The use and limits of ITS data in the analysis of intraspecific variation in Passiflora L. (Passifloraceae)

The discovery and characterization of informative intraspecific genetic markers is fundamental for evolutionary and conservation genetics studies. Here, we used nuclear ribosomal ITS sequences to access intraspecific genetic diversity in 23 species of the genus Passiflora L. Some degree of variation was detected in 21 of these. The Passiflora and Decaloba (DC.) Rchb. subgenera showed significant differences in the sizes of the two ITS regions and in GC content, which can be related to reproductive characteristics of species in these subgenera. Furthermore, clear geographical patterns in the spatial distribution of sequence types were identified in six species. The results indicate that ITS may be a useful tool for the evaluation of intraspecific genetic variation in Passiflora.     

Extensive intraspecific chloroplast DNA (cpDNA) variation in the alpine Draba aizoides L. (Brassicaceae): haplotype relationships and population structure

Chloroplast DNA (cpDNA) sequence variation is currently the most widely used tool for the inference of phylogenetic relationships among plants at all taxonomic levels. Generally, noncoding regions tend to evolve faster than coding sequences and have recently been applied to the study of phylogenetic relationships among closely related taxa. An implicit assumption of many of these studies is that intraspecific cpDNA variation is either absent or low and therefore will not interfere with the reconstruction of interspecific relationships. A survey of cpDNA sequence variation in the common alpine plant species Draba aizoides L. was undertaken to assess levels of intraspecific cpDNA sequence variation. These levels were compared to levels of interspecific sequence divergence between D. aizoides and related alpine Draba species. Intraspecific cpDNA sequence divergence was extensive in D. aizoides, and intraspecific differences were often larger than interspecific differences. cpDNA haplotype relationships were explored using a maximum parsimony approach and minimum-spanning networks. Results from both methods were largely congruent but comparisons provided interesting insights into the presumed evolutionary history of cpDNA haplotypes. A combined effect of cpDNA introgression and complex lineage sorting was inferred to explain the pattern of cpDNA variation found in D. aizoides. Our results suggest that intraspecific cpDNA variation can be extensive and that intraspecific variation needs to be taken into account when inferring phylogenetic relationships among closely related taxa.     

New microsatellite loci for the European bushcricket,Ephippiger ephippiger (Orthoptera: Tettigoniidae)

Ephippiger ephippiger is a model organism for studies of sexual selection and phylogeography but little is known about fine‐scale population structure. Available microsatellite loci have null allele problems so we used an enrichment technique to isolate 21 new microsatellite loci for E. ephippiger. We present primer pairs for 10 polymorphic loci (3–11 alleles per locus). Observed heterozygosities at polymorphic loci ranged from 0.118 to 0.787, but several were significantly lower than expected.     

Phylogeographic histories of representative herpetofauna of the southwestern U.S.: mitochondrial DNA variation in the desert iguana (Dipsosaurus dorsalis) and the chuckwalla (Sauromalus obesus)

To determine whether genetic variation in representative reptiles of the southwestern U.S. may have been similarly molded by the geologic history of the lower Colorado River, we examined restriction site polymorphisms in the mitochondrial DNA (mtDNA) of desert iguanas (Dipsosaurus dorsalis) and chuckwallas (Sauromalus obesus). Observed phylogeographic structure in these lizards was compared to that reported for the desert tortoise (Xerobates agassizi), whose mtDNA phylogeny demonstrates a striking genetic break at the Colorado River. Both the desert iguana and chuckwalla exhibit extensive mtDNA polymorphism, with respective genotypic diversities G = 0.963 and 0.983, close to the maximum possible value of 1.0. Individual mtDNA clones, as well as clonal assemblages defined by specific levels of genetic divergence, showed pronounced geographic localization. Nonetheless, for each species the distributions of certain clones and most major clonal groupings encompass both sides of the Colorado River valley, and hence are clearly incongruent with the phylogeographic pattern of the desert tortoise. Overall, available molecular evidence provides no indication that the intraspecific phylogenies of the southwestern U.S. herpetofauna have been concordantly shaped by a singular vicariant factor of overriding significance.     

Characterization of microsatellite markers for Couch's spadefoot toad (Scaphiopus couchii) and cross‐amplification in other species of the family Scaphiopodidae

I developed 12 di‐ and tetranucleotide microsatellite loci for Couch's spadefoot toad (Scaphiopus couchii). These loci have 3–37 alleles per locus and observed heterozygosities ranging from 0.157 to 0.941 among 85 individuals from four populations. Global and within‐population exact tests do not reveal departure from Hardy–Weinberg expectations and all loci pairs are in linkage equilibrium. These independent markers will be useful for studies of population structure and kinship in this commonly studied amphibian. Additionally, several of these loci may be applicable for studies of other North American toads of the family Scaphiopodidae.     

Variable microsatellite markers for a snapping shrimp (Alpheus armillatus) species complex

Snapping shrimp (Alpheus) are an important ecological component of tropical marine systems and are excellent models for studies of phylogeography and behavioural ecology. Here, I describe 10 variable microsatellite loci for the Alpheus armillatus species complex of snapping shrimp. The primers were tested on 60 individuals representing three evolutionary lineages within the complex, and yielded three to 21 alleles. These loci will be effective tools in future studies on population genetics and mating behaviour in this complex.     

Universal primers for the amplification of chloroplast microsatellites in grasses (Poaceae)

Attempts to design truly universal primers to amplify chloroplast microsatellites have met with limited success due to nonconservation of repeat loci across widely divergent taxa. We have used the complete chloroplast genome sequences of rice, maize and wheat to design five pairs of primers that amplify homologous mononucleotide repeats across the Poaceae (grasses). Sequencing confirmed conservation of repeat motifs across subfamilies and a preliminary study in Anthoxanthum odoratum revealed polymorphism at two loci with a haplotype diversity value of 0.495. These primers provide a valuable tool to study cytoplasmic diversity in this extensively studied and economically important range of taxa.     

A comparison of mtDNA restriction sites vs. control region sequences in phylogeographic assessment of the musk turtle (Sternotherus minor)

A total of nearly 800 base pairs of mitochondrial DNA sequence was assayed in each of 52 musk turtles (Sternotherus minor) collected across the species' range in the south-eastern USA. About one-half of the sequence information in effect was accessed by conventional recognition-site assays of the entire mtDNA molecule; the remainder came from direct sequence assays of a normally hypervariable 5′ section of the noncoding control region. The two assay methods produced essentially nonoverlapping sets of variable character states that were compared with respect to magnitudes and phylogeographic patterns of mtDNA variation. The two assay procedures yielded nearly identical outcomes with regard to: (a) total levels of species-wide mtDNA genetic variation; (b) mean levels of within-locale variation; (c) extremely high population genetic structure; (d) a phylogenetcally significant separation of samples from the north-western half of the species' range vs. those in the south-eastern segment; and (e) considerably lower genetic variability within the north-western clade. The micro- and macro-phylogeographic mtDNA patterns in the musk turtle are consistent with a low-dispersal natural history, and with a suspected longer-term biogeographic history of the species, respectively.     

Three species of Pagodia (Corynexochida,Trilobita) from the Furongian (Cambrian) of northern Anhui,China and their intraspecific variation

More than one thousand cranidia, librigenae, and pygidia of the trilobite Pagodia were collected from the Cambrian (upper Jiangshanian through Stage 10) at Fenghuangshan in northern Anhui, China. They were identified as Pagodia lotos Walcott, 1905, Pagodia depressa (Walcott, 1905), and Pagodia bia Walcott, 1905. Based on one rare nearly complete exoskeleton of P. depressa and comparisons among the three species of Pagodia, the redefinition for those species is attempted as follows. Pagodia lotos: Glabella rectangular, slightly convergent anteriorly; anterior border ridge-like, anterior border furrow deep; pygidium semicircular, axis tapered rearward, with 4 axial rings and a terminal piece. P. depressa: Cranidium subquadrate, highly convex; glabella cylindrical, convergent anteriorly; pygidium semicircular. P. bia: Glabella rectangular, constricted in the midlength; fixigena wide (tr.); anterior border very narrow (sag.) and ridge-like; pygidium semicircular with wide borders; pygidial axis ended closer to the pygidial border furrow. We also discuss their intraspecific variations, commonly seen on the exfoliated specimens: palpebral ridge evident or obliterated, the length (sag.) of anterior area variable, the decoration of exoskeleton surface (with granules variable in size, convex or pitted).     

Genetic variation and origin of red turpentine beetle (Dendroctonus valens LeConte) introduced to the People's Republic of China

Abstract 1 The red turpentine beetle, Dendroctonus valens LeConte, is a recent New World introduction to the People's Republic of China. An outbreak of these beetles has infested over 0.5 million hectares of pine forests. 2 Efforts are underway to suppress this outbreak using biological control measures. However, the wide distribution in the native range of D. valens suggests regional variation of the beetle's biology, predators, and parasitoids. Thus, knowledge of the origin of these beetles can help devise precise and effective control measures. 3 A portion of the mitochondrial cytochrome oxidase subunit I gene was sequenced for 218 individuals from 32 populations throughout the native range of D. valens and in China. 4 Haplotype diversity was high. A total of 131 haplotypes were found and Jukes–Cantor corrected nucleotide difference ranged from 0 to 16%. Haplotype diversity ranged from 0.53 to 0.98 and unique haplotypes were found in most populations. 5 Parsimony and statistical parsimony analyses of these haplotypes support the hypothesis that the introduction of D. valens to China was recent and originated from the Pacific North‐west of the U.S.A. 6 In addition, the high haplotype diversity also suggests a large or multiple introductions. However, based on the genetics of the beetle's reproductive behaviour, this diversity may also be explained by a limited number of individuals or introductions.     

Fungal specific primers for PCR‐amplification of mitochondrial LSU in lichens

Fungal specific primer sequences for the amplification of the large subunit of the mitochondrial ribosomal DNA (mtLSU) are presented in this paper. Fungal specific primers make the separation of fungal and algal cells prior to DNA‐extraction from lichens unnecessary. This is especially useful in crustose and small foliose and fruticose lichens. An example from a complex of closely related species of the crustose lichen genus Biatora shows the usefulness of mtLSU‐sequences for studies of infraspecific variability and lower level systematics of lichenized ascomycetes.     

Plant intraspecific functional trait variation is related to within‐habitat heterogeneity and genetic diversity in Trifolium montanum L.

Intraspecific trait variation (ITV), based on available genetic diversity, is one of the major means plant populations can respond to environmental variability. The study of functional trait variation and diversity has become popular in ecological research, for example, as a proxy for plant performance influencing fitness. Up to now, it is unclear which aspects of intraspecific functional trait variation (iFD_CV) can be attributed to the environment or genetics under natural conditions. Here, we examined 260 individuals from 13 locations of the rare (semi‐)dry calcareous grassland species Trifolium montanum L. in terms of iFD_CV, within‐habitat heterogeneity, and genetic diversity. The iFD_CV was assessed by measuring functional traits (releasing height, biomass, leaf area, specific leaf area, leaf dry matter content, F_v/F_m, performance index, stomatal pore surface, and stomatal pore area index). Abiotic within‐habitat heterogeneity was derived from altitude, slope exposure, slope, leaf area index, soil depth, and further soil factors. Based on microsatellites, we calculated expected heterozygosity (H_e) because it best‐explained, among other indices, iFD_CV. We performed multiple linear regression models quantifying relationships among iFD_CV, abiotic within‐habitat heterogeneity and genetic diversity, and also between separate functional traits and abiotic within‐habitat heterogeneity or genetic diversity. We found that abiotic within‐habitat heterogeneity influenced iFD_CV twice as strong compared to genetic diversity. Both aspects together explained 77% of variation in iFD_CV ( = .77, F_{2, 10} = 21.66, p < .001). The majority of functional traits (releasing height, biomass, specific leaf area, leaf dry matter content, F_v/F_m, and performance index) were related to abiotic habitat conditions indicating responses to environmental heterogeneity. In contrast, only morphology‐related functional traits (releasing height, biomass, and leaf area) were related to genetics. Our results suggest that both within‐habitat heterogeneity and genetic diversity affect iFD_CV and are thus crucial to consider when aiming to understand or predict changes of plant species performance under changing environmental conditions.     

Landscape models for nuclear genetic diversity and genetic structure in white-footed mice (Peromyscus leucopus)

Dramatic changes in the North American landscape over the last 12 000 years haveshaped the genomes of the small mammals, such as the white-footed mouse (Peromyscusleucopus), which currently inhabit the region. However, very recent interactions ofpopulations with each other and the environment are expected to leave the most pronouncedsignature on rapidly evolving nuclear microsatellite loci. We analyzed landscapecharacteristics and microsatellite markers of P. leucopus populations along atransect from southern Ohio to northern Michigan, in order to evaluate hypotheses aboutthe spatial distribution of genetic heterogeneity. Genetic diversity increased to thenorth and was best approximated by a single-variable model based on habitat availabilitywithin a 0.5-km radius of trapping sites. Interpopulation differentiation measured byclustering analysis was highly variable and not significantly related to latitude orhabitat availability. Interpopulation differentiation measured as F_STvalues and chord distance was correlated with the proportion of habitat intervening, butwas best explained by agricultural distance and by latitude. The observed gradients indiversity and interpopulation differentiation were consistent with recent habitatavailability being the major constraint on effective population size in this system, andcontradicted the predictions of both the postglacial expansion and core-peripheryhypotheses.     

Divergence in the chloroplast genome and nuclear rDNA of the rare western australian plant lambertia orbifolia gardner (Proteaceae)

The population genetic structure of the Australian plant Lambertia orbifolia was investigated for chloroplast DNA (cpDNA) and rDNA based on restriction fragment length polymorphism. Variation was assessed in 14-20 individuals from six populations with probes covering the majority of the chloroplast genome and the whole rRNA gene unit. For cpDNA, eight mutations were detected which were distributed over five haplotypes. Nucleotide diversity in the species was high and the majority of this diversity was distributed between populations with diversity within populations restricted to a single population. There was significant differentiation between the two regions in the species distribution with the Narrikup region being distinguished by a single haplotype that was characterized by six unique mutations. Variation in rDNA was detected with three gene length variants present in most individuals. However, the Narrikup region was characterized by homogenization of the gene unit to a single length variant in all individuals. The divergence of the Narrikup region suggests that the disjunction in the species distribution has been present for a long time and the two regions represent separate evolutionary lineages.     

Low levels of allozyme variability in the threatened species Antirrhinum subbaeticum and A. pertegasii (Scrophulariaceae): implications for conservation of the species

AIMS: This study was designed to compare levels of genetic variation and its partitioning in three related species of Antirrhinum, A. subbaeticum, A. pertegasii and A. pulverulentum, and to check the hypothesis that species with small total population size have lower levels of genetic variability than those with bigger ones. This information should contribute to the development of conservation strategies of rare endemic species of Antirrhinum. METHODS: One hundred and seventy-seven plants were screened for variability at 14 allozyme loci by means of horizontal starch gel. Parameters of genetic diversity, and its partitioning, were calculated. An indirect estimate of gene flow was based on the equation: Nm = (1 - GST)/4GST. KEY RESULTS: Genetic variabilities in A. subbaeticum and A. pertegasii were found to be the lowest known for the genus, the within-population genetic diversity being correlated with population size in both species. The distribution of genetic diversity is strikingly different among species, with 85 % of the total variation distributed among populations in A. subbaeticum, 6 % in A. pertegasii and 23 % in A. pulverulentum. Estimated levels of gene flow were negligible for A. subbaeticum (0.04), high for A. pertegasii (3.92), and substantial for A. pulverulentum (0.83). Genetic and geographic distances were negatively correlated in A. pertegasii, whereas no significant correlation was found in the other two species. CONCLUSIONS: Levels of total genetic diversity agree with the hypothesis that species with small total population size have lower levels of genetic variability than those with bigger ones. Strategies for the conservation of the species are recommended, such as preservation of natural populations and avoidance of possible causes of threat, as well as ex situ preservation of seeds, reinforcement of small populations of A. subbaeticum with plants or seeds from the same population, and avoidance of translocations among populations.     

Genetic variation within and between Iliamna corei and I. remota (Malvaceae): implications for species delimitation

Iliamna corei and I. remota are classified as endangered species; however, their designation as separate species has been questioned. In order to address this problem, intersimple sequence repeat (ISSR) data were generated to examine patterns of genetic differentiation within and between these two taxa. ISSRs were used to screen individuals for genetic diversity of I. corei from the single known natural population and two garden populations, and individuals of I. remota from the six natural populations and four garden populations. Using ten primers, 140 informative markers were generated. Ninety-four percent of loci detected revealed polymorphisms. Cluster analysis (neighbour-joining, NJ) revealed that the two species are genetically distinct and that the Illinois populations of I. remota were genetically distinct from the Virginia populations of I. remota . Ordination by Principal Coordinate Analysis (PCoA) supported the findings of the NJ, with a separation of I. corei and I. remota . Analysis of Molecular Variance (AMOVA) revealed that the majority of variation detected was within populations, which is consistent with other self-incompatible plants. The results indicate a correlation between the geographical distributions of the species and gene flow.  © 2006 The Linnean Society of London, Botanical Journal of the Linnean Society , 2006, 151 , 345–354.     

Genetic signature of recent glaciation on populations of a near-shore marine fish species (Syngnathus leptorhynchus)

Continental glaciation has played a major role in shaping the present-day phylogeography of freshwater and terrestrial species in the Northern Hemisphere. Recent work suggests that coastal glaciation during ice ages may have also had a significant impact on marine species. The bay pipefish, Syngnathus leptorhynchus , is a near-shore Pacific coast fish species with an exceptionally wide latitudinal distribution, ranging from Bahia Santa Maria, Baja California to Prince William Sound, Alaska. Survey data indicate that S. leptorhynchus is experiencing a range expansion at the northern limit of its range, consistent with colonization from southern populations. The present study uses six novel microsatellite markers and mitochondrial DNA (mtDNA) sequence data to study the present-day population genetic structure of four coastal populations of S. leptorhynchus . Deficits in mtDNA and nuclear DNA diversity in northern populations from regions glaciated during the last glacial maximum (LGM) [ c . 18 000 years before present ( bp )] suggest that these populations were effected by glacial events. Direct estimates of population divergence times derived from both isolation and isolation-with-migration models of evolution are also consistent with a postglacial phylogenetic history of populations north of the LGM. Sequence data further indicate that a population at the southern end of the species range has been separated from the three northern populations since long before the last interglacial event ( c . 130 000 years bp ), suggesting that topographical features along the Pacific coast may maintain population separation in regions unimpacted by coastal glaciation.     

Social kin associations and genetic structuring of striped dolphin populations (Stenella coeruleoalba) in the Mediterranean Sea

We investigated hierarchical patterns of genetic subdivision, and assessed kinship within and between social groups of striped dolphins (Stenella coeruleoalba) in the Tyrrhenian Sea. A total of 165 samples were analysed at eight microsatellite DNA loci, including outgroup samples from the Adriatic, Scotland and Spain for population-level comparisons. We found population genetic structure within the Mediterranean basin, including small but significant differentiation between the Adriatic and Tyrrhenian Seas (FST=0.0047, P=0.008), and between putative 'inshore' and 'offshore' (FST=0.0217, P=0.005) populations in the Tyrrhenian Sea. Assessment of kinship within and among 12 association groups showed higher average kinship for females within than between groups, and smaller groups showed higher average kinship. Comparisons of relatedness for both sexes showed a significant difference between males and females, with females more likely to associate with adult kin. Together these data emphasize the importance of the social cohesion of kin in small groups to the structuring of striped dolphin populations in this environment.     

Isolation and characterization of microsatellite loci in Penstemon rostriflorus (Plantaginaceae) and cross‐species amplification

Eight novel microsatellite primer pairs are presented for Penstemon rostriflorus, representing the first microsatellite markers available for this genus. Loci were characterized for 20 individuals from two populations in the Great Basin, USA. All loci are polymorphic within P. rostriflorus (seven to 13 alleles per locus; observed heterozygosity between 0.40 and 0.95), and therefore useful for population genetic studies within the species. Cross‐species transferability was tested on 40 additional species of Penstemon, and results indicate that these primers pairs will likely be useful for population genetic studies on many Penstemon species.