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A collection of 5,659 expressed sequence tags (ESTs) from pineapple [ Ananas comosus (L.) Merr.] was screened for simple sequence repeats (EST-SSRs) with motif lengths between 1 and 6 bp. Lower thresholds of 15, 7 and 5 repeat units were used to define microsatellites of the mono-, di-, and tri- to hexanucleotide repeat type, respectively. Based on these criteria, 696 SSRs were identified among 3,389 EST unigenes, together representing 2,840 kb. This corresponds to an average density of one SSR every 4.1 kb of non-redundant EST sequences. Dinucleotide repeats were most abundant (38.4% of all SSRs) followed by trinucleotide repeats (38.1%). Flanking primer pairs were designed for 537 EST-SSR loci, and 49 of these were screened for their functionality in 12 accessions of A . comosus , 14 accessions of 5 additional Ananas species and 1 species of Pseudananas. Distinct PCR products of the expected size range were obtained with 36 primer pairs. Eighteen loci analyzed in more detail were all polymorphic in pineapple, and primer pairs flanking these loci also generated PCR products from a wide range of genera and species from six subfamilies of the Bromeliaceae. The potential to reveal polymorphism in a heterologous target species was demonstrated in Deuterocohnia brevifolia (subfamily Pitcairnioideae).

Short runs of mononucleotide repeats are present in chloroplast genomes of higher plants. In soybean, rice, and pine, PCR (polymerase chain reaction) with flanking primers has shown that the numbers of A or T residues in such repeats are variable among closely related taxa. Here we describe a set of primers for studying mononucleotide repeat variation in chloroplast DNA of angiosperms where database information is limited. A total of 39 (A)n and (T)n repeats (n greater than or equal to 10) were identified in the tobacco chloroplast genome, and DNA sequences encompassing these 39 regions were aligned with orthologous DNA sequences in the databases. Consensus primer pairs were constructed and used to amplify total genomic DNA from a hierarchical set of angiosperms. All 10 primer pairs generated PCR products from members of the Solanaceae, and 8 of the 10 were also functional in most other angiosperm species. Levels of interspecific polymorphism within the genera Nicotiana, Lycopersicon (both Solanaceae), and Actinidia (Actinidiaceae) proved to be high, while intraspecific variation in Nicotiana tabacum, Lycopersicon esculentum, and Actinidia chinensis was limited. Sequence analysis of PCR products from three primer pairs revealed variable numbers of A, G, and T residues in mononucleotide arrays as the major cause of polymorphism in Actinidia. Our results suggest that universal primers targeted to mononucleotide repeats may serve as general tools to study chloroplast variation in angiosperms.

Premise Microsatellite markers were developed for Xerophyta dasylirioides (Velloziaceae), a species endemic to the Malagasy inselbergs, to explore the impact of its island‐like distribution on genetic diversity and gene flow. Methods and Results A total of 7110 perfect microsatellite loci were recovered by shotgun sequencing on an Illumina MiSeq platform. Primer pairs were designed for 40 arbitrarily selected loci. Fifteen primer pairs that generated distinct PCR products were used to genotype 80 individuals of X. dasylirioides from three inselberg populations. All markers were polymorphic, revealing two to 17 alleles in the overall sampling. Levels of observed and expected heterozygosity per locus ranged from zero to 1.000 and from zero to 0.850, respectively. Success rates of cross‐amplification in 10 additional species of Xerophyta (X. croatii, X. decaryi, X. isaloensis, X. labatii, X. lewisiae, X. pinifolia, X. retinervis, X. setosa, X. spekei, X. tulearensis) ranged from zero to 70%. Conclusions Fifteen newly developed microsatellite markers provide a toolkit for assessing population genetic parameters of X. dasylirioides in its unique island‐like habitats.

The distribution of polyploidy along a relatively steep Andean elevation and climatic gradient is studied using the genus Fosterella L.B. Sm. (Bromeliaceae) as a model system. Ecological differentiation of cytotypes and the link of polyploidy with historical biogeographic processes such as dispersal events and range shift are assessed. 4',6-Diamidino-2-phenylindole (DAPI) staining of nuclei and flow cytometry were used to estimate the ploidy levels of 159 plants from 22 species sampled throughout the distribution range of the genus. Ecological differentiation among ploidy levels was tested by comparing the sets of climatic variables. Ancestral chromosome number reconstruction was carried out on the basis of a previously generated phylogeographic framework. This study represents the first assessment of intrageneric, intraspecific and partially intrapopulational cytotype diversity in a genus of the Bromeliaceae family. In Fosterella , the occurrence of polyploidy was limited to the phylogenetically isolated penduliflora and rusbyi groups. Cytotypes were found to be ecologically differentiated, showing that polyploids preferentially occupy colder habitats with high annual temperature variability (seasonality). The combined effects of biogeographic history and adaptive processes are presumed to have shaped the current cytotype distribution in the genus. The results provide indirect evidence for both adaptive ecological and non-adaptive historical processes that jointly influenced the cytotype distribution in the predominantly Andean genus Fosterella (Bromeliaceae). The results also exemplify the role of polyploidy as an important driver of speciation in a topographically highly structured and thus climatically diverse landscape.

Macaranga (Euphorbiaceae) has received much ecological and evolutionary research attention as a genus that includes some of the most conspicuous pioneer trees of Southeast Asian tropical rainforests and because of its manifold associations with ants, including about 30 species that are obligate ant-plants (myrmecophytes). We used sequence data from three chloroplast DNA loci (ccmp5, ccmp6, atpB-rbcL) to assess phylogeographical patterns in species of Macaranga, section Pruinosae, sampled from various regions of Borneo and the Malay Peninsula. Forty-nine chloroplast DNA haplotypes (HT) were identified among 768 specimens from five species, Macaranga gigantea (N = 329; 23 HT), Macaranga pearsonii (N = 347; 21 HT), Macaranga puberula (N = 24; 4 HT), Macaranga hosei (N = 48; 6 HT), and Macaranga pruinosa (N = 20; 5 HT). Forty-one haplotypes were species-specific, whereas eight haplotypes were shared by two, three, or four species and occupied internal positions in a parsimony network. Population genetic parameters based on haplotype frequencies proved to be in a similar range in the non-myrmecophytic M. gigantea and in the ant-associated M. pearsonii, which have overlapping distributions in northern and eastern Borneo. A comparison of G ST and N ST values revealed a strong phylogeographic structure in both species, whereas colonization pathways suggested by the network topology were different. Both species exhibited similar levels of haplotypic diversity and moderate to high levels of population differentiation. There were no obvious indications for an influence of the symbiotic ant partners on the population structure of their host plants.

The genus Dyckia (Bromeliaceae) comprises 147 species that are distributed in Brazil and adjacent countries. Many species are rare and narrow endemics. We used 454 pyrosequencing to isolate 1,587 microsatellite loci in Dyckia marnier - lapostollei . Of 50 loci that were selected for primer design, 15 markers proved to be polymorphic in five populations from three heterologous species, Dyckia dissitiflora , Dyckia pernambucana and Dyckia limae. Numbers of alleles per locus varied from 3 to 30, and expected and observed heterozygosities ranged from 0.21 to 0.53 and from 0.16 to 0.44, respectively, in the overall sample. The 15 new microsatellite markers are promising tools for studying population genetics in Dyckia species.

Suaeda subg. Brezia (Chenopodiaceae/Amaranthaceae) comprises ~45 halophytic species distributed worldwide along coastlines and in saline inland habitats. Thirteen species are currently accepted from the Americas, but species delimitation is difficult due to the scarcity of distinguishing characters. Little is known yet about phylogenetic relationships and biogeography of American Brezia species. Here, we present molecular phylogenies based on DNA sequence data from the nuclear ribosomal internal transcribed spacer (ITS) and the chloroplast rpl32-trnL intergenic region. Our sampling comprised 157 accessions covering all 13 American Brezia species along with 38 accessions from 16 Eurasian taxa. Phylogenetic trees were generated using parsimony and Bayesian methods. Three monophyletic lineages were discerned in the ITS tree: the Suaeda maritima, S. prostrata and S. corniculata group. Most American species proved to belong to the S. corniculata group. Species boundaries were mostly not recovered or even contradicted by the ITS data, which could be a consequence of low sequence variation in terminal clades and/or reticulate evolution. The rpl32-trnL phylogeny was poorly resolved, with the majority of American species being part of a polytomy with few supported internal nodes. Several incongruities were found between the nuclear and chloroplast tree, revealing at least four instances of hybridization and chloroplast capture between distant lineages. Chromosome counts showed that all American species are polyploid with hexaploidy prevailing. We discuss our results in terms of species relationships, hybridization, polyploidy and biogeography with emphasis on the colonization from NE Asia and Europe, and the subsequent spread and diversification in the Americas.

PREMISE OF THE STUDY: Inselbergs are terrestrial, island‐like rock outcrop environments that present a highly adapted flora. The epilithic bromeliad Encholirium spectabile is a dominant species on inselbergs in the Caatinga of northeastern Brazil. We conducted a population genetic analysis to test whether the substantial phenotypic diversity of E. spectabile could be explained by limited gene flow among populations and to assess the relative impact of pollen vs. seed dispersal on the genetic structure of the species. METHODS: Nuclear and chloroplast microsatellite markers were used to genotype E. spectabile individuals from 20 rock outcrop locations, representing four geographic regions: northern Espinhaço Range, Borborema Plateau, southwestern Caatinga and southeastern Caatinga. F‐statistics, structure, and other tools were applied to evaluate the genetic makeup of populations. KEY RESULTS: Considerable levels of genetic diversity were revealed. Genetic structuring among populations was stronger on the plastid as compared with the nuclear level, indicating higher gene flow via bat pollination as compared with seed dispersal by wind. structure and AMOVA analyses of the nuclear data suggested a high genetic differentiation between two groups, one containing all populations from the southeastern Caatinga and the other one comprising all remaining samples. CONCLUSIONS: The strong genetic differentiation between southeastern Caatinga and the remaining regions may indicate the occurrence of a cryptic species in E. spectabile. The unique genetic composition of each inselberg population suggests in situ conservation as the most appropriate protection measure for this plant lineage.

The genus Dyckia (Bromeliaceae) comprises more than 150 terrestrial or epilithic species with a strongly xeromorphic habit. Most of its members belong to the azonal rock vegetation of Neotropical savannas and forests of Brazil and adjacent countries. Dyckia is relatively species-rich compared with its closest relatives Encholirium (27 species) and Deuterocohnia (17 species). Here, we present the first molecular phylogenetic analysis of Dyckia using DNA sequence data from six plastid loci (matK gene, rps16 intron, petD intron, rpl32-trnL, rps16-trnK and trnD-trnT) and a portion of the nuclear gene phyC. A total of 124 accessions were included, corresponding to 79 taxa from six genera. Phylogenetic trees were generated using parsimony, likelihood and Bayesian methods. DNA sequence variation among Dyckia species turned out to be extremely low, and phylogenies were poorly resolved. The monophyly of Dyckia is supported, whereas evidence is provided that Encholirium is paraphyletic. Based on a dated plastid DNA tree, Dyckia experienced a recent radiation starting around 2.9 million years ago. Four major clades could be identified that roughly correspond to the geographic origin of the samples. A parsimony network based on plastid DNA haplotypes shows a star-like pattern, indicating recent range expansions. Our data are compatible with a scenario where Dyckia and Encholirium diverged in northeastern Brazil, whereas one lineage of Dyckia dispersed to southern Brazil from where a rapid colonization of suitable habitats was initiated. We discuss our results in relation to species delimitation in Dyckia.

• Premise of the study: Central European salt habitats are mainly restricted to the maritime coast but scattered occurrences can also be found inland. In inland habitats, human activities have caused losses and reductions in the size of natural salt sites but have also created new anthropogenic habitats around potash mining dumps colonized by halophytic species in the last 30 yr. We aimed to investigate the effects of bottlenecks, isolation, and ongoing habitat fragmentation on the genetic variation of a species commonly growing in these special habitats.• Methods: We used 10 microsatellite markers to compare genetic diversity and differentiation of 31 populations of Suaeda maritima (Chenopodiaceae) from Central European coasts and inland habitats. Two approaches were applied to analyze the tetraploid data based on allele frequencies directly derived from microsatellite data and from transformed binary data.• Key results: In comparison to the coastal populations from the North Sea and the English Channel, significantly reduced genetic variation and increased between-population differentiation was revealed for populations from the German inland and the Baltic Sea coast. Genetic structure analyses clearly separated coastal and inland populations.• Conclusions: Our results indicate that gene flow is restricted among populations from inland salt sites and the Baltic Sea coast, presumably due to their isolation, small sizes, genetic bottlenecks and/or founder events. Patterns of allele distribution indicate some occasional genetic exchange among habitat types in the past. Anthropogenic salt sites may facilitate gene flow among inland salt habitats preventing endangered inland halophyte populations from genetic erosion.