. From June 2007 to November 2008 we sampled small mammals from both banks of the mid-Araguaia River, in the states of Tocantins and Para. Data are given about morphological traits, geographic distribution and natural history of 22 species of small non-volant mammals (eight marsupials and 14 rodents) surveyed at PEC and its surroundings. We also present mitochondrial phylogenetic analyses that allow species identification within the genera: Oecomys, Oligoryzomys and Rhipidomys, and delineate an undescribed species of Thrichomys. Based on morphologic and molecular data, we describe a new species of Rhipidomys previously assigned to R. nitela, which is apparently endemic to the Araguaia-Tocantins basin in the Cerrado. Additionally, our phylogenetic analyses provide support for the role played by the Araguaia River as an important geographic barrier for two sister species of Rhipidomys.

Molecular Phylogeny and Phylogeography of Suidae

猪科动物是最为昌盛的哺乳动物之一,但它们的分类及系统发育关系却尚待解决。为此,用PCR直接测序法测定了一个红河猪(Potamochoerusporcus)、一个马来西亚野猪(Susbarbatus)以及数头欧亚野猪(Susscorfa)线粒体细胞色素b全序列1140bp。结合从GenBank中获得的其他猪科物种胞色素b序列,用邻接法和最大简约法构建了猪科物种的系统发育关系,其结果显示苏拉威西鹿猪与其他猪科物种为姐妹群的关系。非洲物种疣猪和河猪聚为一枝,而欧亚猪属物种聚为另一支。各物种在系统发育关系中的位置与它们的地理分布对应。猪属物种可以被划分为3个种组:分布于菲律宾群岛、苏拉威西岛及其邻近岛屿的猪属物种形成一个种组,包括苏拉威西岛疣猪、菲律宾疣猪和印度尼西亚疣猪;欧亚野猪种组,包括欧亚野猪和姬猪;须猪和爪哇疣猪形成一个种组。

Impacts of climatic factors on evolution of molecular diversity and the natural distribution of wild stocks of the giant freshwater prawn (Macrobrachium rosenbergii)

Natural distributions of most freshwater taxa are restricted geographically, a pattern that reflects dispersal limitation. Macrobrachium rosenbergii is unusual because it occurs naturally in rivers from near Pakistan in the west, across India and Bangladesh to the Malay Peninsula, and across the Sunda Shelf and Indonesian archipelago to western Java. Individuals cannot tolerate full marine conditions, so dispersal between river drainage basins must occur at limited geographical scales when ecological or climatic factors are favorable. We examined molecular diversity in wild populations of M. rosenbergii across its complete natural range to document patterns of diversity and to relate them to factors that have driven evolution of diversity in this species. We found 3 clades in the mitochondrial deoxyribonucleic acid (mtDNA) data set that corresponded geographically with eastern, central, and western sets of haplotypes that last shared a common ancestor 1 × 106 y ago. The eastern clade was closest to the common ancestor of all 3 clades and to the common ancestor with its congener, Macrobrachium spinipes, distributed east of Huxley's Line. Macrobrachium rosenbergii could have evolved in the western Indonesian archipelago and spread westward during the early to mid-Pleistocene to India and Sri Lanka. Additional groups identified in the nuclear DNA data set in the central and western clades probably indicate secondary contact via dispersal between regions and modern introductions that have mixed nuclear and mtDNA genes. Pleistocene sea-level fluctuations can explain dispersal across the Indonesian archipelago and parts of mainland southeastern Asia via changing river drainage connections in shallow seas on wide continental shelves. At the western end of the modern distribution where continental shelves are smaller, intermittent freshwater plumes from large rivers probably permitted larval dispersal across inshore areas of lowered salinity.