Isolation and characterization of 11 polymorphic microsatellite loci in the whitegirdled goby (Pterogobius zonoleucus) and cross-species amplification in the serpentine goby (P. elapoides)

Eleven polymorphic microsatellite markers were developed from a (CA)(n) -enrichment library of the whitegirdled goby (Pterogobius zonoleucus). Polymorphism at these loci ranged from 2 to 12 alleles, and observed and expected heterozygosities from 0.05 to 0.90 and from 0.05 and 0.86, respectively. All loci conformed to Hardy-Weinberg equilibrium, with no significant linkage disequilibrium between all locus pairs. Cross-species amplification tests were successful in P. elapoides, and most loci were polymorphic. These microsatellite markers will be useful in further population genetic studies of both species.     

芍药属牡丹组基于形态学证据的系统发育关系分析

对芍药属牡丹组Paeonia L.sect.Moutan DC.（全部野生种）40个居群进行了基于形态学证据的系统学分析,试图建立组内种间的系统发育关系。利用PAUP （4.0）计算机程序分别构建了建立在25个形态学性状基础上的所有研究类群的距离树（UPGMA、NJ）和最大简约树（MP）。所得树的拓扑结构基本一致,差异只发生在距离树和简约树之间,在由形态和细胞学关系都很近的5个种（牡丹P.suffruticosa、矮牡丹P.jishanensis、卵叶牡丹P.qiui、紫斑牡丹P.rockii和凤丹P.o     

Genetic and morphological differentiation of the mangrove crab Perisesarma guttatum (Brachyura: Sesarmidae) along an East African latitudinal gradient

The genetic structure and morphometric differentiation of mangrove crab Perisesarma guttatum populations were examined among shelf connected locations along a latitudinal gradient on the East African coast. Over 2200 specimens were sampled from 23 mangrove sites for geometric morphometrics analysis. Population genetic analyses of mitochondrial cytochrome c oxidase subunit I (COI) DNA sequences were used to evaluate connectivity among populations. A total of 73 haplotypes were detected, and almost no haplotypes were found in common between two highly supported phylogeographic clades: southern Mozambique (Inhaca Island and Maputo Bay) and a northern clade that included north Mozambique, Tanzania and Kenya. These two clades were identified based on the species' populations pairwise genetic differentiation and geographical location. Φ_ST values were considerably high between the two clades, indicating the presence of significant population genetic structure between Kenya and South Mozambique. However, each clade was composed of genetically similar populations along the latitudinal gradient, and no significant population structure was found within each clade because the Φ_st values were not significant. The morphometric analysis corroborated the division into two clades (i.e. Inhaca Island/Maputo Bay and northern populations) and also detected less shape variation among populations that were few kilometres apart. The significant spatial genetic structuring between the southern and the northern populations of P. guttatum along the geographic gradient under study, combined with morphological differences, suggests that these populations may be considered as cryptic species. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 99 , 28–46.     

Genetic diversity of Echinacea species based upon amplified fragment length polymorphism markers.

The taxonomy of Echinacea is based on morphological characters and has varied depending on the monographer. The genus consists of either nine species and four varieties or four species and eight varieties. We have used amplified fragment length polymorphisms (AFLP) to assess genetic diversity and phenetic relationships among nine species and three varieties of Echinacea (sensu McGregor). A total of 1086 fragments, of which approximately 90% were polymorphic among Echinacea taxa, were generated from six primer combinations. Nei and Li's genetic distance coefficient and the neighbor-joining algorithm were employed to construct a phenetic tree. Genetic distance results indicate that all Echinacea species are closely related, and the average pairwise distance between populations was approximately three times the intrapopulation distances. The topology of the neighbor-joining tree strongly supports two major clades, one containing Echinacea purpurea, Echinacea sanguinea, and Echinacea simulata and the other containing the remainder of the Echinacea taxa (sensu McGregor). The species composition within the clades differs between our AFLP data and the morphometric treatment offered by Binns and colleagues. We also discuss the suitability of AFLP in determining phylogenetic relationships.     

Phylogeny and evolution of basils and allies (Ocimeae, Labiatae) based on three plastid DNA regions

A phylogeny of basils and allies (Lamiaceae, tribe Ocimeae) based on sequences of the trnL intron, trnL-trnF intergene spacer and rps 16 intron of the plastid genome is presented. Several methods were used to reconstruct phylogenies and to assess statistical support for clades: maximum parsimony with equally and successively weighted characters, bootstrap resampling, and Bayesian inference. The phylogeny is used to investigate the distribution of morphological, pericarp anatomy, chemical, and pollen characters as well as the geographical distribution of the clades. Tribe Ocimeae is monophyletic and easily diagnosable with morphological synapomorphies. There are monophyletic clades within Ocimeae that broadly correspond to currently recognised subtribes: Lavandulinae, Hyptidinae, Ociminae, and Plectranthinae. Only Lavandulinae has clear non-molecular synapomorphies. Several currently recognised genera are not monophyletic. Floral morphology consistent with sternotribic pollination is most common in Ocimeae, but there are independent departures from this model. Buzz pollination is likely in some species, the only postulated occurrence of this within Lamiaceae. Quinone diterpenoids and flavones in the leaf exudates differ in their distributions across the phylogeny and this could contribute to differences in the recorded medicinal as well as pesticidal uses of the species in the different clades. Mapping geographic distribution on to an ultrametric phylogenetic tree produced using non-parametric rate smoothing supports an Asiatic origin for Ocimeae. There are several secondary occurrences in Asia arising from the African Ociminae and Plectranthinae clades. Colonisation of Madagascar occurred at least five times, and New World colonisation occurred at least three times.     

An evaluation of the monophyly of Massarina based on ribosomal DNA sequences

The monophyletic status of the genus Massarina was evaluated on the basis of phylogenetic analysis of the partial small subunit gene (SSU), internal transcribed spacers (ITS 1 & 2), and 5.8S gene sequences of the ribosomal DNA. Species of Massarina used in the study clustered into two distinct clades with high bootstrap support in trees generated from maximum parsimony, weighted parsimony, maximum likelihood, and neighbor-joining analyses. The hypothesis that Massarina species belong to a phylogenetically monophyletic group is rejected. Species with narrowly fusiform ascospores form a monophyletic clade with Lophiostoma, a genus highly similar in morphology. The five species currently accepted in Massarina with such spore morphology are here transferred into the genus Lophiostoma. Massarina species with broadly fusiform to ellipsoidal ascospores are retained as Massarina s. str., lectotypified by M. eburnea. Massarina walkeri is presently excluded from both Massarina and Lophiostoma. The transfer of M. papulosa to a new genus Oletheriostrigula is verified.     

A Phylogenetic Analysis of the Plastid matK Gene with Emphasis on Melanthiaceae sensu lato

Abstract: The presented mat K tree primarily agrees well with the previously presented rbc L tree and combined rbc L + atp B + 18SrDNA tree. According to the mat K tree, the monocotyledons are monophyletic with 100 % bootstrap support. Acorus diverges first from all other monocotyledons (90 % bootstrap support) in which two major clades are recognized: one (89 %) consisting of Alismatanae and Tofieldia (Nartheciaceae), and the other (< 50 %) comprising Lilianae, Commelinanae and Nartheciaceae other than Tofieldia. Within the latter major clade, Petrosavia and Japonolirion (Nartheciaceae) (82 %) diverge first from the remaining taxa (< 50 %) in which two clades are formed: one (81 %) consisting of Pandanales, Dioscoreales and Nartheciaceae-Narthecioideae, and the other (< 50 %) comprising Liliales, Asparagales and Commelinanae. In the former clade, Dioscoreales and Narthecioideae are grouped together (88 %). In the latter clade, Asparagales and Commelinanae are grouped together (< 50 %). Differences between the mat K and rbc L tree topologies appear in the positions of Tricyrtis (Calochortaceae) and Dracaenaceae. Differences between the mat K and combined rbc L + atp B + 18SrDNA tree topologies exist in the positions of the Petrosavia-Japonolirion pair (Nartheciaceae) and Pandanales. The stop codon position of the mat K gene appears to be highly variable among the monocotyledons, especially in the Liliales.     

Phylogenetic relationships of the Subclass Peniculia (Oligohymenophorea, Ciliophora) inferred from small subunit rRNA gene sequences

Peniculine ciliates have been recognized as a distinct higher taxon of ciliates for almost 50 years. However, phylogenetic relationships within the Subclass Peniculia are still unsettled. To contribute to our understanding of their phylogeny and provide evidence for the position of Urocentrum turbo, we sequenced its small subunit (SS) rRNA gene and the SSrRNA genes from Lembadion bullinum, Frontonia sp., Paramecium caudatum, Paramecium multimicronucleatum, Paramecium putrinum, and Paramecium woodruffi. Urocentrum turbo was the only one of these species not to exhibit a shortened Helix E10_1, which we conclude characterizes the "higher" peniculines. Except for U. turbo, the peniculines are strongly supported as a monophyletic clade with Lembadion, Frontonia, and Paramecium species forming separate and strongly supported clades by bootstrap analysis using distance matrix, maximum parsimony, and maximum likelihood methods. Urocentrum turbo is associated with different lineages, depending upon the analysis used. The Paramecium species form at least four clades with the Paramecium aurelia subgroup being the most derived. We conclude that the Subclass Peniculia should be divided into two orders, the Order Urocentrida and Order Peniculida, with the latter order having two suborders, the Suborder Frontoniina and Peniculina. We place U. turbo with the peniculines because of shared morphological and stomatogenetic features.     

PHYLOGENY OF THE CONJUGATING GREEN ALGAE (ZYGNEMOPHYCEAE) BASED ON rbc L SEQUENCES

Sequences of the gene encoding the large subunit of RUBISCO (rbcL) for 30 genera in the six currently recognized families of conjugating green algae (Desmidiaceae, Gonatozygaceae, Mesotaeniaceae, Peniaceae, and Zygnemataceae) were analyzed using maximum parsimony and maximum likelihood; bootstrap replications were performed as a measure of support for clades. Other Charophyceae sensu Mattox and Stewart and representative land plants were used as outgroups. All analyses supported the monophyly of the conjugating green algae. The Desmidiales, or placoderm desmids, constitute a monophyletic group, with moderate to strong support for the four component families of this assemblage (Closteriaceae, Desmidiaceae, Gonatozygaceae, and Peniaceae). The analyses showed that the two families of Zygnematales (Mesotaeniaceae, Zygnemataceae), which have plesiomorphic, unornamented and unsegmented cell walls, are not monophyletic. However, combined taxa of these two traditional families may constitute a monophyletic group. Partitioning the data by codon position revealed no significant differences across all positions or between partitions of positions one and two versus position three. The trees resulting from parsimony analyses using first plus second positions versus third position differed only in topology of branches with poor bootstrap support. The tree derived from third positions only was more resolved than the tree derived from first and second positions. The rbcL‐based phylogeny is largely congruent with published analyses of small subunit rDNA sequences for the Zygnematales. The molecular data do not support hypotheses of monophyly for groups of extant unicellular and filamentous or colonial desmid genera exhibiting a common cell shape. A trend is evident from simple omniradiate cell shapes to taxa with lobed cell and plastid shapes, which supports the hypothesis that chloroplast shape evolved generally from simple to complex. The data imply that multicellular placoderm desmids are monophyletic. Several anomalous placements of genera were found, including the saccoderm desmid Roya in the Gonatozygaceae and the zygnematacean Entransia in the Coleochaetales. The former is strongly supported, although the latter is not, and Entransia's phylogenetic position warrants further study.     

A comparison of the intraspecific variability of Phlebotomus sergenti Parrot, 1917 (Diptera: Psychodidae).

Phlebotomus sergenti populations from different areas of the Mediterranean basin are known to exhibit high intraspecific variability. Previous studies of ITS2 revealed the presence of two branches that may represent sibling species. To corroborate this finding by other tools, two colonies of P. sergenti originating from Turkey and Israel, each belonging to a different ITS2 branch, were compared by three different methods: geometric morphometric analysis of wing shape, RAPD (random amplified polymorphic DNA), and cross-mating study. For geometric morphometric analysis, two-dimensional Cartesian coordinates of 16 landmarks from the wings were digitized and analyzed. Significant shape differences were found between colonies but not between sexes within each colony. RAPD results formed two distinctive clades corresponding to the origin of the colony but also showed heterogenity among members of both colonies. In cross-mating studies, viable hybrid F1 and F2 progeny were obtained when both Turkish males/Israeli females and Israeli males/Turkish females were crossed. F1 progeny was included in RAPD analysis and these hybrids formed a distinctive clade with an intermediate position between the two parental clades. No significant differences were found in egg production of crossed sand flies. The cross-mating study showed that there is no reproductive barrier between P. sergenti from different geographical areas. On the other hand, RAPD and geometric morphometric analysis revealed a significant difference between colonies and confirmed the suitability of previous ITS2 analysis for discrimination among sand fly populations. Further development of molecular markers should resolve a possible existence of sibling species within Phlebotomus sergenti.     

Reassessment of morphological characteristics in freshwater eels (genus Anguilla, Anguillidae) shows congruence with molecular phylogeny estimates

The morphology-based phylogeny of freshwater eels, proposed by V. Ege in 1939, has been accepted as the basis of eel classification since that time. However, this has been called into question by recent molecular studies. Most of the morphological characteristics recognized by Ege are morphometric. Since methods for the application of morphometric data to phylogeny construction have not been fully established, it is unclear whether the observed discrepancies between morphological and molecular data arise from intrinsic differences or from flawed analyses. Here, we have used two methods to assemble evolutionary trees from distance matrices constructed according to Ege's data, the neighbor-joining (NJ) method and the minimum network (MinNet) method; the latter is based on an evolutionary algorithm. After reanalysing Ege's morphological data, we found that both methods gave results consistent with those based on molecular data, although not with Ege's original classification. Therefore, we speculate that some morphological features Ege used to subdivide the eel groups may not be synapomorphic as he proposed, but symplesiomorphic or convergent . The method developed here may prove useful for constructing phylogeny for taxon groups where only continuous morphometric characteristics are recognized, such as the freshwater eels.     

Phylogenetic analysis of the SSU rRNA from members of the Chrysophyceae

The nucleotide sequence for the nuclear-encoded small subunit ribosomal RNA gene (SSU rRNA) was determined for 24 species of the Chrysophyceae sensu stricto. These sequences were aligned, using primary and secondary structure, with nine previously published sequences for the Chrysophyceae, 14 for the Synurophyceae, and five for the Eustigmatophyceae (outgroup). Data analyses were the substitution rate calibration distance method using neighbor-joining (TREECON), Kimura 2-parameter neighbor-joining method (PAUP) and the maximum parsimony method (PAUP, PHYLIP). Trees from the analyses were largely congruent, but bootstrap support was weak at many nodes. The analyses recovered clades of uniflagellate and biflagellate organisms associated with current higher level taxonomy (e.g., subclass, order). The genus Ochromonas was polyphyletic, and O. tuberculata in particular was distantly related to the other Ochromonas species in the analysis. The family Paraphysomonadaceae occupied a basal position in three of four analyses. The class Synurophyceae appeared to be embedded within the Chrysophyceae, but bootstrap support was weak (< 50%) in all analyses except the PHYLIP parsimony analysis (= 81%). It was considered premature to place the Synurophyceae back into the Chrysophyceae based upon the analysis of one gene, especially given the ultrastructural and pigment differences between the two groups, but the relationship of these two groups deserves further study.     

Neglected taxonomy of rare desert fishes: congruent evidence for two species of leatherside chub

Conservation biologists rely heavily on taxonomy to set the scope for biological monitoring and recovery planning of rare or threatened species. Yet, taxonomic boundaries are seldom evaluated as falsifiable hypotheses that can be statistically tested. Here, we examine species boundaries in leatherside chub (Teleostei, Cyprinidae), an imperiled desert fish native to the Bonneville Basin and upper Snake River drainages of western North America. Recent molecular data hint that this fish could be composed of two distinct taxa that are geographically separated into northern and southern species. To formally test this hypothesis, we evaluated leatherside chub using several different categories of species concepts, including criteria dependent on phylogenetic, morphological, and ecological data. We found that leatherside chub is composed of two reciprocally monophyletic clades (candidate species) characterized by numerous fixed genetic differences for both mitochondrial and nuclear DNA markers; mtDNA sequence divergence between the two clades approached 8%. The candidate species also showed significant differences in cranial shape, revealed by morphometric analysis. Finally, controlled growth and foraging experiments using representative populations from each clade show that candidate species appear to be locally adapted to the thermal environments where they now occur. Combined, these three lines of evidence support the hypothesis that leatherside chub is composed of two species. Moreover, all lines of evidence place these two species within the genusLepidomeda, a group consisting of three additional species of endangered spinedace fishes, and one extinct species, all native to the Colorado River system. Hence, we elevate the two clades of leatherside chub to distinct species status (Lepidomeda copeiin the north andL. aliciaein the south), and argue that each warrants independent conservation and recovery action.     

CONFIDENCE LIMITS ON PHYLOGENIES: THE BOOTSTRAP REVISITED

Abstract— The bootstrap, a non-parametric statistical analysis, can be used to assess confidence limits on phylogcnics. The method most widely used tests the monophyly of individual clades. This paper proposes additional applications of the bootstrap which provide useful information about phylogeny even when many clades are found not to be supported with confidence (as often occurs in practice). In such cases it is still possible to place a constraint on the phylogenetic position of taxa by examining the relative size of the smallest monophyletic groups that contain them. In addition, the taxonomic composition of these larger clades can be determined, as well as the relative likelihood of their occurrence. The distinction between hypotheses about membership in particular clades and hypotheses about entire topologies is also discussed. To investigate the latter, the bootstrap is used to estimate the sampling distribution of tree similarity indices. All methods are illustrated by reference to a large data set on the angiosperm family Asteraccae, selected from the literature.     

Morphometric and meristic characteristics of two groups ofLethenteron reissneri

Morphometric and meristic characteristics were compared between two genetically divergent groups ofLethenteron reissneri, collected from Hokkaido and Honshu islands, Japan. Whereas significant differences were found in some morphometric and meristic characters, the ranges overlapped somewhat in all of the characters examined between the two groups. Thus, it was difficult to distinguish exactly between the two groups using only the morphological characters. In sympatric populations of the two groups, no significant differences were found in almost all of the morphometric and meristic characters examined. The two groups ofL. reissneri, which are reproductively isolated from each other, should be regarded as sibling species. Assortative mating, based on morphological features, appears unlikely as the reproductive isolating mechanism between the two groups.     

Application of molecular genetics and geometric morphometrics to taxonomy and conservation of cave beetles in central Italy

Integration of molecular genetic techniques and geometric morphometrics represent a valuable tool in the resolution of taxonomic uncertainty and the identification of significant units for conservation. We combined mitochondrial DNA cytochrome c oxidase subunit II gene sequence data and geometric morphometric analysis to examine taxonomic status and identify units for conservation in four species of the hypogean beetle Duvalius (Coleoptera, Trechinae) using mainly museum specimens collected in central Italy. Previous taxonomic studies based on morphological traits described several subspecies often inhabiting geographically distinct caves. Phylogenetic analysis identified two well supported monophyletic lineages and a number of different clades with relatively small genetic differences, suggesting a short divergence time in line with known geological history of the study area. Geometric morphometrics, on the other hand, recovered a high level of distinctiveness among specimens. Both genetic and morphometric analyses did not entirely corroborate former taxonomic nomenclature, suggesting possible rearrangements and the definition of evolutionary significant units. Beetles of the genus Duvalius are protected by regional laws and the majority of taxa considered in this study inhabit caves located outside protected areas. Our study advocates the importance of devoting protection efforts to networks of cave ecosystems rather than single locations or species.     

美国红鱼生长激素基因的克隆及其遗传进化分析

采用PCR方法从美国红鱼肝脏中扩增出美国红鱼生长激素基因。该基因序列长为1 497 bp,与5个已报道的鲈亚目鱼类生长激素基因结构比较发现,他们的外显子和内含子之比均为6∶5,而且相对应的6个外显子大小一致。用Clustal X软件对美国红鱼和其他13种鲈形目鱼类生长激素基因序列进行比对,选取同源性较高的551 bp的编码区序列构建NJ和MP系统进化树,结果发现由其构建的MP和NJ树与根据形态特征构建的系统发育树基本一致,特别是在鲈形目鱼类科间及科以上分类阶元的系统发育研究方面比较一致。研究结果表明鱼类生长激素基因序列能够较真实反映近缘鱼类之间的关系,是进行鲈形目鱼类系统进化分析的较好遗传标志。     

Geographic variation in the molecular,morphological and reproductive characters of the ayuPlecoglossus altivelis (plecoglossidae) in the Japan-Ryukyu Archipelago

In an attempt to clarify geographic variation in the ayuPlecoglossus altivelis, the molecular, morphological and reproductive characters of populations from various localities in the Japan-Ryukyu Archipelago were examined. An electrophoretic analysis of enzymes encoded by 18–28 loci and univariate and multivariate analyses of 12 meristic and 17 morphometric characters indicated that there are two distinct groups of ayu in the archipelago: that of the Japan Islands and that of the Ryukyu Islands. Within the Japan Islands group, a landlocked population of Lake Biwa was found to diverge somewhat from other amphidromous populations genetically and morphologically. The Lake Biwa population was noted as being by far the most fecund, departing from a north-south cline of fecundity observed in amphidromous populations. Though no virtual divergence was observed among amphidromous populations in the Japan Islands in molecular data, divergence among these populations was considerable in morphological and fecundity data. Differences in geographic variation patterns are attributed to qualitative differences inherent in molecular and organismal characters: organismal characters are responsive to selection and also directly altered by environmental influences in part but molecular characters are hardly so. The evolution of different characters may have proceeded at different rates among geographic populations of ayu.     

Spot-fixed fin digging behavior in foraging of the benthophagous maiden goby, Pterogobius virgo (Perciformes: Gobiidae)

 Several patterns of feeding behaviors have been documented in benthophagous fishes. The foraging behavior of the maiden goby, Pterogobius virgo, was studied at Kurahashi Island in the Seto Inland Sea, Japan. Pterogobius virgo foraged mostly on polychaetes by volume from among several available prey items by digging in the sandy bottom. The digging behavior comprised swing of only pectoral fins or of both pectoral fins and body. Pectoral fin swing exposed the cryptic prey within the bottom, and fins and body swing exposed the prey and washed the sediment away. The swings were repeatedly and continuously conducted at a site during the daytime, making a pit several centimeters deep in which the fish was located. After the prey was exposed, the fish immediately and rapidly picked up the prey. Polychaetes were abundant prey in the sediment, occurring in the layer 3–5 cm deep from the bottom surface in the study area. In this goby, spot-fixed fin digging, the first documentation of feeding habits in gobies, may be effective for feeding on the most valuable prey, i.e., polychaetes, which may be otherwise unavailable for this fish. Received: April 24, 2001 / Revised: April 26, 2002 / Accepted: May 7, 2002