Víctor N. Suárez-Santiago

Phylogeny of the Centaurea group (Centaurea, Compositae) - geography is a better predictor than morphology.

The Centaurea group is part of the Circum-Mediterranean Clade (CMC) of genus Centaurea subgenus Centaurea, a mainly Mediterranean plant group with more than 200 described species. The group is traditionally split on morphological basis into three sections: Centaurea, Phalolepis and Willkommia. This division, however, is doubtful, especially in light of molecular approaches. In this study we try to resolve this phylogenetic problem and to consolidate the circumscription and delimitation of the entire group against other closely related groups. We analyzed nuclear (internal transcribed spacer of the ribosomal genes) and chloroplast (rpl32-trnL intergenic spacer) DNA regions for most of the described species of the Centaurea group using phylogenetic and network approaches, and we checked the data for recombination. Phylogeny was used to reconstruct the evolution of the lacerate-membranaceous bract appendages using parsimony. The magnitude of incomplete lineage sorting was tested estimating the effective population sizes. Molecular dating was performed using a Bayesian approach, and the ancestral area reconstruction was conducted using the Dispersal-Extinction-Cladogenesis method. Monophyly of the Centaurea group is confirmed if a few species are removed. Our results do not support the traditional sectional division. There is a high incongruence between the two markers and between genetic data and morphology. However, there is a clear relation between geography and the structure of the molecular data. Diversification in the Centaurea group mainly took place during the Pliocene and Pleistocene. The ancestral area infered for the Circum-Mediterranean Clade of Centaurea is the Eastern Mediterranean, whereas for the Centaurea group it is most likely NW-Africa. The large incongruencies, which hamper phylogenetic reconstruction, are probably the result of introgression, even though the presence of incomplete lineage sorting as an additional factor cannot be ruled out. Convergent evolution of morphological traits may have led to incongruence between morphology-based, traditional systematics and molecular results. Our results also cast major doubts about current species delimitation.

Study on Cortinarius subgenus Telamonia section Hydrocybe in Europe, with especial emphasis on Mediterranean taxa.

In this paper we have attempted to clarify the taxonomy and nomenclature of thirteen taxa of the genus Cortinarius subgenus Telamonia (sections Hydrocybe, Fraternii) well represented in the southwestern Mediterranean area of Europe (C. atrocoeruleus, C. bombycinus, C casimiri, C. contrarius, C. decipiens, C. fraternus, C. gallurae, C. hoffmannii, C. petroselineus, C. sertipes, C. subturibulosus, C. urdaibaiensis and C. vernus). To this end we have performed a combined study of morphological and molecular data (rDNA ITS sequences). The morphological analysis was carried out on 114 collections and the molecular analysis involved 31 of the 114 collections, including 11 type collections (types for C. casimiri and C. fraternus were not available). In addition, a study of spores under field emission scanning electron microscopy (FESEM) was conducted. The results of the combined analysis allowed us to asign the studied material to five species (C. casimiri s.l., C. decipiens s.l., C. gallurae, C. subturibulosus s.l. and C. vernus s.l.). Thus, all collections from more continental areas, which were originally identified as six different taxa (C. atrocoeruleus, C. contrarius, C. decipiens, C. fraternus, C. sertipes, C. flexipes fo. sertipes) corresponded to C. decipiens sensu lato, a widely distributed, genetically and morphologically variable species. Cortinarius casimiri is also found in such habitats, but it is confirmed as distinct taxon. Collections from Mediterranean sclerophyllous communities correspond to C. gallurae, C. vernus sensu lato and C. subturibulosus sensu lato. Due to close phylogenetic relationships we propose the new combinations C. casimiri var. hoffmannii (=C. decipiens var. hoffmannii non C. hoffmannii) and C. subturibulosus var. bombycinus (=C. bombycinus), and the new variety C. vernus var. nevadavernus (=C. vernus H. Lindstr. & Melot sensu auct.).

Evolutionary history of fumitories (subfamily Fumarioideae, Papaveraceae): An old story shaped by the main geological and climatic events in the Northern Hemisphere

Fumitories (subfamily Fumarioideae, Papaveraceae) represent, by their wide mainly northern temperate distribution (also present in South Africa) a suitable plant group to use as a model system for studying biogeographical links between floristic regions of the Northern Hemisphere and also the Southern Hemisphere Cape region. However, the phylogeny of the entire Fumarioideae subfamily is not totally known. In this work, we infer a molecular phylogeny of Fumarioideae, which we use to interpret the biogeographical patterns in the subfamily and to establish biogeographical links between floristic regions, such as those suggested by its different inter- and intra-continental disjunctions. The tribe Hypecoeae is the sister group of tribe Fumarieae, this latter holding a basal grade of monotypic or few-species genera with bisymmetric flowers, and a core group, Core Fumarieae, of more specious rich genera with zygomorphic flowers. The biogeographical analysis shows a subfamily that originated in East Asia at the end of the Early Cretaceous. From here, ancestral range expansions followed three different directions, one at the beginning of the Late Cretaceous by the ancestor of tribe Hypecoeae towards central Asia, and two during the Cretaceous-Palaeogene transition towards western North America and Indochina by the ancestor of the tribe Fumarieae. The ancestor of Core Fumarieae expanded its range from East Asia into the Himalayas before to the middle Eocene. The uplifts of the Qinghai-Tibetan Plateau together with the zonal climate pattern of the Palaeogene are suggested to be responsible both for the accelerated diversification rate resulting in the origin of the basal lineages of Core Fumarieae as well as for the westward migration of the ancestor of Fumarieae s.str. into the Irano-Turanian region. From here, this latter group reached South Africa during late Eocene and Mediterranean basin during Oligocene. There were two colonization waves of the Mediterranean following two different routes: a northern route during the early Oligocene by the subtribe Sarcocapninae, probably facilitated by the land bridge resulting of the Mediterranean microplate accretion; and a southern route into North Africa, through the Gomphotherium land bridge, taken by the subtribe Fumariinae between late Oligocene and middle Miocene.

Phylogeny of the tribe Fumarieae (Papaveraceae s.l.) based on chloroplast and nuclear DNA sequences: Evolutionary and biogeographic implications

PREMISE OF THE STUDY Little research has been done at the molecular level on the tribe Fumarieae (Papaveraceae). Papaveraceae is a model plant group for studying evolutionary patterns despite the lack of a reference phylogeny for this tribe. We investigated the phylogenetic relationships within the tribe to complete the molecular data for this family in order to help understand its character evolution and biogeographic pattern. METHODS We used maximum-parsimony and Bayesian approaches to analyze five DNA regions for 25 species representing 10 of the 11 Fumarieae genera and five outgroups. Evolutionary pathways of four characters (habit, life span, type of fruit, and number of seeds per fruit) were inferred on the phylogeny using parsimony. The ancestral distribution areas were reconstructed using dispersal-vicariance analysis. KEY RESULTS Fumarieae is monophyletic and includes three groups that agree with the morphology-based subtribes: Discocapninae, Fumariinae, and Sarcocapninae. Within subtribes, the relationships among genera were different from those obtained with morphological data. Annual life span, nonchasmophytic habit, and a several-seeded capsule were the basal character states for the tribe. The ancestor occupied a continuous area between West Eurasia and Africa. Vicariances explain the divergence between lineages Discocapninae (South Africa) and Fumariinae-Sarcocapninae (Mediterranean), and the disjunction of Fumariinae (Mediterranean-Central Asia). CONCLUSIONS Molecular phylogeny confirms the subtribal classification of Fumarieae based on morphology. However it provides different results regarding the relationships among genera within each subtribe, which affects the inference of the evolutionary pathway followed by the four selected characters. The disjunct distribution of the tribe is explained by different vicariance scenarios.

Polyploidy, the major speciation mechanism in Muscari subgenus Botryanthus in the Iberian Peninsula

Currently, three species ofMuscari subg. Botryanthus are recognized in the Iberian Peninsula: two diploids (2n = 18), M. atlanticum and M. cazorlanum, and one morphologically variable species with three different ploidy levels, M. neglectum (2n = 36, 45, 54). We have made a comparative study of numerous Iberian populations to clarify the taxonomy and evolution of this group. To this end we carried out morphological and cytogenetic analyses, and phylogenetic analysis of the internal transcribed spacers of nuclear ribosomal DNA. Comparative and UPGMA analyses of the morphological characteristics show that the different ploidy levels of M. neglectum represent different species. We describe the pentaploid and hexaploid levels as two new species, M. olivetorum (2n = 45) and M. baeticum (2n = 54), each with an exclusive combination of morphological characters and a characteristic ecological behavior pattern. Phylogenetic study of ITS shows that the two new species are not autopolyploids from M. neglectum but allopolyploids. These findings are supported by the additivity of the three ITS variants found in M. olivetorum with the ITS of M. neglectum and M. baeticum, and also by morphology. Possible parents for both new species are proposed. Absence of homogenization between homeologous M olivetorum nrDNA loci is explained by the absence of sexual reproduction and by nucleolar dominance, indicating that this is a recent species.

Tragopogon lainzii , a New Species of Tragopogon (Asteraceae) Segregated from T. dubius : Evidence from Morphological and Molecular Data

Abstract Tragopogon dubius is one of the most widespread species of Tragopogon, extending across much of Eurasia. Traditionally, T. dubius has been considered a morphologically homogeneous species that includes all Tragopogon collections with yellow flowers and swollen peduncles under capitula. Here we describe a new species of Tragopogon from the Iberian Peninsula, T. lainzii, which has heretofore been included in T. dubius. To this end, we performed comparative morphological, cytogenetic, and molecular analyses on many populations of both species. Our results show that T. dubius is not a homogeneous species and that different lineages exist across its broad geographic distribution. Moreover, we show that hybridization has occurred in the wild between sympatric populations of T. dubius, T. lainzii, and T. porrifolius.

Differential spreading of HinfI satellite DNA variants during radiation in Centaureinae

BACKGROUND AND AIMS Subtribe Centaureinae appears to be an excellent model group in which to analyse satellite DNA and assess the influence that the biology and/or the evolution of different lineages have had on the evolution of this class of repetitive DNA. Phylogenetic analyses of Centaureinae support two main phases of radiation, leading to two major groups of genera of different ages. Furthermore, different modes of evolution are observed in different lineages, reflected by morphology and DNA sequences. METHODS The sequences of 502 repeat units of the HinfI satellite DNA family from 38 species belonging to ten genera of Centaureinae were isolated and compared. A phylogenetic reconstruction was carried out by maximum likelihood and Bayesian inference. KEY RESULTS Up to eight different HinfI subfamilies were found, based on the presence of a set of diagnostic positions given by a specific mutation shared by all the sequences of one group. Subfamilies V-VIII were mostly found in older genera (first phase of radiation in the subtribe, late Oligocene-Miocene), although some copies of these types of repeats were also found in some species of the derived genera. Subfamilies I-IV spread mostly in species of the derived clade (second phase of radiation, Pliocene to Pleistocene), although repeats of these subfamilies exist in older species. Phylogenetic trees did not group the repeats by taxonomic affinity, but sequences were grouped by subfamily provenance. Concerted evolution was observed in HinfI subfamilies spread in older genera, whereas no genetic differentiation was found between species, and several subfamilies even coexist within the same species, in recently radiated groups or in groups with a history of recurrent hybridization of lineages. CONCLUSIONS The results suggest that the eight HinfI subfamilies were present in the common ancestor of Centaureinae and that each spread differentially in different genera during the two main phases of radiation following the library model of satellite DNA evolution. Additionally, differential speciation pathways gave rise to differential patterns of sequence evolution in different lineages. Thus, the evolutionary history of each group of Centaureinae is reflected in HinfI satellite DNA evolution. The data reinforce the value of satellite DNA sequences as markers of evolutionary processes.

Multilamellar bodies linked to two active plasmalemma regions in the pollen grains of Sarcocapnos pulcherrima

The presence of visible multilamellar bodies in the cytoplasm of pollen grains of at least seven species of the family Papaveraceae has led us to study the behaviour of these bodies during pollen-grain ontogeny and in growing pollen tubes of Sarcocapnos pulcherrima C. Morales & R. Garcia germinated in vitro. Our transmission-electron-microscope (TEM) studies in pollen grains show that the multilamellar bodies may be classified as: 1) small, isolated and placed in the region of apertures in the cytoplasm; and 2) large, in clusters and in contact with the active plasmalemma apertures only when tubules are being formed in the apertural intine. Similar types of multilamellar bodies to those observed in the pollen apertures can be seen near the apex of the growing pollen tube (small and isolated) and in contact with the apex plasmalemma (large and clustered). Our results support the hypothesis that the multilamellar bodies are functionally linked to moments when the cytoplasmic membrane is very active. We have also linked the multilamellar bodies to Golgi vesicles as they both react positively to acid-phosphatase (AP) staining and also to the plasmalemma by the thiocarbohydrazide-silver proteinate-staining (TCH-Sp) electron-contrasting technique.

Satellite-DNA diversification and the evolution of major lineages in Cardueae (Carduoideae Asteraceae)

In a previous work, we characterized the HinfI satellite DNA family in the subtribe Centaureinae (Cardueae) demonstrating that a “library” of eight HinfI subfamilies would exist in the common ancestor of all Centaureinae, which were differentially amplified in different lineages. Now, we extend our study by analyzing a total of 219 additional repeats from fifteen species belonging to Carlininae, Echinopsinae and Carduinae, and comparing them to those of Centaureinae. Most HinfI sequences belonged to the subfamily II, although a few sequences of other subfamilies were detected in some species. Additionally, a new subfamily characteristic of several Carduinae species was discovered. Although phylogenetic trees grouped sequences by subfamily affinity instead of species provenance, when comparing repeats of the same subfamily, the degree of divergence between any pair of sequences was related to the evolutionary distance between the species compared in most cases. Exceptions were in comparisons between sequences of some Centaureinae species, and between sequences of some Carduinae species and those of Centaureinae. Our results demonstrate that: (1) At least nine HinfI subfamilies would exist in the common ancestor of Cardueae, each one differentially amplified in different lineages; (2) After differential spreading, sequences of each subfamily evolved concertedly through molecular drive, resulting in the gradual divergence of repeats between different species; (3) The rate to which concerted evolution occurred was different between lineages according to the evolutionary history of each one.

Isolation and characterization of eight polymorphic microsatellite loci for the critically endangered Arenaria nevadensis (Caryophyllaceae)

We report on the isolation and characterization of eight microsatellite markers from enriched libraries for Arenaria nevadensis, one of the most critically endangered plant species in the Iberian Peninsula. These are the first microsatellite loci reported for Arenaria species. The number of alleles ranged from two to eight, and the expected heterozygosity from 0.067 to 0.873. These markers will be useful for characterizing the genetic diversity in A. nevadensis and understanding its population structure, and will provide important genetic data for the conservation and recovery of this species.