Flow cytometric and cytogenetic analyses of Iberian Peninsula Festuca spp.

Festuca L. has an important diversification centre in the Iberian Peninsula. We used chromosome counting, fluorescence (FISH) and genomic in situ hybridization (GISH), and DNA flow cytometry (FCM) to clarify the taxonomic position of several taxa, to search for phylogenetic relationships and to assess the extent and pattern of genome variation in fescues. The chromosome number of Festuca duriotagana var. barbata is determined for the first time and new ploidy level estimations are given for F. rothmaleri and F. summilusitana. In the latter species, besides the reported decaploid level, dodecaploidy was found in some populations, which points to the existence of an unrecognized taxon. Moreover, these differences were confirmed by FCM and a high positive correlation was found with the type of substrate where F. summilusitana was growing. For each section, a decrease of genome size with increase of polyploidy was observed. In general, in situ hybridization techniques failed to reveal phylogenetic relationships among the selected species. In FISH, a variation in the number of rDNA sites was observed in some species. GISH results indicate that F. henriquesii is not a progenitor of the studied polyploid species.     

A new species of Haplophyllum A. Juss. (Rutaceae) from the Iberian Peninsula: evidence from morphological, karyological and molecular analyses

BACKGROUND AND AIMS: The discovery of a new species, Haplophyllum bastetanum F.B. Navarro, V.N. Suárez-Santiago & Blanca sp. nov., in the south-east of Spain has prompted the comparative study of species of the Iberian Peninsula, and others related, through morphological, cytogenetic, molecular, distributional and ecological characterization. METHODS: The morphological study involved a quantitative analysis of the species present in the Iberian Peninsula and a comparative analysis of the morphological characteristics between H. bastetanum and other related species. Mitotic analyses were made with root meristems taken from germinating seeds. Phylogenetic analyses of the internal transcribed spacer sequences of nuclear ribosomal DNA were performed using neighbour-joining (NJ) and maximum-parsimony methods. KEY RESULTS: Haplophyllum bastetanum is a diploid species (2n = 18) distinguished primarily for its non-trifoliate glabrous leaves, lanceolate sepals, dark-green petals with a dorsal band of hairs, and a highly hairy ovary with round-apex locules. The other two Iberian species (H. linifolium and H. rosmarinifolium) are tetraploid (2n = 36) and have yellow petals. Both phylogenetic methods generated a well-supported clade grouping H. linifolium with H. rosmarinifolium. In the NJ tree, the H. linifolium-H. rosmarinifolium clade is a sister group to H. bastetanum, while in the parsimony analysis this occurred only when the gaps were coded as a fifth base and the characters were reweighted according to the rescaled consistency index. This latter group is supported by the sequence divergence among taxa. CONCLUSIONS: The phylogenies established from DNA sequences together with morphological and cytogenetic analyses support the separation of H. bastetanum as a new species. The results suggest that the change in the number of chromosomes may be the key mechanism of speciation of the genus Haplophyllum in the Iberian Peninsula. An evolutionary scheme for them is propounded.     

Multivariate and cladistic analyses of the purple-flowered species ofErysimum (Cruciferae) from the Iberian Peninsula

A taxonomically difficult purple-flowered group within the genusErysimum, restricted to the Iberian Peninsula, is analyzed by multivariate and cladistic analyses. 51 specimens have been scored for 14 characters. Both principal components and discriminant analyses provide support to the recognition of the five species considered by the author, namelyE. linifolium, E. lagascae, E. baeticum, E. popovii, andE. cazorlense. Cladistic analysis, using 7 characters resulted in a single most parsimonious cladogram containing no homoplasies. The pattern of morphologic divergence follows a clear NW.-SE. trend, which is congruent with the topology of the cladogram. This trend significantly affects growth-form as well as fruit characters, both providing the main grounds for species recognition. The different behavior and significance of several characters in both kinds of analysis is discussed. The co-occurrence of morphologically similar individuals differing in the flower color is discussed, too. Possible explanations for this phenomenon involve hybridization in a wide sense or, alternatively, rejecting the assumption of monophyly for the group.     

Karyology of theScorzonera (Compositae) species from the Iberian Peninsula

A karyological study of 15 taxa ofScorzonera L. from the Iberian Peninsula has been made. The chromosome numbers found inS. hispanica var.pinnatifida, S. baetica, S. reverchonii, S. angustifolia, S. laciniata var.calcitrapifolia and var.subulata (2n = 14) are new. Diploid cytotypes with 2n = 14 and 2n = 12 prevail, andS. hispanica var.crispatula is the only taxon which exhibits autopolyploidy (2n = 14, 28). x = 7 is considered to be the base chromosome number within the genus, with x = 6 being derived from it by translocation. This and detailed karyotype analyses allow to group the Iberian Peninsula species ofScorzonera into three groups.     

A new species of Muscari subgenus Leopoldia from the Iberian Peninsula

The biological analysis of several populations confirms the existence in the Iberian Peninsula of two species of Muscari subgenus Leopoldia: M. comosum and M. matritensis (the latter described as new). Karyologically, they differ in at least a translocation rearrangement, in the presence of heterochromatin, which is lacking in M. matritensis, and in some chromosomal characters. At the genic level, the distance between M. comosum and M. matritensis is high (0.378). Morphologically, they show some differences in floral structure which are correlated with different reproductive systems, allogamy in M. comosum and autogamy in M. matritensis. The possible evolutionary relationships between these taxa is discussed.     

Behaviour of pollinator insects within inflorescences of Scrophularia species from Iberian Peninsula

Several hypotheses have been proposed to explain the pattern of pollinator visits to vertical inflorescences. These range from a response to a pattern of resources to merely instinctive behaviour. In dichogamous plants, such behaviour has been associated with promoting outcrossing and avoiding geitonogamy. We here analyse behaviour of the principal pollinator groups in five protogynous species of Scrophularia with different flower sizes (S. sambucifolia, S. grandiflora, S. lyrata, S. scorodonia and S. canina), and the distribution of sexual phases along the inflorescences. The results in all cases show that pollinators follow a pattern of ascending visits accompanied by movements between flowers of the same whorl (horizontal movements). The relative frequency of these horizontal movements depends on the flower size, with a higher frequency in species with large flowers. In vertical movements of the three more common pollinator groups to several plant species (bumblebees, wasps and small bees), the behaviour was essentially independent of flower size, with bumblebees having the highest ratio of ascents to descents. Behaviour of the pollinators, together with the absence of a definite pattern of distribution of the sexual phases along the inflorescence, implies that geitonogamy is not avoided in any of the Scrophularia species studied.     

Two new species of Galium (Rubiaceae) from the Iberian Peninsula

Two new species of the genus Galium are described from the Iberian Peninsula: G. moralesianum Ortega-Olivencia & Devesa, and G. talaveranum Ortega-Olivencia & Devesa. The first grows on limestone or dolomitic-limestone outcrops in south-east Spain (the Segura and Gádor mountain ranges) and shows morphological similarities with G. boissieranum Ehrend. & Krendl, which is endemic to southern Spain (mountains of Málaga). The second inhabits the margins of water courses, alder stands and cork-oak dehesas (parkland-type systems) of the west of the Iberian Peninsula, and is morphologically reminiscent of G. mollugo L. The chromosome numbers of these two species and of G. boissieranum are given.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 143 , 177−187.     

The non-omphalinoid species of Arrhenia in the Iberian Peninsula

A taxonomic study of the species with nutant, pleurotoid and cyphelloid basidiomata of the genus Arrhenia in the Iberian Peninsula is presented. This study is based on the examination of recent specimens collected in the field and from dried herbarium collections. Five species and one variety of this genus are recognized on the basis of morphological and ecological features: Arrhenia acerosa var. acerosa, A. acerosa var. tenella, A. auriscalpium, A. lobata, A. retiruga and A. spathulata. Arrhenia auriscalpium is new to Spain, where it is not restricted to alpine zones of the Eurosiberian region and is more common in the Mediterranean region than previously reported. In contrast, A. acerosa var. tenella currently is known from the Eurosiberian subalpine belt and represents the first report to the Iberian Peninsula, and A. retiruga is known only from the mesomediterranean to supramediterranean belts of the Mediterranean region. Arrhenia acerosa var. acerosa, A. lobata and A. spathulata are widespread in both the Eurosiberian and the Mediterranean regions. Non-Iberian materials of A. acerosa var. tenella and A. spathulata also were studied for comparison. Cyphella cochlearis var. subsessilis, Dictyolus lagunae, Leptoglossum muscigenum var. azonum and Pleurotellus acerosus var. tenellus are lectotypified. Cyphella cochlearis var. subssesilis, Dictyolus lagunae and Leptoglossum muscigenum var. azonum are synonymized with Arrhenia spathulata, and Cyphella cochlearis var. auriformis is placed in synonymy with A. auriscalpium. These taxa are illustrated, described and discussed, based on Iberian material with emphasis on features of the basidioma, pigment of the pileipellis, presence or absence of clamps and shape and size of the basidiospores. A diagnostic key also is given.     

Flow cytometric and karyological analysis of polysomaty and polyploidization during callus formation from leaf segments of various potato genotypes

Summary Flow cytometry and karyological analysis were used to study polysomaty and polyploidization during the first 15 days of callus formation in leaf segments from shoot cultures and greenhouse-grown plants of various lines and genotypes of Solanum tuberosum and S. phureja. The greenhouse-grown plants showed a higher degree of polysomaty (77% and 49% of polyploidized nuclei) than the shoot cultures (< 3%). During the in vitro culture period, polyploidization occurred through endoreduplication. Segments of the five shoot cultures showed up to 87%, 53%, 59%, 45% and 56% polyploidization, respectively; the DNA content of corresponding interphase nuclei amounted to 8C, 16C, 16C, 16C and 8C, and the chromosome numbers to 96. Segments from the two greenhouse-grown genotypes showed up to 87% and 84% polyploidization; the DNA content amounted to 32C and 16C, and the chromosome numbers to 192 and 96. The number of reduplication cycles was species-dependent; the degree of polyploidization was dependent on the initial ploidy level of the genotypes. Cell proliferation did not take place at a constant rate. The maximum frequencies of metaphases (52–171 per segment) occurred after 1 week of culture and were correlated with the ploidy level of the genotypes. Cells were triggered to mitosis rather than to endoreduplication. Cell cycles with normal monochromosomes could be shorter than 1 day, and those with diplochromosomes lasted at least 1 day. Polysomaty, degree of polyploidization and abnormal nuclear processes are discussed in relation to the origin of genetic instability early in culture.     

Two new sorediate species of lichens in the Catillariaceae from the Iberian Peninsula

Abstract:Two new species of lichens are described from the Iberian Peninsula. Halecania giraltiae is the first sorediate saxicolous species known in the genus and has been collected growing on lichens and sometimes directly on rock in localities from northern Portugal to central Spain. Catillaria fungoides, a sorediate, corticolous representative of the C. nigroclavata group, is known from southern Portugal and northern Spain.     

Flow cytometric and cytogenetic analyses in human spontaneous abortions

Cytogenetic and flow cytometric analyses were performed on 38 human spontaneous abortions in an attempt to obtain information on karyotype abnormalities and to compare the two approaches of analysis. In 19 cases, it was not possible to perform cytogenetic analysis because too long a time had passed between surgical sampling and cell culture, and in vitro culture failed. Of the 19 cases analyzed, 10/19 showed a normal karyotype and 5/19 showed a single trisomy (2/5 trisomies involved chromosome 16, 1/5 trisomy involved chromosome 18, 1/5 trisomy involved chromosome 20, and 1/5 was Klinefelter syndrome). Of the remaining 4/19 cases, 2/19 showed a polyploid condition (1 tetraploidy and 1 triploidy), 1/19 a double trisomy (chromosomes 13 and 21), and 1/19 a pentasomy of the sex chromosomes (49,XXXXY). Flow cytometric analysis was performed on all abortive samples. The samples were subdivided, when possible, into two portions conventionally named amniotic and chorionic, using the amniotic membrane as an anatomical reference. Maternal blood lymphocytes were used as a diploid standard for each sample. In the 19 cases not analyzed by the cytogenetic approach, flow cytometric analysis showed 9 diploid and 10 aneuploid DNA distributions. In the remaining 19 cases, analyzed with both approaches, the comparison of DNA estimations using cytogenetic and flow cytometric analyses showed good agreement. In the cases with karyotype abnormalities, flow cytometric measurement provided evidence of an alteration of DNA content with respect to the diploid standard. Flow cytometric analysis showed a diploid distribution, whereas cytogenetic analysis revealed chromosomal abnormalities in only 4/19 cases. These discordant results could be related to mosaic conditions or maternal cell contamination. Moreover, cytogenetic and flow cytometric analyses were performed on 2 amniotic cell cultures, and concordant results were obtained. The results obtained suggest that a combination of these techniques is beneficial in attempts to obtain information about DNA content alterations, even when cultures fail, and in screening studies of human abortions.     

Rinodina turfaceoides,a new corticolous,blastidiate species from the Iberian Peninsula

Abstract:A new corticolous lichen, Rinodina turfaceoides, is described from the Serra de Estrela (Portugal). It is characterized mainly by the blastidiate thallus, large apothecia covered by a thick, yellow-orange pruina, large Physcia -type ascospores and chemistry. The new species is compared with the closely related but mainly muscicolous, R. turfacea, as well as with other corticolous Rinodina species with vegetative propagules, including R. colobinoides and R. disjuncta. Habit photographs of the new species and illustrations of its ascospores are also provided.     

Flow cytometric comparison of haemocytes from three species of bivalve molluscs

Haemocyte subpopulations from three bivalve species (the clams Ruditapes philippinarum and Mercenaria mercenaria and the oyster, Crassostrea virginica) were characterised using light-scatter flow cytometry and a standard set of methods. Two parameter (forward and side scatter) plots for the three species were very similar and resembled plots for mammalian white blood cells. Two haemocyte groups (granulocytes and agranulocytes) were found in both the haemolymph and the extrapallial fluid of the clams while those two groups and an additional third group were found in the haemolymph of the oyster. All subpopulations were sorted on to glass slides, identified, photographed, and measured microscopically. Sorting of the bivalve granulocyte and agranulocyte groups indicated varying degrees of heterogeneity within each population in terms of either size or granularity, or both. However, subsorting of selected regions within the major groupings produced highly pure haemocyte populations. The comparison showed both similarities and differences among species. For instance, a distinct subpopulation of small granulocytes was present only in oysters and a subpopulation of spindle-shaped haemocytes, only in M. mercenaria. The haemocyte subpopulations delineated by light-scatter flow cytometry underscore questions about cell lineages, but the instrument also offers a powerful technique for answering them.     

Scilla merinoi (Liliaceae), a new species from Galicia, northwestern Iberian Peninsula

A new species, Scilla merinoi , from Galicia is described and illustrated. The similarities between the new taxon and two related species, S. odorata and S. verna , are discussed. Together with a morphological comparison of the three species, scanning electron micrographs of seeds and a map of the new species' known distribution are presented.     

Contributions to the karyological study of the genus Ranunculus L. subgenus Batrachium (DC.) A Gray from the Iberian Peninsula

DIOSDADO. J. C, PASTOR, J. E. & VALDÉS, B. 1993. Contributions to the karyological study of the genus Ranunculus L. subgenus Batrachium (DC.) A. Gray from the Iberian Peninsula. The somatic and gametic chromosome numbers and detailed chromosome morphology are presented for eight taxa of aquatic ranunculi from the Iberian Peninsula. Within the subgenus Batrachium diploid and tetraploid levels have been observed (2 n = 16, 32) the commoner of which is the diploid. From the karyological data relationships have been established between the subgenus Batrachium and subgenus Ranunculus.     

Karyology of Limonium (Plumbaginaceae) species from the Balearic Islands and the western Iberian Peninsula

Somatic chromosome numbers, conventional karyotype features and idiograms are reported for 27 Limonium species inhabiting the Western Mediterranean basin (Iberian Peninsula and the Balearic Islands). The chromosome numbers of Limonium barceloi (2 n  = 36), L. ejulabilis (2 n  = 24), L. inexpectans (2 n  = 26), L. interjectum (2 n  = 24), and L. scopulorum (2 n  = 25) were determined for the first time. In addition, new aneuploid and/or polyploid cytotypes are reported in L. alcudianum (2 n  = 26), L. bonafei (2 n  = 26), L. camposanum (2 n  = 26) , L. companyonis (2 n  = 26), L. dufourii (2 n  = 26), L. geronense (2 n  = 36), L. marisolii (2n = 54), L. migjornense (2 n  = 50), and L. pseudodictyocladon (2 n  = 16). A group of polyploid species showed karyotypes comprising homologous chromosomes in groups of three ( L. antonii-llorensii, L. ejulabilis, L. interjectum, L. virgatum, and L. wiedmanii ) , four ( L. geronense ), or six ( L. marisolii ), which suggests an autopolyploid origin. Other polyploid species were characterized by the presence of two different chromosome sets ( x  = 8 and x  = 9) in the genome. The species L. alcudianum , L . bonafei , L. camposanum , L. companyonis , L. dufourii , L. gibertii , L. girardianum , L. inexpectans , L. leonardi-llorensii , L. magallufianum , L. migjornense , L. minoricense , and L. scopulorum showed various combinations of paired and unpaired x  = 8 and x  = 9 chromosome sets, suggesting that they are allopolyploids. © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 257–272.     

Lichenized species of Omphalina (Tricholomataceae) in the Iberian Peninsula

Abstract:A taxonomic revision of the lichenized species of the genus Omphalina in the Iberian Peninsula is presented, based upon fungal and lichen herbarium material, as well as on fresh collections. Relevant morphological and ecological features are discussed and a diagnostic key is provided. Four lichenized species, O. ericetorum, O. hudsoniana, O. meridionalis and O. velutina, are recognized. A squamulose vegetative thallus (Coriscium -type) is present only in O. hudsoniana, while the other three species form a thallus of hyphal globules (Botrydina -type). Anatomical features of the vegetative thallus cannot be used to distinguish between these last three species. Whilst O. hudsoniana is known only from the montane belt of the Eurosiberian Region and O. meridionalis seems to be restricted to the Mediterranean Region, living in more or less humid sites of the Supramediterranean belt, the other two species, O. ericetorum and O. velutina, are widespread.     

Indicator species of broad-leaved oak forests in the eastern Iberian Peninsula

The degraded state, scattered distribution and substrate type of Quercus pyrenaica forests in eastern Spain make them a subject of particular interest, as well as a priority for conservation efforts. An extension of Dufrêne and Legendre's indicator value (IndVal) analysis was applied to one hundred relevés. This extension of the IndVal method allowed us, by considering the preference of species for combinations of relevé groups, to obtain those species that represent local, but also regional, characteristics (as a new practical use). Our objective was to identify the species that best indicate the main ecological features of these broad-leaved oak forests. The usefulness of our analysis was evidenced at two levels: local indicator species, showing the characteristic species of individual sites and including the presence of endemics in the area; and indicator species of combinations of two or more sites, reflecting the affinities or relationships between these scattered forests.Most singular forests showed a higher number of local indicator species. The indicator species of each site combination represent the main ecological traits shared by these groups – such as proximity to the coast, land uses or postglacial migration pathways – thereby defining their relationships. Indicator value analysis with site group combinations (Multipatt function) proved to be a useful tool for the identification of different species that could serve both to determine the local forest conservation status and aid in its preservation, as well as contributing to an understanding of the life history of larger territories.     

Flow cytometric DNA analyses of epithelial dysplasia of the esophagus

OBJECTIVE: To investigate, with flow cytometry, DNA aneuploidy as a marker of early carcinogenesis in dysplastic esophageal lesions. STUDY DESIGN: DNA content of exfoliated cells from 789 cases of esophageal dysplasia (including mild dysplasia, 195 cases; moderate dysplasia, 383 cases; and severe dysplasia, 211 cases) was determined with a FACS 420 flow cytometer. RESULTS: Cellular DNA content was closely related to the severity of dysplasia. The carcinogenesis rate in patients with dysplasia showed that DNA aneuploidy was significantly higher than in patients showing DNA diploidy. CONCLUSION: DNA aneuploidy in dysplastic lesions is a very important early signal of carcinogenesis. Patients with dysplastic lesions showing DNA aneuploidy should be treated and closely followed.     

Floral rewards in someScrophularia species (Scrophulariaceae) from the Iberian Peninsula and the Balearic Islands

The genusScrophularia in the Iberian Peninsula and Balearic Islands comprises two sections,Scrophularia andCanina G. Don. Analyses were carried out on flower production, flower duration and their sexual phases, pollen and nectar production together with observations on their pollinators. Nectar production is correlated with corolla size and pollen production with anther size. The taxa of sect.Scrophularia show greater nectar and pollen production than those of sect.Canina. Also, those of the first section produce more ovules per ovary than those of the second group, production being correlated with the ovary size.Some observations on floral and reproductive biology in some species ofScrophularia from the Iberian Peninsula and the Balearic Islands. I.