Population differentiation and species cohesion in two closely related plants adapted to neotropical high-altitude 'inselbergs', Alcantarea imperialis and Alcantarea geniculata (Bromeliaceae)

Isolated granitic rock outcrops or 'inselbergs' may provide a window into the molecular ecology and genetics of continental radiations under simplified conditions, in analogy to the use of oceanic islands in studies of species radiations. Patterns of variability and gene flow in inselberg species have never been thoroughly evaluated in comparison to related taxa with more continuous distribution ranges, or to other species in the same kingdom in general. We use nuclear microsatellites to study population differentiation and gene flow in two diploid, perennial plants adapted to high-altitude neotropical inselbergs, Alcantarea imperialis and Alcantarea geniculata (Bromeliaceae). Population differentiation is pronounced in both taxa, especially in A. imperialis. Gene flow in this species is considerably lower than expected from the literature on plants in general and Bromeliaceae in particular, and too low to prevent differentiation due to drift (N(e)m < 1), unless selection coefficients/effect sizes of favourable alleles are great enough to maintain species cohesion. Low gene flow in A. imperialis indicates that the ability of pollinating bats to promote gene exchange between inselbergs is smaller than previously assumed. Population subdivision in one inselberg population of A. imperialis appears to be associated with the presence of two colour morphs that differ in the coloration of rosettes and bracts. Our results indicate a high potential for inselbergs as venues for studies of the molecular ecology and genetics of continental radiations, such as the one that gave rise to the extraordinary diversity of adaptive strategies and phenotypes seen in Bromeliaceae.     

A set of polymorphic microsatellite loci for Vriesea gigantea and Alcantarea imperialis (Bromeliaceae) and cross‐amplification in other bromeliad species

Fifteen polymorphic microsatellite markers were isolated and characterized in two species of Bromeliaceae: Vriesea gigantea and Alcantarea imperialis. The number of alleles observed for each locus ranged from three to 16. The loci will be used for studies of the genetic structure of natural populations, reproductive biology, and evolutionary relationships among and within these genera. A cross‐amplification test in 22 taxa suggests that the markers will be useful for similar applications in numerous other bromeliad species.     

Within-population spatial genetic structure in four naturally fragmented species of a neotropical inselberg radiation, Alcantarea imperialis, A. geniculata, A. glaziouana and A. regina (Bromeliaceae)

Studies of organisms on 'terrestrial islands' can improve our understanding of two unresolved issues in evolutionary genetics: the likely long-term effects of habitat fragmentation and the genetic underpinnings of continental species radiations in island-like terrestrial habitats. We have addressed both issues for four closely related plant species of the adaptive radiation Bromeliaceae, Alcantarea imperialis, A. geniculata, A. regina and A. glaziouana. All four are adapted to ancient, isolated inselberg rock outcrops in the Brazilian Atlantic rainforest and are thus long-term fragmented by nature. We used eight nuclear microsatellites to study within-population spatial genetic structure (SGS) and historical gene dispersal in nine populations of these species. Within-population SGS reflected known between-species differences in mating systems. The strongest SGS observed in A. glaziouana (Sp=0.947) was stronger than literature estimates available for plants. Analysis of short- and long-distance components of SGS identified biparental inbreeding, selfing and restricted seed dispersal as main determinants of SGS, with restricted pollen dispersal by bats contributing in some localities. The ability of Alcantarea spp. to colonize isolated inselbergs probably stems from their flexible mating systems and an ability to tolerate inbreeding. Short-ranging gene dispersal (average sigma=7-27 m) is consistent with a loss of dispersal power in terrestrial island habitats. Population subdivision associated with sympatric colour morphs in A. imperialis is accompanied by between-morph differences in pollen and seed dispersal. Our results indicate a high potential for divergence with gene flow in inselberg bromeliads and they provide base-line data about the long-term effects of fragmentation in plants.     

Isolation and characterization of microsatellite loci in Pitcairnia albiflos (Bromeliaceae), an endemic bromeliad from the Atlantic Rainforest, and cross-amplification in other species

Eight microsatellite markers were isolated from Pitcairnia albiflos, an endangered endemic bromeliad species restricted to 'inselberg' rock outcrops in the state of Rio de Janeiro, Brazil. The number of alleles observed for each locus ranged from two to 12. Average observed and expected heterozygosities were 0.408 and 0.663, respectively. The cross-amplification test in 16 taxa suggests that the markers will be useful in numerous related bromeliad species. The loci will be used to study genetic structure and reproductive biology in fragmented inselberg populations and the origin and maintenance of barriers to gene flow between sympatric Pitcairnia species.     

A cladistic phylogeny of the genus Littorina (Gastropoda): implications for evolution of reproductive strategies and for classification

Gross anatomical characters of all 18 species of Littorina are used to construct a phylogenetic hypothesis for the genus, by the method of cladistic analysis. The resulting cladogram suggests that of the four subgenera of Littorina, one (Littorina) is paraphyletic. It is uncertain whether the genus Mainwaringia should be included in Littorina. It is shown that the non-planktotrophic Littorina species in the northern Atlantic comprise a monophyletic group, with the sister-species L. kurila and/or L. subrotundata in the northern Pacific. Invasion of the Atlantic by a minimum of two Pacific species, across the Arctic migration route established during the late Cenozoic, is sufficient to account for the modern distribution of the subgenera Littorina and Neritrema. The importance of the cladogram as a basis for hypotheses of adaptation is illustrated by a discussion of spawn and development in Littorina.     

Population structure and gene flow of the Atlantic walrus (Odobenus rosmarus rosmarus) in the eastern Atlantic Arctic based on mitochondrial DNA and microsatellite variation

The population structure of the Atlantic walrus, Odobenus rosmarus rosmarus , was studied using 11 polymorphic microsatellites and restriction fragment length polymorphism detected in the NADH-dehydrogenase ND1, ND2 and ND3/4 segments in mtDNA. A total of 105 walrus samples were analysed from northwest (NW) Greenland, east (E) Greenland, Svalbard and Franz Joseph Land. Two of the 10 haplotypes detected in the four samples were diagnostic for the NW Greenland sample, which implied that the group of walruses in this area is evolutionary distinct from walruses in the other three areas. One individual sampled in E Greenland exhibited a Pacific haplotype, which proved a connection between the Pacific walrus and walruses in eastern Greenland. The Franz Joseph Land, Svalbard and E Greenland samples shared the most common haplotype, indicating very little differentiation at the mtDNA level. Gene flow ( Nm ) estimates among the four areas indicated a very restricted exchange of female genes between NW Greenland and the more eastern Atlantic Arctic samples, and a closer relationship between the three samples composing the eastern Atlantic Arctic. The genetic variation at 11 polymorphic microsatellite loci grouped individuals into three populations, NW Greenland, E Greenland and a common Franz Joseph Land–Svalbard population, which were connected by moderate gene flow.     

A genetic analysis of some components of reproductive isolation between two closely related species,Spodoptera latifascia (Walker) and S. descoinsi (Lalanne-Cassou and Silvain) (Lepidoptera: Noctuidae)

The two closely related noctuid moths Spodoptera latifascia and S. descoinsi have been found in sympatry in some parts of French Guiana. They differ by i) ecological preferences, ii) some anatomical features of the genital tract, iii) the relative amounts of the two main components of the female pheromonal blend, and iv) the temporal pattern of sexual behaviour. Viable and fertile interspecific hybrids can be obtained in the laboratory, thus permitting a genetic study of some of these traits. The anatomical differences cause perturbations in mating efficiency. Experimental results show that these differences are hereditarily transmitted, but their exact mode of inheritance remains unknown. The timing of initiation of female calling activity is very likely to be under polygenic control. Conclusive evidence showed that a pair of major autosomal alleles determined the pheromone composition. Moreover, some of the data suggest that the descoinsi allele would become lethal in certain mosaic latifascia/descoinsi genomic environments. The ecological factor is likely to be the major component of reproductive isolation between the two species in natural conditions. However in contact zones, the differences in pheromone composition and in timing of sexual activity are probably involved in preventing interspecific hybridization. Two speciation scenarios are proposed, both sympatric. In one of them, the primary event would be a fragmentation of the tropical forest followed by disruptive selection associated with habitat diversification. In the other, the speciation process might have been initiated by the occurrence of mutations acting on the pheromone mediated mate recognition system.     

Isolation and characterization of microsatellites in the woody shrub, Banksia sphaerocarpa var. caesia (Proteaceae)

Microsatellite markers were developed for the Australian bird-pollinated woody shrub Banksia sphaerocarpa var. caesia to study gene flow among populations in a highly fragmented landscape. Eight loci were developed, and in a sample of 40 individuals from one population, the number of alleles per locus ranged from five to 21 and observed heterozygosities ranged from 0.385 to 0.914. All eight loci showed independent inheritance. Analysis of open-pollinated progeny arrays confirmed Mendelian inheritance at seven loci, while null alleles were suspected at the remaining locus.     

Conservation implications of high genetic variation in two closely related and highly threatened species of Crambe (Brassicaceae) endemic to the island of Gran Canaria: C. tamadabensis and C. pritzelii

We used data from 12 allozyme loci for two endemic Brassicaceae from Gran Canaria (the endangered narrow endemic Crambe tamadabensis and its more widespread congener C. pritzelii) to assess whether their genetic diversity patterns reflect their phylogenetic closeness and contrasting population sizes and distribution areas, and to derive conservation implications. Genetic diversity values are high for both species and slightly higher in C. tamadabensis, despite its narrow distribution in north‐western Gran Canaria. At odds with the generally high interpopulation diversity levels reported in Canarian endemics, values of G_ST in C. tamadabensis and C. pritzelii are rather low (0.067 and 0.126, respectively). We construe that the higher genetic structure detected in C. pritzelii is mainly a result of unbalanced allele frequencies and low population sizes at the edges of its distribution. The overall high allozyme variation detected in C. tamadabensis and C. pritzelii is nevertheless compatible with an incipient but consistent genetic differentiation between the two species, modulated by recurrent bottlenecks caused by grazing and drift. Our data suggest that conservation efforts aimed at maintaining the existing genetic connectivity in each species and ex situ conservation of seeds are the best strategies to conserve their genetic diversity.     

Genetic variation and differentiation in the lizard, Podarcis wagleriana (Reptilia: Lacertidae)

The electrophoretic variation at 26 presumptive gene loci was investigated in populations of Podarcis ivagleriana from Sicily, the Aegadian Islands, and the Aeolian Islands. For interspecific comparison, samples of the closely related lizard P. sicula from the same geographic area were also used. Population heterogeneity analyses carried out by the estimation of F-statistics and Nei's standard genetic distance, showed a high genetic homogeneity within P. sicula, but a noticeable genetic differentiation within P. wagleriana. In the latter species, Nei's D ranged from 0 to 0.212, and this is because the Aeolian populations were quite distinct from those inhabiting Sicily and the Aegadian Islands. Fixed differences identified at three loci (Ck, Ada, Gp-4) contributed to a relatively high value of Nei's standard genetic distance between the two population groups (D= 0.147). This value is very similar to those found comparing pairs of well-recognized biological species included in the genera Podarcis and Lacerta. Estimation of the time of evolutionary divergence shows that the Aeolian and Sicilian populations of P. wagleriana have been isolated geographically for a long time (0.7 Myr according to Nei's formula; 2 Myr according to Sarich's calibration), indicating evolutionary divergence at the species level. Based on genetic and biogeographic data, it is suggested the recognition of full specific status for the Aeolian populations, for which the name P. rajfonei comb. nova (Aeolian wall lizard) is proposed. Electrophoretic data and comparative species distributions suggest that (1) Podarcis sicula recently colonized the Aeolian Islands, and (2) it has competed successfully with P. rajfonei in this area, greatly reducing the range of the latter and causing the extinction of most of its populations. In fact, P. sicula is widespread in the Aeolian Archipelago, while P. rajfonei is confined at present to one large island (Vulcano) and three tiny islands (Strombolicchio, Scoglio Faraglione, La Canna). This can be considered a classic example of competitive exclusion of a native form (P. rajfonei) by a species accidentally introduced by man (P. sicula).     

Genetic diversity, gene flow and drift in Bavarian red deer populations (Cervus elaphus)

The red deer (Cervus elaphus) populationof Bavaria in Southern Germany was severelyreduced during the 19th century due toover-hunting. The species has since recoveredwithin designated areas. Subsequent habitatfragmentation presumably has changed thegenetic structure of Bavarian red deer.In order to assess the genetic diversity, weanalysed samples obtained from nine differentBavarian and two adjacent (Thueringen andCzech-Republic) red deer populations,genotyping 19 microsatellite loci. Our analysesrevealed moderate and significant differencesin diversity. Referring to assignment tests,the genetic differentiation of Bavarian reddeer was sufficient to assign an individual'sorigin to the correct population at an averageof 91.6%. The correlation of genetic andgeographic distance matrices revealed noevidence for isolation by distance. Thecoalescent model analysis suggests that thegenetic structure used to be characterized by adrift–gene flow equilibrium and is nowinfluenced by drift and disruption of the geneflow. Only three of the examined populationsshowed a probability of less than 10% that twogenes within these populations share a commonancestor (F _IS-value). Twopopulations had high F _IS values,indicating the influence of drift.Additionally, the intrapopulation indicesrevealed a low variability in thesepopulations. The estimated effective populationsizes (N _e) were generally in thesame range as the actual population sizes. Theinbreeding rates, based on the estimated N _e, and the inbreeding coefficientssuggested that the Bavarian red deerpopulations are in a stable state.     

Genetic variation and diadromy in some native New Zealand galaxiids (Teleostei: Galaxiidae)

We examined genetic variation at 13 isozyme loci in three diadromous and two non-diadromous species of galaxiid fish from streams in South and Stewart Islands of New Zealand. Analysis of allele frequency data revealed higher F_s7 and Nei's D values among conspecific populations of the non-diadromous species. We propose that the genetic structurings of these species are affected by their migratory behaviours. We suggest that the derived condition of non-diadromous behaviour may increase scope for speciation, but also the probability of extinction. This is exacerbated by population fragmentation over the last century.     

Genetic structure and phylogeographic relationships of Fagus sylvatica L. woods in Lazio (Central Italy)

The genetic characterisation of beech woods in Lazio was performed by microsatellite approach. The phylogeographic relationship existing among stands was investigated, allowing us to identify a putative glacial refugium for this species. This work provides information for management policies and conservation strategies of F. sylvatica germplasm resources in central Italy.     

Genetic variation and structure in Scandinavian red deer (Cervus elaphus): influence of ancestry,past hunting,and restoration management

In the 19^th century, the red deer (Cervus elaphus) population in Sweden experienced a rapid decline in numbers and distribution. A small population was, however, remnant in the southernmost province (Skåne) of the country, presumably corresponding to the nominate form of red deer (Cervus elaphus elaphus Linnaeus, 1758). After management, reintroductions, and supplementary release during the 20^th century the Swedish C. elaphus population recovered. The recovery was partially uncontrolled, and included introductions of C. elaphus of continental origin. In northern central Sweden (Jämtland) the current C. elaphus population may stem from natural colonization from Norway and/or from specimens of Swedish origin that have escaped from enclosures. To evaluate the status of the current, partially separated populations, we investigated variation at microsatellite markers in 157 C. elaphus specimens from ten locations in Sweden and Norway. Analyses suggest that the highest‐likelihood phylogenetic structure among the individuals sampled is described four distinct genetic clusters: (1) animals from the province of Västergötland in south‐western Sweden; (2) deer from the southernmost province of Skåne; (3) deer from the provinces Jämtland, Blekinge, and Västmanland; and (4) Norwegian deer. Cervus elaphus from a captive herd at the Skåne Zoo cluster with deer from Skåne or deer from Västergötland, depending on the method of analysis. A number of populations in Sweden may genetically match the nominate form of red deer (C. e. elaphus). The recently established C. elaphus population in Jämtland seems to stem mainly from escapees from enclosures, with a mixed ancestry from the wild remnant population in Skåne and continental deer, whereas the influx from Norway is minor, if any. Our results show the need for a detailed assessment of genetic differentiation, and emphasize the value of local management plans when planning and managing introductions. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 43–53.     

Three ways to distinguish species: using behavioural,ecological, and molecular data to tell apart two closely related ants,Camponotus renggeri and Camponotus rufipes (Hymenoptera: Formicidae)

The closely related Camponotus renggeri and Camponotus rufipes (subgenus Myrmothrix) often live in sympatry in the Brazilian ‘cerrado’ savannah, and are distinguished by nuances in their blackish body colour and by the colour of the legs. Variation in morphological characters, however, makes species separation difficult and it has been suggested that the two species should be merged into one. As appropriate species identification is essential for studies in ecology and evolutionary biology, here we examine how natural history data (habitat preference, nesting biology) and molecular tools (nuclear and mitochondrial markers) perform in distinguishing sympatric populations of C. renggeri and C. rufipes. In our study area, C. rufipes was only seen in cerrado sensu stricto (scrub of shrubs and trees), whereas C. renggeri occurred in cerrado sensu stricto and cerradão (closed woodland). Camponotus renggeri nested underground or in fallen/erect dead trunks, whereas C. rufipes constructed distinctive nests of dry straw. Nest persistence through time was higher in C. rufipes, especially in the hot/rainy season. Nest distribution was random in C. renggeri and aggregated in C. rufipes. Molecular data consistently showed that, regardless of the source of genetic variation, the uppermost hierarchical level of divergence is observed between species, unambiguously differentiating the individuals identified as C. renggeri and C. rufipes as two independent evolutionary lineages. Mitochondrial data throughout the species' geographical ranges further confirmed a consistent genetic divergence between C. renggeri and C. rufipes along their distribution in Brazil. Our integrated approach combining morphological traits with natural history and molecular data confirms that C. renggeri and C. rufipes are valid species that can be separated in our study area relatively well. © 2015 The Linnean Society of London     

Genetic and geographic differentiation in the Rio Negro tuco-tuco (Ctenomys rionegrensis): inferring the roles of migration and drift from multiple genetic markers

Among tuco-tucos, Ctenomys rionegrensis is especially amenable to the study of the forces driving population differentiation because of the restricted geographic range it occupies in Uruguay. Within this limited area, the Rio Negro tuco-tuco is limited to sandy soils. It nonetheless exhibits remarkable variation in pelage color, including melanic, agouti, and dark-backed individuals. Two hypotheses have been put forth to explain this pattern: (1) local differentiation and fixation of alternative pelage types by genetic drift under limited gene flow; or (2) fixation by natural selection that may take place even in the presence of gene flow. A previous allozyme study rejected the genetic drift hypothesis on the basis of high inferred levels of migration. New estimates of gene flow from microsatellites and mitochondrial cytochrome b sequences were obtained for C. rionegrensis populations to further test these hypotheses. Much lower levels of gene flow were estimated with these more sensitive markers. Microsatellite-based estimates of gene flow are close to zero and may come closest to estimating current levels of migration. A lack of equilibrium between migration and genetic drift is also strongly suggested by the absence of an isolation-by-distance pattern found in all three genetic datasets. The microsatellite genotype data show that the species is strongly structured geographically, with subpopulations constituting distinct genetic entities. If current levels of gene flow are very low, as indicated by the new data, the local fixation of alternative alleles, including those responsible for pelage color polymorphism, is possible by drift alone. A scenario is thus proposed in which the species expanded in the recent past from a more restricted geographic range and has subsequently differentiated in near isolation, with genetic drift possibly playing a primary role in overall genetic differentiation. The local fixation of pelage color types could also be due to drift, but selection on this trait cannot be ruled out without direct analysis.     

Genetic variation in natural populations of Anthurium sinuatum and A. pentaphyllum var. pentaphyllum (Araceae) from north‐east Brazil using AFLP molecular markers

Genetic variation was investigated using AFLP markers in 12 populations of Anthurium sinuatum and A. pentaphyllum var. pentaphyllum (Araceae) in north‐east Brazil, Amazonia and the Brazilian Atlantic forest. Two unique genetic patterns characterized the populations of A. sinuatum as a group, but no correlation between genetic and geographical interpopulation distance was found; the Amazonian population was not separated from that in Ceará. The isolated Ceará brejo populations of A. sinuatum were genetically distinct, but genetic diversity levels were similar to populations elsewhere, with no evidence of genetic erosion. Anthurium pentaphyllum populations were significantly different from each other; Bayesian genetic structural analysis found no common genetic pattern, but revealed genetic clusters unique to subgroups and individual populations in the Atlantic forest and French Guiana. Anthurium pentaphyllum and A. sinuatum can be distinguished genetically, but individuals of both species formed intermediate genetic clusters that blurred their distinction. We suggest that genetic mixing of A. sinuatum and A. pentaphyllum has occurred in north‐east Brazil, possibly connected with cycles of humid forest expansion. The weak genetic structure in A. sinuatum is consistent with the natural fragmentation of continuous forest areas, possibly during the Holocene. This study highlights the scientific importance of the highly threatened brejo forests for tropical American biogeography. © 2009 The Linnean Society of London, Botanical Journal of the Linnean Society, 2009, 159 , 88–105.     

Genetic variation and population structure in Korean endemic species: III.Hosta minor (Liliaceae)

Hosta minor, an insect-pollinated and rhizomatous herbaceous perennial, occurs in eastern and southern Korean Peninsula. AlthoughH. minor is a Korean endemic species and most populations of the species are scattered, the species maintains considerably high levels of genetic variation within the species, with a moderate level of variation (14%) found among populations. Nineteen of the 29 putative loci resolved (66%) were polymorphic within the species, the mean number of alleles per locus was 2.10 across all populations and averaged 1.68 within populations. In addition, genetic diversity was a considerably higher (for species and population level, mean estimates of genetic diversity were 0.275 and 0.230, respectively) than the average for other long-lived herbaceous perennials. Indirect estimate of the number of migrants per generation (Nm=1.03, calculated from mean G_ST) was moderate. Factors contributing to the high levels of genetic diversity found within populations ofH. minor include population maintenance via sexual and asexual modes of reproduction, primarily outcrossing breeding system, long generation time, probable ancient polyploid origin of the species, and moderate levels of seed dispersal by wind. Human disturbance in South Korea such as road and apartment constructions appears to be the major threat to this genetically diverse species.