Genetic diversity of cultured populations of giant freshwater prawn (Macrobrachium rosenbergii) in China using mtDNA COI and 16S rDNA markers

Partial sequences of mitochondrial DNA 16S rDNA and COI genes (395 bp and 498 bp respectively) were sequenced from samples of ten cultured populations of Macrobrachium rosenbergii (Giant Freshwater Prawn – GFP) in Zhejiang, Guangdong and Guangxi Provinces in China and two wild populations of GFP from Mekong River and Dongnai River in Viet Nam. Five haplotypes of 16S rDNA were identified in the 360 samples. The wild populations displayed nucleotide diversity (π) of 0.0008 and 0.0003, and genetic diversity (h) of 0.3030 and 0.1310 in the Mekong River and Dongnai River respectively. The cultured populations displayed no significant genetic diversity. COI sequences identified 17 haplotypes based on 21 polymorphic sites. At this marker, the 12 populations showed a range of h from 0.1290 to 0.6940 and π from 0.0003 to 0.0073. The largest genetic distance (Da) among the 12 populations was 0.0065 (between ZJB and BT/DN populations) and the lowest Da was 0.0003 (between GDD and GDA populations). The wild populations had higher genetic diversity than the cultured populations, but three of cultured populations from Zhejiang (ZJA, ZJB and ZJC) had π higher than wild populations, because they originated from Thailand, Bangladesh and the Mekong River in Viet Nam.     

Genetic Diversity and Population Structure for the Conservation of Giant Spiny Frog(Quasipaa spinosa) Using Microsatellite Loci and Mitochondrial DNA

The giant spiny frog(Quasipaa spinosa) is an endangered species with a relatively small distribution limited to southern China and Northern Vietnam. This species is becoming increasingly threatened because of over-exploitation and habitat degradation. This study provides data on the genetic diversity and population genetic structure of the giant spiny frog to facilitate the further development of effective conservation recommendations for this economically important but threatened species. We examined 10 species-specific microsatellite loci and Cyt b genes(562 bp) collected from 13 wild populations across the entire range of this species. Results of 10 microsatellite loci analysis showed a generally high level of genetic diversity. Moreover, the genetic differentiation among all 12 populations was moderate to large(overall F_(ST) = 0.1057). A total of 51 haplotypes were identified for Cyt b, which suggests high haplotype nucleotide diversities. Phylogeographic and population structure analyses using both DNA markers suggested that the wild giant spiny frog can be divided into four distinct major clades, i.e., Northern Vietnam, Western China, Central China, and Eastern China. The clades with significant genetic divergence are reproductively isolated, as evidenced by a high number of private alleles and strong incidence of failed amplification in microsatellite loci. Our research, coupled with other studies, suggests that Q. spinosa might be a species complex within which no detectable morphological variation has been revealed. The four phylogenetic clades and some subclades with distinct geographical distribution should be regarded as independent management units for conservation purposes.     

In Memoriam: Professor Walter James Harman, PhD (1928–2002)

The Mediterranean spiny lobsters, the common spiny lobster Palinurus elephas (Fabricius, 1758) and the pink spiny lobster P. mauritanicus Gruvel, 1911, are important target species for commercial fisheries. In this study, we focus our attention on the DNA sequence variation of the mitochondrial cytochrome oxydase I gene (COI) in the two species of Palinurus. Spiny lobster DNA samples from four Mediterranean localities were analysed to examine the genetic variability at both the intra- and interspecific level. Furthermore, the phylogenetic relationships within the family Palinuridae (among the two species of Palinurus, most of the species of Panulirus and all the species of Jasus) are examined.     

DNA barcodes for globally threatened marine turtles: a registry approach to documenting biodiversity

DNA barcoding is a global initiative that provides a standardized and efficient tool to catalogue and inventory biodiversity, with significant conservation applications. Despite progress across taxonomic realms, globally threatened marine turtles remain underrepresented in this effort. To obtain DNA barcodes of marine turtles, we sequenced a segment of the cytochrome c oxidase subunit I (COI) gene from all seven species in the Atlantic and Pacific Ocean basins (815 bp; n = 80). To further investigate intraspecific variation, we sequenced green turtles (Chelonia mydas) from nine additional Atlantic/Mediterranean nesting areas (n = 164) and from the Eastern Pacific (n = 5). We established character-based DNA barcodes for each species using unique combinations of character states at 76 nucleotide positions. We found that no haplotypes were shared among species and the mean of interspecific variation ranged from 1.68% to 13.0%, and the mean of intraspecific variability was relatively low (0–0.90%). The Eastern Pacific green turtle sequence was identical to an Australian haplotype, suggesting that this marker is not appropriate for identifying these phenotypically distinguishable populations. Analysis of COI revealed a north–south gradient in green turtles of Western Atlantic/Mediterranean nesting areas, supporting a hypothesis of recent dispersal from near equatorial glacial refugia. DNA barcoding of marine turtles is a powerful tool for species identification and wildlife forensics, which also provides complementary data for conservation genetic research.     

Integrative taxonomy and molecular phylogeny of genus aplysina (demospongiae: verongida) from mexican pacific

Integrative taxonomy provides a major approximation to species delimitation based on integration of different perspectives (e.g. morphology, biochemistry and DNA sequences). The aim of this study was to assess the relationships and boundaries among Eastern Pacific Aplysina species using morphological, biochemical and molecular data. For this, a collection of sponges of the genus Aplysina from the Mexican Pacific was studied on the basis of their morphological, chemical (chitin composition), and molecular markers (mitochondrial COI and nuclear ribosomal rDNA: ITS1-5.8-ITS2). Three morphological species were identified, two of which are new to science. A. clathrata sp. nov. is a yellow to yellow-reddish or -brownish sponge, characterized by external clathrate-like morphology; A. revillagigedi sp. nov. is a lemon yellow to green, cushion-shaped sometimes lobate sponge, characterized by conspicuous oscules, which are slightly elevated and usually linearly distributed on rims; and A. gerardogreeni a known species distributed along the Mexican Pacific coast. Chitin was identified as the main structural component within skeletons of the three species using FTIR, confirming that it is shared among Verongida sponges. Morphological differences were confirmed by DNA sequences from nuclear ITS1-5.8-ITS2. Mitochondrial COI sequences showed extremely low but diagnostic variability for Aplysina revillagigedi sp. nov., thus our results corroborate that COI has limited power for DNA-barcoding of sponges and should be complemented with other markers (e.g. rDNA). Phylogenetic analyses of Aplysina sequences from the Eastern Pacific and Caribbean, resolved two allopatric and reciprocally monophyletic groups for each region. Eastern Pacific species were grouped in general accordance with the taxonomic hypothesis based on morphological characters. An identification key of Eastern Pacific Aplysina species is presented. Our results constitute one of the first approximations to integrative taxonomy, phylogeny and evolutionary biogeography of Eastern Pacific marine sponges; an approach that will significantly contribute to our better understanding of their diversity and evolutionary history.     

Genetic structure of the endangered species <Emphasis Type="Italic">Pinna nobilis</Emphasis> (Mollusca: Bivalvia) inferred from mtDNA sequences

This study examines the population genetic structure of the endangered bivalve Pinna nobilis (Mollusca: Bivalvia), based on novel mtDNA sequences (partial COI and 16S rDNA mtDNA genes). The analyzed nucleotide sequences of COI were 729 bp in size, coding for a 243 amino acid peptide, while the analyzed nucleotide sequences of 16S rDNA were 489 bp in size. These sequences of P. nobilis were the first DNA sequences of the species submitted to any Genetic Data Base. Population samples from four geographic regions from Greece, as well as a population sample of Atrina fragilis (as an outgroup) were used. High values of haplotypic diversity were found in the population samples of P. nobilis, based on the COI sequences. A single base in the analyzed 16S rDNA sequences was different in all analyzed individuals from a single population sample (Chios island) differentiating it from the other ones. These mtDNA sequences could be informative for further genetic analyses of the endangered species, contributing in conservation plans for its protection and/or aquaculture investigations.     

Increased inter-colony fusion rates are associated with reduced COI haplotype diversity in an invasive colonial ascidian Didemnum vexillum

Considerable progress in our understanding of the population genetic changes associated with biological invasions has been made over the past decade. Using selectively neutral loci, it has been established that reductions in genetic diversity, reflecting founder effects, have occurred during the establishment of some invasive populations. However, some colonial organisms may actually gain an ecological advantage from reduced genetic diversity because of the associated reduction in inter-colony conflict. Here we report population genetic analyses, along with colony fusion experiments, for a highly invasive colonial ascidian, Didemnum vexillum. Analyses based on mitochondrial cytochrome oxidase I (COI) partial coding sequences revealed two distinct D. vexillum clades. One COI clade appears to be restricted to the probable native region (i.e., north-west Pacific Ocean), while the other clade is present in widely dispersed temperate coastal waters around the world. This clade structure was supported by 18S ribosomal DNA (rDNA) sequence data, which revealed a one base-pair difference between the two clades. Recently established populations of D. vexillum in New Zealand displayed greatly reduced COI genetic diversity when compared with D. vexillum in Japan. In association with this reduction in genetic diversity was a significantly higher inter-colony fusion rate between randomly paired New Zealand D. vexillum colonies (80%, standard deviation ±18%) when compared with colonies found in Japan (27%, standard deviation ±15%). The results of this study add to growing evidence that for colonial organisms reductions in population level genetic diversity may alter colony interaction dynamics and enhance the invasive potential of newly colonizing species.     

Detection of Panulirus argus Virus 1 (PaVI) in the Caribbean spiny lobster using fluorescence in situ hybridization (FISH)

Panulirus argus Virus 1 (PaV1) is the first virus known to be pathogenic to a wild lobster. It infects the Caribbean spiny lobster P. argus from the Florida Keys, and has a predilection for juveniles. The monitoring of the virus in wild populations and study of its behavior in the laboratory require the development of reliable diagnostic tools. A sensitive and specific fluorescence in situ hybridization (FISH) assay was developed for detection of PaV1. The lower detection limit using a 110 bp DNA probe in a dot-blot hygridization for PaV1 DNA was 10 pg of cloned template PaV1 DNA and 10 ng of genomic DNA extracted from the hemolymph of diseased spiny lobster. The fluorescein (FITC)-labeled probe specifically hybridized to PaVl-infected cells in the hepatopancreas, hindgut, gills, heart, foregut, and nerve tissues. FITC staining was observed around the inner periphery of the nuclear membrane, with lighter staining in a more dispersed pattern within the nucleus. The probe did not hybridize with host tissues of uninfected spiny lobsters, nor did it cross-react with 4 other virus samples tested. This assay will facilitate our understanding of the pathogenesis of the viral disease and help in monitoring efforts directed at determining the prevalence of PaV1 in juvenile nurseries for this lobster.     

Genetic Variation and Population Structure of Hair Crab (<Emphasis Type="Italic">Erimacrus isenbeckii</Emphasis> ) in Japan Inferred from Mitochondrial DNA Sequence Analysis

Genetic variation and population structure of hair crab (Erimacrus isenbeckii) were examined using nucleotide sequence analysis of 580 base pairs (bp) in the 3′ portion of the mitochondrial cytochrome c oxidase subunit I gene (COI) of 20 samples collected from 16 locales in Japan (the Hokkaido and Honshu Islands) and one in Korea. A total of 27 haplotypes was defined by 23 variable nucleotide sites in the examined COI region. Pairwise population F _ST estimates and neighbor-joining tree inferred distinct genetic differentiation between the representative samples from the Pacific Ocean off the Eastern Hokkaido Island and the Sea of Japan, while others were intermediate between these two groups. AMOVA also showed a weak but significant differentiation among these three groups. The present results suggest a moderate population structure of hair crab, probably influenced by high gene flow between regional populations due to sea current dependent larval dispersal of this species.     

Mechanisms of global diversification in the brown booby (Sula leucogaster) revealed by uniting statistical phylogeographic and multilocus phylogenetic methods

Recent theoretical and empirical research suggests that statistical models based on coalescent theory can improve both phylogeographic and phylogenetic inference. An approach that involves elements of both statistical phylogeography (e.g. Isolation with Migration analyses) and multilocus phylogenetic inference (e.g. *beast) may be particularly useful when applied to populations with relatively old divergence times. Here, we use such an approach in the globally distributed brown booby (Sula leucogaster). We sampled 215 individuals from all major breeding areas and genotyped them at eight microsatellite and three nuclear intron loci. We found that brown booby populations were highly differentiated and that colonies can be grouped into four major genetic populations (Caribbean Sea, Central Atlantic Ocean, Indo-Central Pacific and Eastern Pacific). These populations apparently diverged in the absence of gene flow and, with one exception, currently exchange few to no migrants. The Eastern Pacific population diverged from all other populations approximately one million years ago [90% highest posterior density: 330,000-2,000,000 years ago] and exhibits a distinct male plumage, relative to other populations. However, recent gene flow from the Indo-Central Pacific into the Eastern Pacific appears to have occurred, suggesting that approximately one million years of genetic isolation and divergence in male plumage colour are not sufficient to prevent interbreeding. Gene flow following secondary contact of the Indo-Central Pacific and Eastern Pacific populations was not detected in previous mitochondrial DNA (mtDNA) studies, and the contrast between the mtDNA results and our current results highlights the advantage of a multilocus phylogeographic approach.     

A molecular phylogenetic analysis of strombid gastropod morphological diversity

The shells of strombid gastropods show a wide variety of forms, ranging from small and fusiform to large and elaborately ornamented with a strongly flared outer lip. Here, we present the first species-level molecular phylogeny for strombids and use the resulting phylogenetic framework to explore relationships between species richness and morphological diversity. We use portions of one nuclear (325 bp of histone H3) and one mitochondrial (640 bp of cytochrome oxidase I, COI) gene to infer relationships within the two most species-rich genera in the Strombidae: Strombus and Lambis. We include 32 species of Strombus, representing 10 of 11 extant subgenera, and 3 of the 9 species of Lambis, representing 2 of 3 extant subgenera. Maximum likelihood and Bayesian analyses of COI and of H3 and COI combined suggest Lambis is nested within a paraphyletic Strombus. Eastern Pacific and western Atlantic species of Strombus form a relatively recent monophyletic radiation within an older, paraphyletic Indo-West Pacific grade. Morphological diversity of subclades scales positively with species richness but does not show evidence of strong phylogenetic constraints.     

DNA sequences and cross-breeding experiments in the hawthorn spider mite Amphitetranychus viennensis reveal high genetic differentiation between Japanese and French populations

Sequence variation of the complete second internal transcribed spacer (ITS2, 445 bp) of nuclear ribosomal DNA and part of the mitochondrial cytochrome oxidase I gene (COI, 350 bp) was examined in Amphitetranychus viennensis (Zacher) mites (Acari:Tetranychidae) from four French and four Japanese locations. Sequence analysis consistently revealed the separation of the samples in two major groups: French mites differed from Japanese by 3.8–4.1% of the nucleotide divergence in COI sequences. These two groups also displayed distinct ITS2 consensus sequences (2.1% nucleotide divergence). A few variations, not affecting the diagnostic sites around the consensus sequence, were revealed among cloned copies of the same individual. Reciprocal crosses and backcrosses between one French and two Japanese populations disclosed strong reproductive incompatibility. However, fertile hybrid females were obtained, indicating the conspecificity of the tested mites. Despite the presence of Wolbachia in the French strain, but not in the Japanese ones, our crosses did not display the unidirectional incompatibility typically produced by this microorganism, but rather a bidirectional – although asymmetrical – incompatibility pattern. The post-zygotic incompatibilities in A. viennensis cannot be explained by the presence of Wolbachia but to some extent by mite genome divergence resulting from limited gene exchange between allopatric populations. Experiments of Wolbachia elimination by antibiotic treatment and subsequent crosses with cured strains are still needed to fully understand the reproductive incompatibility patterns in this mite species.     

Assessment of three mitochondrial loci variability for the crown-of-thorns starfish: a first insight into Acanthaster phylogeography

Acanthaster planci (L.) is one of the major threats to coral reefs, whose genetic diversity has been mainly studied with allozymes. Allozymes revealed the low genetic differentiation between A. planci populations in the Indo-Pacific area. We obtained sequences of A. planci from Kenya, Mayotte and Madagascar at the three loci cytochrome oxydase subunit I (COI), 16S rDNA (16S) and five tRNAs, analysed together with available sequences of Acanthaster from the Pacific Ocean. The level of genetic diversity varied among the three loci, tRNAs being on average three times less divergent than COI and 16S genes. The genus Acanthaster appeared monophyletic, the two species A. brevispinus (Fisher) and A. planci forming distinct clades in agreement with data from morphology and systematics. The A. planci clade split into a West Indian Ocean group and a Pacific group, in agreement with allozyme data on population differentiation. To cite this article: K. Gérard et al., C. R. Biologies 331 (2008).     

Global population structure of the spiny dogfish Squalus acanthias,a temperate shark with an antitropical distribution

The spiny dogfish (Squalus acanthias) is a temperate, coastal squaloid shark with an antitropical distribution in the Atlantic and Pacific oceans. The global population structure of this species is poorly understood, although individuals are known to undergo extensive migrations within coastal waters and across ocean basins. In this study, an analysis of the global population structure of the spiny dogfish was conducted using eight polymorphic nuclear microsatellite markers and a 566‐bp fragment of the mitochondrial ND2 gene region. A low level of genetic divergence was found among collections from the Atlantic and South Pacific basins, whereas a high level of genetic divergence was found among Pacific Ocean collections. Two genetically distinct groups were recovered by both marker classes: one exclusive to North Pacific collections, and one including collections from the South Pacific and Atlantic locations. The strong genetic break across the equatorial Pacific coincides with major regional differences in the life‐history characters of spiny dogfish, suggesting that spiny dogfish in areas on either side of the Pacific equator have been evolving independently for a considerable time. Phylogeographic analyses indicate that spiny dogfish populations had a Pacific origin, and that the North Atlantic was colonized as a result of a recent range expansion from the South American coast. Finally, the available data strongly argue for the taxonomic separation of the North Pacific spiny dogfish from S. acanthias and a re‐evaluation of the specific status of S. acanthias is warranted.     

Isolation of extremophiles with the detection and retrieval of<Emphasis Type="Italic"> Shewanella</Emphasis> strains in deep-sea sediments from the west Pacific

Tests to detect the presence of piezophilic Shewanella strains in the deep-sea sediments of the west, mid- and east Pacific at different depths were done by amplification of previously identified pressure-regulated operons (ORF1,2 and ORF3). The operon fragments were detected in all the deep-sea sediment samples, indicating the broad presence of piezophilic deep-sea Shewanella species or related species in the deep-sea sediments across the Pacific. Extremophiles were isolated from the deep-sea sediment of the west Pacific under atmospheric pressure. Two psychrophilic/psychrotrophic strains, WP2 and WP3, were assigned to the Shewanella genus as determined by their 16S rDNA sequences. WP2 and WP3 were both capable of amplifying pressure-regulated operons; the sequences of the pressure-regulated operons of WP2 and WP3 share high identity between each other, but have more differences from those of S. benthica and S. violacea. The major fatty acids of WP2 and WP3 are 3OH-i-13:0, 14:0, i-15:0, 16:0, 16:1, 18:1, and 20:5. Combined phenotypic analysis, 16S rDNA sequences, and DNA–DNA hybridization results suggest that WP2 and WP3 are two new deep-sea Shewanella species.Communicated by K. Horikoshi     

Molecular characterisation and evolution of the hemocyanin from the European spiny lobster,<Emphasis Type="Italic"> Palinurus elephas</Emphasis>

The hemocyanin of the European spiny lobster Palinurus elephas (synonym: Palinurus vulgaris) is a hexamer composed by four closely related but distinct subunits. We have obtained the full cDNA sequences of all four subunits, which cover 2275-2298 bp and encode for native polypeptides of 656 and 657 amino acids. The P. elephas hemocyanin subunits belong to the alpha-type of crustacean hemocyanins, whereas beta- and gamma-subunits are absent in this species. An unusual high ratio of non-synonymous versus synonymous nucleotide substitutions was observed, suggesting positive selection among subunits. Assuming a constant evolution rate, the P. elephas hemocyanin subunits emerged from a single hemocyanin gene around 25 million years ago. The alpha-type hemocyanins of P. elephas and the American spiny lobster Panulirus interruptus split around 100 million years ago. This is about five times older than the assumed divergence time of the species and suggests that the genera may have split with the formation of the Atlantic Ocean. The application of the Bayesian method for phylogenetic inference allows for the first time a solid reconstruction of the evolution of the decapod hemocyanins, showing that the beta-subunit types diverged first and that the crustacean pseudo-hemocyanins are associated with the gamma-type subunits.     

Genetic Structure of Marchalina hellenica (Hemiptera: Margarodidae) Populations from Turkey: Preliminary mtDNA Sequencing Data

The scale insect Marchalina hellenica (Gennadius) (Hemiptera: Margarodidae) contributes to the production of pine honey in Turkey and Greece via the honeydew excreted when it feeds on pine trees. Although it is an insect of prime economic importance, there is no information on its genetic structure. Preliminary data were obtained based on sequencing analysis of 12s rDNA and COI mtDNA gene segments from samples from four areas of Turkey. Sequences of the 12s rDNA gene segment from Greek samples available in GenBank were also included. No variability was detected concerning the COI mtDNA gene segment analysis, although 13 haplotypes were revealed based on the 12s rDNA gene segment. The most distant population was from Mudanya-Bursa Province (Turkey). Further research is necessary on the genetic structure and variability of M. hellenica populations from the two neighboring countries.     

基于线粒体COI基因序列探讨泥蚶的遗传分化

采用PCR技术对我国沿海地区7个泥蚶群体的线粒体COI基因部分序列进行了测定和遗传分析。在来自7个群体的38个泥蚶样本均得到660 bp的COI基因片段序列,共103个多态位点,组成17种单倍型;数据分析表明：7个群体形成了二大类群：福建以北(包括福建)的5个群体（江苏盐城、浙江奉化、浙江乐清养殖和自然群体、福建福鼎）形成一个类群,类群内的遗传距离为0.0016;福建以南的类群（广东湛江、海南海口）形成一个类群,遗传距离为0.0006;二个类群之间的遗传距离为0.1529,表现为高度的分化。因此我国沿海泥蚶已分化形成福建以南和以北二大类群,二大类群之间的遗传分化已达到亚种水平。     

Genetic diversity and population connectivity of the Asian green mussel Perna viridis in South China Sea,inferred from mitochondria DNA markers

The Asian green mussel Perna viridis is ecologically and economically important in the coastal regions of China. In order to characterize the genetic diversity and population connectivity of P. viridis in South China Sea, a 664 bp region of mitochondrial COI gene and a 293 bp region of 16S rRNA gene were sequenced and analyzed for 78 and 92 individuals from four populations in South China Sea, respectively. A total of 15 haplotypes were defined by 14 variable nucleotide sites in COI gene, and 7 haplotypes by 6 variable nucleotide sites in 16S rRNA gene. High haplotype diversity and low nucleotide diversity were observed in COI gene, while moderate haplotype diversity and low nucleotide diversity were observed in 16S rRNA gene. Pairwise F_ST values of COI gene were all negative and those of 16S rRNA gene ranged from −0.01409 to 0.10289. The results showed that no significant genetic divergence (or shallow genetic structure) and high levels of population connectivity among the four populations of P. viridis in South China Sea.     

基于线粒体COI基因的海蜇不同地理群体遗传特征

对采自辽东湾和韩国江华岛的2个海蜇群体56个个体的线粒体COI基因序列进行扩增,并结合GenBank上其他15个海蜇样品的同源序列对其进行序列变异分析,研究海蜇群体的遗传特征和群体遗传结构状况.在所分析的71个个体中共检测到28个多态位点,定义了32种单倍型.海蜇群体呈现出高的单倍型多样性(0.91±0.06～0.94±0.01)和中等或较低的核苷酸多样性[(0.60±0.34)%～(0.68±0.40)%].与其他几种大型水母相比,海蜇群体的遗传多样性水平较高.系统分析结果显示,海蜇群体间具有2个明显的单倍型谱系分支.比较海蜇不同群体间的遗传分化指数(F_ST)和分子变异分析(AMOVA)发现,在本研究取样范围内不同海蜇群体间存在明显的遗传结构.不同海蜇群体存在复杂的遗传模式,海蜇独特的生活史特征、其分布海域的海流状况及人为因素如增殖放流等可能是造成海蜇这种复杂遗传模式的原因.