Polymorphism of white muscle myosin and parvalbumins in the genus Barbus (Teleostei: Cyprinidae)

Muscle proteins were investigated in two large European barbels, Barbus barbus and B. meridionalis , and in four small tropical barbels native to SE Asia: B. conchonius , B. tetrazona , B. sachsi and B. titteya . Polyacrylamide gel electrophoresis was used to analyse myosin heavy and light chains and parvalbumin isotypes from white trunk muscle. Each species could be biochemically identified. The myosin subunit and parvalbumin isotype patterns obtained for the two European barbels were similar. The Asian barbels, on the other hand, not only differed from the European species but displayed a greater diversity within their group. These biochemical results are largely in agreement with morphological and genetic data, but fail to substantiate suggested close relationships between Asian barbel species.     

Microsatellite variation and genetic structure of brook trout (Salvelinus fontinalis) populations in Labrador and neighboring Atlantic Canada: evidence for ongoing gene flow and dual routes of post-Wisconsinan colonization

In conservation genetics and management, it is important to understand the contribution of historical and contemporary processes to geographic patterns of genetic structure in order to characterize and preserve diversity. As part of a 10-year monitoring program by the Government of Newfoundland and Labrador, Canada, we measured the population genetic structure of the world's most northern native populations of brook trout (Salvelinus fontinalis) in Labrador to gather baseline data to facilitate monitoring of future impacts of the recently opened Trans-Labrador Highway. Six-locus microsatellite profiles were obtained from 1130 fish representing 32 populations from six local regions. Genetic diversity in brook trout populations in Labrador (average H(E)= 0.620) is within the spectrum of variability found in other brook trout across their northeastern range, with limited ongoing gene flow occurring between populations (average pairwise F(ST)= 0.139). Evidence for some contribution of historical processes shaping genetic structure was inferred from an isolation-by-distance analysis, while dual routes of post-Wisconsinan recolonization were indicated by STRUCTURE analysis: K= 2 was the most likely number of genetic groups, revealing a separation between northern and west-central Labrador from all remaining populations. Our results represent the first data from the nuclear genome of brook trout in Labrador and emphasize the usefulness of microsatellite data for revealing the extent to which genetic structure is shaped by both historical and contemporary processes.     

Strong genetic differentiation and postglacial origin of populations in the marine midge Clunio marinus (Chironomidae,Diptera)

The marine midge Clunio marinus (Chironomidae, Diptera) is characterized by a one‐dimensional distribution along the European Atlantic coast, where its lunar and circadian emergence rhythms are genetically adapted to the local tidal regimes, resulting in a series of ‘temporal races’. Clunio marinus is restricted to rocky coasts and thus the temporal races occur in different rocky patches. We studied 10 populations of Clunio marinus from five different regions, spanning the major rocky mainland coasts from Spain to Norway, using amplified fragment length polymorphisms (AFLP), microsatellites and mitochondrial cytochrome oxidase I (COI) sequences. Star‐like patterns of COI haplotypes within regions indicate postglacial colonization. A high degree of shared polymorphisms in AFLP markers suggests colonization from a single source, implying postglacial evolution of timing adaptations in relation to the local tidal regime. In contrast, no COI haplotypes are shared among regions. We hypothesize that different levels of differentiation of nuclear vs. mitochondrial markers in the source region were carried forward during postglacial expansion. Despite the recent origin of populations, all markers reveal distinct genetic differentiation between rocky coasts on a scale of 650–1150 km. Differentiation between rocky coasts is not correlated to timing adaptations, suggesting that geographic isolation is prevalent between rocky coasts and that this facilitated the evolution of local timing adaptations. At the same time there is little genetic differentiation within rocky coasts on a scale of 2–6 km; leaving open the possibility that within rocky coasts with large variation in tidal regimes, temporal adaptations evolved in the face of gene flow.     

Heterogeneous colonization pattern of European Cyprinids, as highlighted by the dace complex (Teleostei: Cyprinidae)

The dace (Leuciscus leuciscus), with a very large geographic distribution all over Europe, represents an interesting species model for studies of the global mechanisms underlying aquatic system biodiversity. To assess the congruence with the past colonization process hypothesis of the freshwater fauna in Western Europe, we investigated the evolutionary history of this species, by integrating morphological variation (eight meristic characters), mitochondrial (cytochrome b, 16S rDNA and control region, over a total of 2169 bp) and nuclear (12 allozymes loci) phylogenetic relationships, and investigating population dynamics via expansion, migration, bottleneck, and divergence time analyses. We carried out nested clade phylogeographic analysis for a total of 663 specimens from 31 populations taken from all over the distribution area. Unlike previous studies, we found that L. leuciscus is currently constituted by five lineages belonging to two clades (yielding 6.3% of pairwise divergence). The relationships between these lineages were accounted for by complex biogeographical patterns due to Pliocene and Pleistocene paleoclimatic events, validating the identification of new glacial refuges for freshwater fish in Western Europe. Finally, we demonstrated hybridization between L. leuciscus and Leuciscus idus.     

New insights on postglacial colonization in western Europe: the phylogeography of the Leisler's bat (Nyctalus leisleri)

Despite recent advances in the understanding of the interplay between a dynamic physical environment and phylogeography in Europe, the origins of contemporary Irish biota remain uncertain. Current thinking is that Ireland was colonized post-glacially from southern European refugia, following the end of the last glacial maximum (LGM), some 20 000 years BP. The Leisler''s bat (Nyctalus leisleri), one of the few native Irish mammal species, is widely distributed throughout Europe but, with the exception of Ireland, is generally rare and considered vulnerable. We investigate the origins and phylogeographic relationships of Irish populations in relation to those across Europe, including the closely related species N. azoreum. We use a combination of approaches, including mitochondrial and nuclear DNA markers, in addition to approximate Bayesian computation and palaeo-climatic species distribution modelling. Molecular analyses revealed two distinct and diverse European mitochondrial DNA lineages, which probably diverged in separate glacial refugia. A western lineage, restricted to Ireland, Britain and the Azores, comprises Irish and British N. leisleri and N. azoreum specimens; an eastern lineage is distributed throughout mainland Europe. Palaeo-climatic projections indicate suitable habitats during the LGM, including known glacial refugia, in addition to potential novel cryptic refugia along the western fringe of Europe. These results may be applicable to populations of many species.     

Variability in the number of tooth rows in the pharyngeal dentition of Barbus intermedius (Teleostei; Cyprinidae): genetic,hormonal and environmental factors

The variability of pharyngeal dentition in a natural population of B. intermedius and effects of genetic, hormonal and environmental factors on the number of tooth rows in the pharyngeal dentition in offspring from wild‐caught parents have been investigated. It was revealed that: (i) about 10% of fish from natural population have four‐rowed dentition instead of three‐rowed dentition characteristic for this species; (ii) the presence of the additional tooth row is not an abnormality of tooth replacement since it occurs symmetrically on both sides; (iii) occurrence of the fourth row of teeth is heritable since laboratory‐reared offspring from parents with four‐rowed dentition have the same dentition. Even if one of the parents had four‐rowed dentition the percentage of four‐rowed individuals in the progeny was significantly higher than in progeny from parents with normal (three rowed) dentition; (iv) the number of tooth rows appears to be hormonally controlled: high levels of thyroid hormone result in a decrease in the number of tooth rows to two. In contrast, deficiency of this hormone results in an increase to four rows; (v) no differences in the number of tooth rows were found in fish reared under 17°C, 24°C, 30°C and room temperature (20–26°C).     

Isolation and characterization of polymorphic microsatellite loci for the dace complex: Leuciscus leuciscus (Teleostei: Cyprinidae)

Ten novel polymorphic microsatellites were isolated from the dace complex (Leuciscus leuciscus), which is a European cyprinid species. Four multiplex polymerase chain reaction sets were optimized in order to genotype 26 polymorphic loci in all, including 16 previously published cyprinid-specific loci. The level of genetic diversity was assessed in 142 dace individuals. We also successfully applied 26 of the microsatellites to 10 related species. These primers thus will be useful to assess population structure of the dace and other cyprinid species, with application for conservation issues and phylogeographical approaches.     

Isolation and characterization of polymorphic microsatellite loci in the freshwater fishes Telestes souffia and Telestes muticellus (Teleostei: Cyprinidae)

Ten novel polymorphic microsatellites (seven with perfect motifs) were isolated from vairone species (Telestes souffia and Telestes muticellus), which are endangered European cyprinid species. Together with 11 previously published cyprinid-specific loci, five multiplex sets were optimized, allowing the genotyping of 21 polymorphic loci. The level of genetic diversity was assessed in 97 individuals from the two species T. souffia and T. muticellus. We also successfully applied the 21 microsatellites to nine related species. These primers will thus be useful in assessing population structure of the vairone and other cyprinid species, with application for conservation issues and phylogeographical approaches.     

Conservation genetic studies of the endangered Cape Fear Shiner, Notropis mekistocholas (Teleostei: Cyprinidae)

Genetic variation at ten microsatellite lociand one anonymous-nuclear locus was assayed forthree geographic samples of the criticallyendangered North American cyprinidNotropis mekistocholas (Cape Fear shiner). Despite low abundance of this species, therewas little suggestion of small populationeffects; allele diversity and heterozygositywere relatively high, F_IS values withinsamples were non-significant, and genotypeswere distributed in frequencies according toHardy-Weinberg expectations. Geneticdivergence among samples was minimal despitethe presence of dams, constructed in the early1900s, that separate the sample sites. Thissuggests that recent gene flow has beensufficient to inhibit genetic divergence orthat gene flow has been reduced but there hasbeen insufficient time for genetic divergenceto develop. Tests of heterozygosity excesswere non-significant, suggesting that N.mekistocholas in the localities sampledhave not undergone recent reductions ineffective population size. Future studiesemploying larger sample sizes to provide morerobust tests of population structure andtemporally separated samples to estimatecontemporaneous N_e are warranted.     

Levels of genetic variability in sculpins (Gottidae: Teleostei) of the North American Pacific coast and an assessment of potential correlates

We examined levels of allozymic variation in intertidal sculpins of the genera Oligocottus, Clinocottus and Artedius. Variability in these species, as assessed by the proportion of loci polymorphic and average heterozygosity per locus, was generally low but within the range of values observed for other fish groups. In reviewing the ecological and life-history attributes of these sculpins, we found only one outstanding correlate of allozyme variability–larger-bodied species tended to have lower levels of variation. Also, the level of heterozygosity was not related in any obvious way to historical/ evolutionary aspects such as position in phylogenetic trees or the inferred relative ages of species. Our study of a relatively well-defined lineage of fishes that co-occur in the same geographical region and in the same, or broadly similar, habitats, underscores the difficulty of detecting correlations between genetic variability and seemingly relevant ecological features of species.     

Differentiation and levels of genetic variation in northern European lynx (Lynx lynx) populations revealed by microsatellites and mitochondrial DNA analysis

The European lynx (Lynx lynx) hasexperienced significant decline in populationnumbers over large parts of its formerdistribution area in central and northernEurope. In Scandinavia (Sweden and Norway), thespecies has been subject to intense hunting and inthe early 20th century the population size mayhave been as low as about 100 animals. Duringthe rest of the century there have beenalternating periods of restricted hunting andtotal protection. Future management of theScandinavian lynx population will requireinsight into what effects demographicbottlenecks may have had on genetic variabilityand structure. For this purpose, 276 lynxesfrom Sweden, Norway, Finland, Estonia andLatvia were analysed for polymorphism at 11feline microsatellite loci and at themitochondrial DNA (mtDNA) control region.Scandinavian lynxes were found to be fixed fora single mtDNA haplotype, while this and threeadditional haplotypes were seen in Finland andthe Baltic States (Estonia and Latvia); thehaplotypes were all very similar, onlydiffering at 1–4 sites within a 700 bp regionsequenced. Microsatellite variability wasmoderate (H_e = 0.51–0.62) with lowerheterozygosity and fewer alleles in Scandinaviathan in Finland and the Baltic States together,though significant so only for the latter.Heterozygosity data in Scandinavia wereconsistent with a recent population bottleneck.Various analyses (e.g. F_st, individual-basedtree, assignment test) revealed distinctgenetic differentiation between Scandinavianlynxes and animals from Finland and the BalticStates. Some structure was evident withinScandinavia as well, suggesting an isolation bydistance. The observed partition of geneticvariability between Scandinavia and the easterncountries thereof indicates that lynxpopulations from the two regions may need to beseparately managed. We discuss what factors canhave contributed to the population geneticstructure seen in northern European lynxpopulations of today.     

A re‐examination of the molecular phylogeny and biogeography of the genus Schizothorax (Teleostei: Cyprinidae) through enhanced sampling,with emphasis on the species in the Yunnan–Guizhou Plateau,China

The mitochondrial DNA cytochrome b sequences of 36 Schizothorax species from 51 localities in the Yunnan–Guizhou Plateau (YGP) and its adjacent areas were analysed. Maximum parsimony, Maximum likelihood and Bayesian phylogenetic analyses were performed to examine the relationships of Schizothorax species. A hypothesis of the phylogenetic relationships of the species is given. A relaxed molecular clock based on Bayesian evolutionary analysis was used to tentatively calculate the divergence times of Schizothorax. Samples from the YGP were tentatively grouped into three geographically distributed clades: the Tsangpo‐Irrawaddy, the Mekong‐Salween and the Trans‐Jinsha River (including Jinsha, Red, Nanpan and Beipan Rivers). Calibration of the molecular clock revealed that two geological periods, the late Miocene about 10 million years before present (Myr BP) and the Pliocene (4.0 Myr BP), were important times in the vicariant speciation of Schizothorax. The phylogenetic history of the species is congruent with events caused by the uplift of the Tibetan Plateau and the YGP. The divergence of Schizothorax species in YGP began in the Pliocene. Our phylogenetic trees did not support the hypothesis that the paleo Jinsha River was drained through the Yangtze River‐Jianchuan Lake‐Erhai Lake to the Red River. Schizothorax in the Beipan River were derived from the Jinsha River.     

A polymorphic microsatellite from the Squalius alburnoides complex (Osteichthyes, Cyprinidae) cloned by serendipity can be useful in genetic analysis of polyploids

A new microsatellite locus (SAS1) for Squalius alburnoides was obtained through cloning by serendipity. The possible usefulness of this new species-specific microsatellite in genetic studies of this hybrid-species complex, was explored. The polymorphism exhibited by SAS1 microsatellite is an important addition to the set of microsatellites previously used in genetic studies in S. alburnoides complex, that mostly relied in markers described for other species. Moreover, the SAS1 microsatellite could be used to identify the parental genomes of the complex, complementing other methods recently described for the same purpose..     

Geographic pattern of genetic variation in the European globeflower Trollius europaeus L. (Ranunculaceae) inferred from amplified fragment length polymorphism markers

The distribution of genetic variation and the phylogenetic relationships between 18 populations of the arctic-alpine plant Trollius europaeus were analysed in three main regions (Alps, Pyrenees and Fennoscandia) by using dominant AFLP markers. Analysis of molecular variance revealed that most of the genetic variability was found within populations (64%), although variation among regions (17%) and among populations within regions (19%) was highly significant (P < 0.001). Accordingly, the global fixation index FST averaged over loci was high (0.39). The among-population differentiation indicates restricted gene flow, congruent with limited dispersal of specific globeflower's pollinating flies (Chiastocheta spp.). Within-population diversity levels were significantly higher in the Alps (mean Nei's expected heterozygosity HE = 0.229) than in the Pyrenees (HE= 0.197) or in Fennoscandia (HE = 0.158). This finding is congruent with the species-richness of the associated flies, which is maximum in the Alps. We discuss the processes involved in shaping observed patterns of genetic diversity within and among T. europaeus populations. Genetic drift is the major factor acting on the small Pyrenean populations at the southern edge of T. europaeus distribution, while large Fennoscandian populations result probably from a founder effect followed by demographic expansion. The Alpine populations represent moderately fragmented relics of large southern ancestral populations. The patterns of genetic variability observed in the host plant support the hypothesis of sympatric speciation in associated flies, rather than recurrent allopatric speciations.     

Population genetic structure of the lemon shark (Negaprion brevirostris) in the western Atlantic: DNA microsatellite variation

DNA microsatellite markers were used to characterize the population genetic structure of the lemon shark, Negaprion brevirostris, in the western Atlantic. This study demonstrates for the first time the usefulness of microsatellites to study population genetic structure and mating systems in the Chondricthyes. Lemon sharks (mostly juveniles) were sampled non-destructively from four locations, Gullivan Bay and Marquesas Key in Florida, Bimini, Bahamas, and Atol das Rocas, Brazil. At least 545 individuals were genotyped at each of four dinucleotide loci. The number of alleles per locus ranged from 19 to 43, and expected heterozygosities ranged from 0.69 to 0.90. Relatively little genetic structure was found in the western Atlantic, with small but significant values for estimators of F(ST) and R(ST) among populations, theta (0.016) and rho (0.026), respectively. No sharp discontinuities were found between the Caribbean sites and Brazil, and most alleles were found at all four sites, indicating that gene flow occurs throughout the western Atlantic with no evidence for distinct stocks.     

Habitat-dependent geographical variation in ontogenetic allometry of the shiner perch Cymatogaster aggregata Gibbons (Teleostei: Embiotocidae)

Studies of intraspecific morphological variation in fishes have traditionally focused on freshwater rather than marine species. In addition, such studies typically focus on adults, although causes and intensities of selective pressures most likely vary through an individual's lifetime. In this study, body and head shape of a marine species, shiner perch Cymatogaster aggregata Gibbons were compared among localities along the Pacific Northwest coast of North America. Evidence was found for intraspecific variation in ontogenetic allometry, and for a closer correlation of body shape with environment rather than geographical proximity. This correlation with environment was more evident in younger fish, thereby demonstrating the importance of analysing multiple life stages. A common garden experiment suggests both environmental and genetic bases for the observed differences. Recognizing intraspecific ecomorphological complexity and its specificity to habitat and/or life stage can have important consequences for understanding the role of local adaptation and population dynamics in macroecology.     

The effects of genetic drift during range expansion on geographical patterns of variation: a computer simulation of the colonization of Australia by Bufo marinus

A computer simulation is performed of allele frequency changes resulting from genetic drift at eleven loci during the probable course of colonization of Australia by populations of the Giant Toad, Bufo marinus. The history of twelve populations for which allele frequency data are available is modelled. Account is taken of the likely pattern of relationship among the populations, the effective size of the populations (as indicated by the observed variance of allele frequencies) and the probable isolation of the populations from each other following their separation. In all simulations, allele frequencies at some loci show a significant association with latitude while others do not. In six of ten simulations, there is a significant association between degree of variation at a locus and the presence of a latitudinal cline of allele frequencies. There are also indications of this kind of association in simulations of a uni-directional range expansion. These results demonstrate that such associations, which were also observed in the data from the actual populations, can result from genetic drift during a range expansion, and therefore cannot be taken as evidence of the action of natural selection.     

Substantial loss of genetic variation in a single generation of Senegalese sole (Solea senegalensis) culture: implications in the domestication process

The flatfish Solea senegalensis represents an important resource in modern mariculture and is largely raised in South Spain and Portugal. Substantial progress has been achieved in its domestication, though suitable reproduction and zootechnical conditions still remain unknown. Difficulties to obtain breeders from wild lead many companies to set up broodstocks with first generation (G1) progeny without genetic guidelines. To study the genetic processes underlying the early stage of domestication of this species, the genetic structure of four representative broodstocks from Southern Spain has been assessed by means of eight microsatellite loci . Data revealed a substantial reduction in levels of genetic variability on just one generation in stocks totally or partially composed of G1 individuals when compared with those solely integrated by individuals from wild. The genetic relatedness within the four stocks has been established, having detected close relationships between individuals from cultured origin, thus suggesting that the loss of variability is apparently due to setting up broodstocks by blind selecting of family related G1 individuals. The high proportion of siblings in these stocks can have negative consequences in future generations due to inbreeding effects. The relationships between the four broodstocks were traced, having found a common origin between two of them, which come from the same donor hatchery, thus enhancing the homogenization of the existent resources for S. senegalensis among the various stocks. This finding alerts about the risks in exchanging fish between hatcheries without knowing their genetic origin. From this study, the use of microsatellites is strongly recommended to control the genetic composition of S. senegalensis broodstocks, aimed to maintain standards of genetic health and improve their reproduction capacity, which is a key issue in the domestication process.     

Genetic diversification and population structure of <Emphasis Type="Italic">Barbus cyri</Emphasis> De Filippi, 1865 (Teleostei: Cyprinidae) in northern Iran inferred from the mitochondrial D-loop gene sequence

A genetic survey of Barbus cyri populations from two biogeographical endorheic basins (Caspian Sea and Urmia Lake) was carried out using a mitochondrial marker (partial D-loop) in order to ascertain intra- and inter-population genetic diversity, population demography and to address their genetic structure which is the key to conservation action planning. Analyses were conducted on sequences obtained from 68 individuals collected from 10 sampling sites, from two basins. By means of morphological characteristics all specimens collected from the Caspian Sea basin were ascribed to Barbus cyri. Genetic diversity values (h and π) of sampling groups were all different from 0 (in Babolrud River population) to 0.857 (in Kalibar River population). Population connectivity and colonization patterns of the studied area were inferred from an analysis of molecular variance distribution and evolutionary relationships among haplotypes. The results point to different levels of isolation among sampling groups due to ecological and anthropogenic factors and the effect of an artificial barrier on genetic variability and conservation status of the population. Finally, this study confirms the uncertainty associated with systematic classification of Barbus spp. based on morphological characters due to the phenotypic plasticity of the species.