Polymorphism in Pomatoceros

Allelic variation at 13 sites for the intertidal Pomatoceros lamarckii (Quatrefages) and at six sites for the sublittoral P. triqueter (L.) in North-West Europe, when examined at 19 metabolic enzyme loci, showed significant differences between sites in the frequencies of the various enzyme morphs. There was no readily interpretable geographic or environmental trend. The Hardy-Weinberg equilibrium was maintained at almost all loci and all sites. The genetic distance between populations was very small in comparison with the difference between the two species.
Colour morphs of P. triqueter showed a clear latitudinal cline with blue and brown morphs predominating at more northerly latitudes, and grey, orange and red morphs at more southerly latitudes.
Interlocality variations in allele frequency, though statistically significant, were never sufficient to alter the order of frequency of the two or three commonest alleles, so that the frequency of all but rare alleles showed a consensus throughout the ranges studied for each species. Accounting for local genetic patchiness in gene frequencies in species with pelagic larvae raises problems if the alleles are selectively neutral; accounting for wide geographic uniformity raises problems if alleles are selected for or against by environmental factors.     

Assortative mating among rock-dwelling cichlid fishes supports high estimates of species richness from Lake Malawi

It has been estimated that Lake Malawi, Africa, contains 500–650 endemic species of cichlid fishes, the largest number of vertebrate species endemic to any comparable sized area on the planet. As many of these putative species cannot be distinguished anatomically, these estimates of species richness depend to a great extent on the assumption that sympatrically occurring male colour morphs represent biological species. We have tested this assumption using a combination of behavioural observations of courtship and microsatellite DNA analysis for six putative species of the Pseudotropheus ( Tropheops ) complex and three of the Pseudotropheus ( Maylandia ) complex occurring sympatrically at Nkhata Bay. We were unable to demonstrate assortative courtship for the species pairs Pseudotropheus ( Maylandia ) zebra / P . 'gold zebra' or P. ( Tropheops ) 'band'/ P. ( T .) 'rust' because we were unable to distinguish between the females of these taxa. All other taxa showed clear assortative courtship, except for P. ( T .) 'deep', a deep-water species which was rarely observed. Fixation indices (θ_ST for the infinite allele model, and R _ST for the stepwise mutation model) calculated from six microsatellite DNA loci demonstrated significant deviations from panmixia in all pairwise comparisons of putative species, indicating little or no gene flow between populations. All taxa showed high levels of allelic diversity providing evidence that genetic bottlenecking may have been of limited importance in the speciation process. Assortative mating among taxa differing only in male colouration is supportive of theories that speciation in these fishes has been driven by sexual selection by female choice.     

Evaluating Genetic Diversity for the Conservation of the Threatened Galapagos Endemic Calandrinia galapagosa (Portulacaceae)

Calandrinia galapagosa (Portulacaceae) is a threatened plant endemic to San Cristobal Island, Galapagos. The species is threatened by goat herbivory: less than 1300 individuals remain in nine populations, three of which have been fenced for protection. It is not known whether these three populations are genetically representative of the species at large or whether the three distinct color morphs exhibited by the species represent different taxonomic units. We measured the frequency of 113 polymorphic amplified fragment length polymorphism markers in 189 specimens representing three different phenotypes from all nine populations to relate the genetic structure of the species to both its geological history and morphological variation. The analysis revealed high diversity within populations and low, but significant, differentiation between populations. In addition, no genetic diversification was observed across color morphs. We conclude that (1) regular gene flow has historically occurred between populations, and (2) individual color morphs are not separate taxa. We identify another population that should be fenced to ensure maximal protection of genetic variation. Our research serves as a case study of using population molecular genetics for optimizing protection strategies for rare plants in tropical archipelagoes affected by introduced herbivores.     

A taxometric analysis ofLimonium (Plumbaginaceae) in Western Europe

A taxometric analysis ofLimonium species in Western Europe has provided evidence for the subgeneric groupings proposed by other authors. A cluster analysis suggests that the origin of sexual and agamospermous species has taken place separately. Sexual and agamospermous species are usually clustered separately. The lack of a sexual process in many agamospermous species, as indicated by the distribution of incompatibility morphs and by very low pollen fertility, suggests that the evolution of sexual and agamospermous taxa has been by different processes. It is suggested that following the origin of agamospermous species or species groups by hybridisation, subsequent evolution in the agamospermous taxa has occurred by asexual means. Evidence from the distribution of pollen/stigma incompatibility morphs, especially the existence of self-compatible agamospermous species supports this hypothesis.     

Phylogeny, genetic variability and colour polymorphism of an emerging animal model: the short-lived annual Nothobranchius fishes from southern Mozambique

Nothobranchius are a group of small, extremely short-lived killifishes living in temporary savannah pools in Eastern Africa and that survive annual desiccation of their habitat as dormant eggs encased in dry mud. One mitochondrial (COI) and three nuclear (CX32.2, GHITM, PNP) loci were used to investigate the phylogenetic relationship of Nothobranchius species from southern and central Mozambique. This group shows marked variation in captive lifespan at both the inter- and intraspecific levels; lifespan varies from a few months to over a year. As their distribution encompasses a steep gradient between semi-arid and humid habitats, resulting in contrasting selection pressures on evolution of lifespan and associated life history traits, Mozambican Nothobranchius spp. have recently become a model group in studies of ageing, age-related disorders and life history evolution. Consequently, intraspecific genetic variation and male colour morph distribution was also examined in the recovered clades. Using Bayesian species tree reconstruction and single loci analyses, three large clades were apparent and their phylogenetic substructure was revealed at the inter- and intra-specific levels within those clades. The Nothobranchius furzeri and Nothobranchius orthonotus clades were strongly geographically structured. Further, it was demonstrated that male colour has no phylogenetic signal in N. furzeri, where colour morphs are sympatric, but is associated with two reciprocally monophyletic groups in Nothobranchius rachovii clade, where colour morphs are parapatric. Finally, our analysis showed that a polymorphism in the Melanocortin1 receptor gene (which controls pigmentation in many vertebrates and was a candidate gene of male colouration in N. furzeri) is unrelated to colour phenotypes of the study species. Our results raise significant implications for future comparative studies of the species and populations analysed in the present work.     

Phylogenetic analysis of RAD‐seq data: examining the influence of gene genealogy conflict on analysis of concatenated data

One of the major issues in phylogenetic analysis is that gene genealogies from different gene regions may not reflect the true species tree or history of speciation. This has led to considerable debate about whether concatenation of loci is the best approach for phylogenetic analysis. The application of Next‐generation sequencing techniques such as RAD‐seq generates thousands of relatively short sequence reads from across the genomes of the sampled taxa. These data sets are typically concatenated for phylogenetic analysis leading to data sets that contain millions of base pairs per taxon. The influence of gene region conflict among so many loci in determining the phylogenetic relationships among taxa is unclear. We simulated RAD‐seq data by sampling 100 and 500 base pairs from alignments of over 6000 coding regions that each produce one of three highly supported alternative phylogenies of seven species of Drosophila. We conducted phylogenetic analyses on different sets of these regions to vary the sampling of loci with alternative gene trees to examine the effect on detecting the species tree. Irrespective of sequence length sampled per region and which subset of regions was used, phylogenetic analyses of the concatenated data always recovered the species tree. The results suggest that concatenated alignments of Next‐generation data that consist of many short sequences are robust to gene tree/species tree conflict when the goal is to determine the phylogenetic relationships among taxa.     

Molecular phylogeny and larval morphological diversity of the lanternfish genus Hygophum (Teleostei: Myctophidae)

Larvae of the deep-sea lanternfish genus Hygophum (Myctophidae) exhibit a remarkable morphological diversity that is quite unexpected, considering their homogeneous adult morphology. In an attempt to elucidate the evolutionary patterns of such larval morphological diversity, nucleotide sequences of a portion of the mitochondrially encoded 16S ribosomal RNA gene were determined for seven Hygophum species and three outgroup taxa. Secondary structure-based alignment resulted in a character matrix consisting of 1172 bp of unambiguously aligned sequences, which were subjected to phylogenetic analyses using maximum-parsimony, maximum-likelihood, and neighbor-joining methods. The resultant tree topologies from the three methods were congruent, with most nodes, including that of the genus Hygophum, being strongly supported by various tree statistics. The most parsimonious reconstruction of the three previously recognized, distinct larval morphs onto the molecular phylogeny revealed that one of the morphs had originated as the common ancestor of the genus, the other two having diversified separately in two subsequent major clades. The patterns of such diversification are discussed in terms of the unusual larval eye morphology and geographic distribution.     

Three species of Isotoma (Collembola, Isotomidae) based on morphology, isozymes and ecology

Morphological markers and isozymes were used for identifying three presumed species of the Isotoma genus. Morphological traits separated three taxa of the genus. 10 isozymes determined by at least 11 loci were analysed from each taxon, and 2 loci were taxon-specific, supporting the hypothesis that the three taxa represented three species. The genetic variation found within the taxa measured as fraction of polymorphic loci at the 99% level was 0.82, 0.55 and 0.55 with the corresponding observed heterozygosity 0.15, 0.09 and 0.12. Two populations of the same taxon from Denmark and Norway, respectively, were very closely related. Additional ecological criteria, obtained from a literature survey, also revealed pronounced differences between the three taxa. Due to these facts three distinct species are proposed, namely I. anglicana Lubbock 1862, I. riparia Nicolet 1841 and I. viridis Bourlet 1839.     

Dental polymorphism and systematics in Saccodon, a neotropical genus of freshwater fishes (Parodontidae, Characoidei)

Population samples of algae-grazing, mountain stream characins of the genus Saccodon of Panama and northwestern South America are reported with up to four sharply differentiated dental morphs. The morphs differ strikingly in the form and numbers of functional teeth and of preformed replacement teeth. Recognition of the polymorphic nature of these variations necessitates new concepts of taxa at species and generic levels. Two species with similar polymorphism are involved, Saccodon dariensis from Panamá and Colombia, and Saccodon wagneri from the Guayas basin on the Pacific coast of Ecuador. A total of rive morphs have been discovered, three of which are homologous in the two species.     

Evidence of genetic and reproductive isolation between two morphs of subtropical-dominant coral Acropora solitaryensis in the non-reef region of Japan

We reveal the existence of a cryptic species of Acropora solitaryensis ( Veron and Wallace, 1984 ), a dominant species in high-latitude coral communities. Although some morphs, such as arborescent table (AR), solid plate (PL), and intermediate (IM) forms, had been known in this species, it was unclear whether these are reproductively isolated from one another. Here, potential reproductive exchange between two representative morphs, AR and PL, were examined using genetic and reproductive methods. Molecular phylogenetic analyses using both mitochondrial and nuclear molecular markers (mitochondrial control region and mini-collagen intron, respectively) indicated that AR is clearly distinct from PL, suggesting that gene flow between the morphs is absent. In cross-fertilization experiments, gametic compatibility between AR and PL was extremely low, suggesting prezygotic isolation of these morphs. These results strongly suggest that AR and IM forms are variations of A. solitaryensis, whereas PL form may be an undescribed species. In addition, AR was closely related genetically to A. pruinosa, which is a high-latitude species with arborescent form, and AR and A. pruinosa were able to hybridize, although with lower fertilities than observed in intra-specific crosses. The two species are thus likely to have speciated not in tropical regions, but in non-reef regions due to habitat segregation.     

POPULATION GENETIC STRUCTURE IN DIPLOID BLUEBERRY VACCINIUM SECTION CYANOCOCCUS (ERICACEAE)

Vaccinium section Cyanococcus comprises diploid, tetraploid, and hexaploid species (x = 12) all of which are highly self-sterile. In order to assess the distribution of genetic variation within and among the diploids, population genetic analyses of allozyme data were conducted on three species: V. elliottii, V. myrtilloides, and V. tenellum. Populations were located throughout the range of these taxa in eastern North America. Data were collected at 20 loci, 12 of which are mendelian based on formal genetic analyses. Consistent with expectations for out-crossing taxa, these species exhibited high levels of variation within populations. Eighteen loci were polymorphic with the number and frequency of alleles varying among taxa. Mean values for the number of alleles per polymorphic locus, proportion of polymorphic loci, and expected heterozygosity were 2.8, 48.2, and 0.148, respectively. High infraspecific genetic identities exceeding 0.9 indicated that these taxa are homogeneous. All populations were in Hardy-Weinberg equilibrium with slight heterozygote excess observed in V. myrtilloides and V. tenellum. Although total genetic diversity was lower than that observed for other predominantly out-crossing species, it was apportioned similarly. The majority could be attributed to differences among individuals within populations. Conspecific populations were relatively undifferentiated with genetic differentiation similar to other self-incompatible species.     

Flickers of speciation: Sympatric colour morphs of the arc‐eye hawkfish,Paracirrhites arcatus,reveal key elements of divergence with gene flow

One of the primary challenges of evolutionary research is to identify ecological factors that favour reproductive isolation. Therefore, studying partially isolated taxa has the potential to provide novel insight into the mechanisms of evolutionary divergence. Our study utilizes an adaptive colour polymorphism in the arc‐eye hawkfish (Paracirrhites arcatus) to explore the evolution of reproductive barriers in the absence of geographic isolation. Dark and light morphs are ecologically partitioned into basaltic and coral microhabitats a few metres apart. To test whether ecological barriers have reduced gene flow among dark and light phenotypes, we evaluated genetic variation at 30 microsatellite loci and a nuclear exon (Mc1r) associated with melanistic coloration. We report low, but significant microsatellite differentiation among colour morphs and stronger divergence in the coding region of Mc1r indicating signatures of selection. Critically, we observed greater genetic divergence between colour morphs on the same reefs than that between the same morphs in different geographic locations. We hypothesize that adaptation to the contrasting microhabitats is overriding gene flow and is responsible for the partial reproductive isolation observed between sympatric colour morphs. Combined with complementary studies of hawkfish ecology and behaviour, these genetic results indicate an ecological barrier to gene flow initiated by habitat selection and enhanced by assortative mating. Hence, the arc‐eye hawkfish fulfil theoretical expectations for the earliest phase of speciation with gene flow.     

Genetic divergence among three morphs of Acentrogobius pflaumii (Gobiidae) around Japan and their identification using multiplex haplotype-specific PCR of mitochondrial DNA

Genetic divergence among three morphs of a goby, Acentrogobius pflaumii, was investigated on the basis of sequence variations in a partial section of the mitochondrial cytochrome oxidase subunit?I gene. A total of 95 specimens, representing the three morphs, collected almost sympatrically from three sites around the Japanese archipelago, were sequenced, the sequence divergences between the morphs (8.62–15.50%) being much greater than within each morph (0.44–0.57%). In addition, each morph formed a distinct cluster in a neighbor-joining tree. Together with an earlier morphological study, such evidence indicated that each morph clearly represented a distinct species. Based on the determined gene sequences, three morph-specific primers were designed for multiplex haplotype-specific polymerase chain reaction (PCR) to enable quick and easy identification of each morph in future studies, especially for early life ecology given morphological similarities at these stages.     

AFLP genome scans suggest divergent selection on colour patterning in allopatric colour morphs of a cichlid fish

Genome scan-based tests for selection are directly applicable to natural populations to study the genetic and evolutionary mechanisms behind phenotypic differentiation. We conducted AFLP genome scans in three distinct geographic colour morphs of the cichlid fish Tropheus moorii to assess whether the extant, allopatric colour pattern differentiation can be explained by drift and to identify markers mapping to genomic regions possibly involved in colour patterning. The tested morphs occupy adjacent shore sections in southern Lake Tanganyika and are separated from each other by major habitat barriers. The genome scans revealed significant genetic structure between morphs, but a very low proportion of loci fixed for alternative AFLP alleles in different morphs. This high level of polymorphism within morphs suggested that colour pattern differentiation did not result exclusively from neutral processes. Outlier detection methods identified six loci with excess differentiation in the comparison between a bluish and a yellow-blotch morph and five different outlier loci in comparisons of each of these morphs with a red morph. As population expansions and the genetic structure of Tropheus make the outlier approach prone to false-positive signals of selection, we examined the correlation between outlier locus alleles and colour phenotypes in a genetic and phenotypic cline between two morphs. Distributions of allele frequencies at one outlier locus were indeed consistent with linkage to a colour locus. Despite the challenges posed by population structure and demography, our results encourage the cautious application of genome scans to studies of divergent selection in subdivided and recently expanded populations.     

Genetic Diversity and Species-Diagnostic Markers of Mud Crabs (Genus Scylla) in Eastern Thailand Determined by RAPD Analysis

Genetic diversity of three mud crab species, Scylla serrata (Forsk?l), S. oceanica (Dana), and S. tranquebarica (Fabricius), collected from two locations in eastern Thailand (Chanthaburi and Trat) was examined by randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR). Ninety-one reproducible RAPD fragments, generated by UBC456, UBC457, and YNZ22, were polymorphic. The percentage of polymorphic bands within populations ranged from 47.92% to 77.59%. Species-specific RAPD markers were also observed and used to construct a molecular diagnostic key in these taxa. Large genetic differences between species were found (D(ij) = 0.425 to 0.751), whereas those between populations within each species were much lower (D(ij) = 0.171 to 0.199). The neighbor-joining tree based on genetic distances among pairs of individuals indicated three distinct groups, corresponding to S. serrata, S. oceanica, and S. tranquebarica. No genotypes were shared among these three species. This suggests the absence of genetic exchanges between sympatric mud crab species in eastern Thailand. Therefore, mud crabs in this area should be recognized as three different species rather than a single panmictic species exhibiting different morphs.     

Ancient allopolyploid speciation in Geinae (Rosaceae): evidence from nuclear granule-bound starch synthase (GBSSI) gene sequences

A nuclear low-copy gene phylogeny provides strong evidence for the hybrid origin of seven polyploid species in Geinae (Rosaceae). In a gene tree, alleles at homologous loci in an allopolyploid species are expected to be sisters to orthologues in the ancestral taxa rather than to each other. Alleles at a duplicated locus in an autopolyploid, however, are expected to be more closely related to each other than they are to any orthologous copies in closely related species. We cloned and sequenced about 1.9 kilobases from the 5' end of the GBSSI-1 gene from two diploid, one tetraploid, and six hexaploid species. Each of the three loci in the hexaploid species forms a separate group, two of which are more closely related to copies in other species than they are to each other. This finding indicates that the hexaploid lineage evolved through two consecutive allopolyploidization events. Based on the GBSSI-1 gene tree, we hypothesized that there was an initial hybridization between a diploid species from the ancestral lineage of Coluria and Waldsteinia and an unknown diploid species to form the tetraploid Geum heterocarpum lineage. Backcrossing of G. heterocarpum with a representative of the unknown diploid lineage then resulted in a hexaploid lineage that has radiated considerably since its origin, comprising at least 40 extant species with various morphologies. A penalized likelihood analysis indicated that Geinae may be about 17 million years old, implying that the hypothesized allopolyploid speciation events are relatively ancient. Six of the 22 cloned Geinae GBSSI-1 copies in this study, which all are duplicate copies in polyploid taxa, may have become pseudogenes. We compared the GBSSI-1 phylogeny with one from chloroplast data and explored implications for the evolution of some fruit characters.     

Re-examination of a proposed case of stasipatric speciation: phylogeography of the Australian morabine grasshoppers (Vandiemenella viatica species group)

Karyotypic differences have been used for delimiting populations or species, although whether these mutations provide strong barriers to gene flow between populations and promote speciation remains contentious. In this study, we assessed whether 11 chromosomal races of Australian morabine grasshoppers ( Vandiemenella viatica species group) represent genetically distinct populations by analyses of cytological and allozyme (35 loci) data and DNA sequences of the elongation factor-1 alpha ( EF -1α), anonymous Mvia11 , and mitochondrial cytochrome c oxidase subunit I ( COI ) loci. While the Vandiemenella chromosomal taxa generally represent genetically distinct units, a substantial portion of the total genetic variation in our samples was not explained by the chromosomal variation. Mantel tests indicated that Vandiemenella populations were spatially structured and have maintained gene flow at a local scale within each of the taxa. The group was subdivided into 13 genetic clusters; four chromosomal taxa comprised single exclusive clusters, while others comprised more than one cluster or clusters shared with other taxa. Boundaries of these cryptic population subdivisions correspond with several biogeographical barriers, such as straits, gulfs, the Murray River, and an ancient mega-lake, Lake Bungunnia. The viatica species group was previously proposed to have diversified without major geographical separation based on the stasipatric speciation model; however, the present study suggests the involvement of allopatric fragmentation. Given extensive nonmonophyly of chromosomal taxa and incomplete barriers to gene flow among taxa, all Vandiemenella chromosomal taxa and genetically distinct populations within chromosomal taxa, except Vandiemenella pichirichi , should be regarded as populations of one species: Vandiemenella viatica .     

Multiple origins of self-fertilization in tristylous Eichhornia paniculata (Pontederiaceae): Inferences from style morph and isozyme variation

Populations of Eichhornia paniculata (Pontederiaceae) exhibit a wide range of mating systems, from predominant outcrossing to high levels of self-fertilization. The origin of self-fertilization in this tristylous species is associated with the loss of style-length morphs from populations and the spread of self-pollinating, floral variants. We examined geographic variation in style morph and allozyme frequencies to determine whether the loss of style morphs and transition to selfing could have multiple origins in E. paniculata. Surveys of floral variation in 167 populations from six states in northeastern Brazil revealed that at least one style morph was absent from 29.3%. Non-trimorphic populations occurred in all states and ranged in frequency from 9% in Ceará to 68% in Alagoas. Selfing variants occurred in 8.5% and 55% of trimorphic and non-trimorphic populations, respectively, and were distributed among five of six states with primary concentrations in Alagoas and Pernambuco. A comparison of electrophoretic variation at 24 isozyme loci in 28 trimorphic, 13 dimorphic and 3 monomorphic populations indicated that non-trimorphic populations contained 84% of the allelic variation present in trimorphic populations and were markedly differentiated from one another. Analyses of genetic distance and the distribution of rare alleles indicated that non-trimorphic populations were often more similar to neighbouring trimorphic populations than to one another. Populations with selfing variants occurred at low frequency in three genetically distinct parts of the range. These results, in combination with genetic and morphological evidence suggest that style morphs are lost repeatedly from populations of E. paniculata and that selfing variants may have originated on at least three separate occasions in northeastern Brazil.