Pollen morphology of some taxa of Vicia L. subgenus Vicilla (Schur) Rouy (Fabaceae) from Turkey

The pollen morphology of the ten taxa belonging to Cassubicae Radzhi, Cracca Dumort., Ervilia (Link) Koch, Lentopsis Kupicha, Trigonellopsis Rech. f. and Variegatae Radzhi sections of the genus Vicia L. subgenus Vicilla (Schur) Rouy (Fabeae, Fabaceae) in Turkey has been examined by use of light microscopy and scanning electron microscopy. Twelve morphometric characters are analysed with one-way analysis of variance and Tukey’s honestly significant difference test for multiple comparisons. Polar axis and equatorial diameter of pollen grains range from 24.64 to 44.19 and from 15.90 to 30.42 μm, respectively. The pollen grains are prolate–spheroidal to perprolate (P/E = 1.09–2.07), but the prolate shape occurs in the majority of the taxa. The regular pollen grains of all taxa are trizonocolporate, isopolar, and released in monads. Ornamentation of the mesocolpium is psilate–perforate in V. tenuifolia subsp. dalmatica and V. tenuifolia subsp. tenuifolia (sect. Cracca), reticulate–rugulate in V. villosa subsp. villosa (sect. Cracca), and reticulate–perforate in the remaining taxa. The apocolpium and colpus area are reticulate–perforate in V. hirsuta (sect. Cracca), V. ervilia (sect. Ervilia) and V. lunata subsp. grandiflora (sect. Trigonellopsis), and psilate, psilate–slightly perforate or perforate in the other taxa. The results obtained from analyses of pollen grains from the studied taxa of the subgenus Vicilla reveal significant differences in general morphology and some morphometric attributes, and ornamentation types. However, it is concluded that pollen morphology has proved to be uninformative for differentiating the subgenera Vicilla and Vicia.     

Phylogenetic relationships between and within sections Hypechusa, Narbonensis and Peregrinae of genus Vicia (Fabaceae) based on evidence from isozymes and morphology

Previous studies of vetch species belonging to sections Hypechusa, Narbonensis and Peregrinae have revealed controversies and open questions that deserve revisiting with the use of contemporary approaches. The paper extends the previous work of Leht and Jaaska (2002) by describing variation of 16 isozymes and 86 morphological characters among 17 Vicia species belonging to sections Hypechusa, Narbonensis and Peregrinae of the type subgenus in comparison with V. bithynica of section Pseudolathyrus. Three more isozymes and 12 new morphological characters are included to improve the resolution of phylogenetic relationships. The maximum parsimony trees based on isozymes and morphology revealed essentially the same four clusters of species that correspond to series Hypechusa and Hyrcanicae of section Hypechusa, to sections Narbonensis and Peregrinae of current taxonomy, except that the isozyme tree placed V. lutea of series Hypechusa into a separate clade. Isozyme variation in V. mollis, V. noeana, V. sericocarpa and V. ciliatula of section Hypechusa is described for the first time. Twenty-three species-specific isozymes that may be used for species identification by analysis of seeds are described. Two new isozymes that distinguish V. serratifolia from V. narbonensis are described that further support their specific status. Series Hypechusa and Hyrcanicae of section Hypechusa, sections Narbonensis and Peregrinae of traditional taxonomy are monophyletic groups that deserve sectional status within the genus Vicia. The phylogenetic position and sectional placement of V. lutea warrants further study with independent sets of DNA characters.     

Nuclear DNA contents, rDNAs, and karyotype evolution in subgenus Vicia: III. The heterogeneous section Hypechusa

Summary. Nuclear DNA contents, automated karyotype analyses, and sequences of internal transcribed spacers from ribosomal genes have been determined in the species belonging to section Hypechusa of the subgenus Vicia. Karyomorphological results and phylogenetic data generated from the comparison of rDNA (genes coding for rRNA) sequences showed that sect. Hypechusa is not monophyletic; however, some monophyletic units are apparent (one including Vicia galeata, V. hyrcanica, V. noeana, and V. tigridis, another including V. assyriaca, V. hybrida, V. melanops, V. mollis, and V. sericocarpa), which partly correspond to morphology-based infrasectional groups. The relationships among these species and the species in sections Faba, Narbonensis, Bithynicae, and Peregrinae have been also investigated. Correspondence and reprints: Dipartimento di Agrobiologia ed Agrochimica, Università della Tuscia, via San Camillo de Lellis, 01100 Viterbo, Italy.     

Pollen morphology of the genus Salvia L. (Lamiaceae) in Turkey

Pollen grains of 30 taxa of the genus Salvia, belonging to sections Salvia, Horminum, Drymosphace, Plethiosphace and Hemisphace from Turkey were examined by light (LM) and scanning electron microscopy (SEM). Detailed pollen morphological characteristics are provided for these taxa. Among the studied taxa, S. verticillata subsp. verticillata from sect. Hemisphace has the smallest pollen grains, and S. pachystachys from sect. Salvia possesses the largest ones. The basic shape of the pollen grains in most taxa is suboblate, oblate-spheroidal or prolate-spheroidal. However subprolate pollen grains are recorded for S. macrochlamys from sect. Salvia. The grains are hexacolpate in all taxa, but in S. recognita from sect. Salvia also octacolpate pollen was found. Three distinct exine sculpturing types exist, reticulate-perforate (the common type), reticulate-granulate and bireticulate. The reticulate-perforate and bireticulate sculpturing patterns can be divided into subtypes based on the number of perforations and the number of secondary lumina in each primary lumen. Pollen morphological characteristics of the taxa studied are compared and discussed on the basis of taxonomical concepts. In some cases, these characters are useful in distinguishing the sections. For instance, the presence of 1-2 large central secondary lumina per primary lumen is a significant character of sect. Horminum separating it from the other sections. As well, the presence of holes on colpus membrane ornamentation can be used as a diagnostic taxonomic character for sectional division between sect. Hemisphace and others. S. ballsiana from sect. Salvia is clearly distinct from the other taxa examined by its unique pollen morphology. Further, for several macromorphologically similar taxa pollen structures provide additional evidence to delimite them from each other.     

Palynological investigations on some Hypericum taxa (Hypericaceae) growing naturally in Turkey

This study examined the detailed pollen morphological structures of sixteen Hypericum taxa (four endemic, twelve non endemic) including eight sections showing the natural distribution in Turkey: H. sect. Ascyreia (H. calycinum L.), H. sect. Heterophyllum (H. heterophyllum Vent.), H. Sect. Taeniocarpium (Hypericum confertum Choisy subsp. confertum, H. venustum Fenzl, H. linaroides Bosse), H. sect. Drosocarpium (H. montbretii Spach, H. bithynicum Boiss.), H. sect. Crossophyllum (H. adenotrichum Spach, H. orientale L.), H. sect. Olympia (H. olympicum L. subsp. olympicum), H. sect. Origanifolia (H. origanifolium Willd., H. avicularifolium Jaub and Spach subsp. depilatum (Freyn and Bornm.) Robson var. depilatum, H. avicularifolium Jaub. and Spach subsp. byzantinum) and H. sect. Hypericum (H. tetrapterum Fries, H. perforatum L., H. triquetrifolium Tura). These taxa were studied under light microscope and scanning electron microscope for the first time. Of the taxa studied, H. tetrapterum has the smallest pollen grains (on average 15.85–17.20 × 15.45–16.05 μm); and H. olympicum subsp. olympicum the largest grains (on average 22.90–23.10 × 21.40–22.10 μm). The pollen grains of 15 taxa are subprolate and one taxon is prolate-spheroidal. The regular pollen grains of all 16 taxa are 3-zonocolporate. Ornamentation is microreticulate; lumina regularly spaced in eight taxa, tectum perforatum; tectal perforations regularly spaced in seven taxa and tectum perforatum; and tectal perforations grouped together in one taxon. Endoaperture is cruciform porus, with short lateral and meridional extensions in ten taxa, cruciform porus, transversally elongated, with very small lateral extensions in one taxon and lalongate colpus in five taxa. Basic pollen types are ten taxa in type X, five taxa in type IV and one taxon in type II.     

Taxonomic relationships between V. faba and its relatives based on nuclear and mitochondrial RFLPs and PCR analysis

Summary The taxonomic relationships between 52 accessions of 12 Vicia species and three accessions of Lathyrus were examined using nuclear RFLP- and PCR-generated data. Two hundred and sixty informative restriction fragments or amplification products were analysed by single linkage analysis, average cluster analysis, and the Wagner parsimony method. Dendrograms constructed from each type of analysis showed similar overall topologies and could be divided into three parts corresponding respectively to the Lathyrus outgroup, the species grouped in the section Faba/narbonemis complex, and the species belonging to the sections Hypechusa and Peregrinae. With few exceptions, the majority of accessions belonging to one species grouped together before branching to other species. An analysis of mitochondrial DNA phenotypes was both consistent with and complemented the results from the nuclear data. Overall, the species relationships show a good correlation with the classification of Maxted et al. but suggest that V. faba is more closely aligned to species from the sections Hypechusa and Peregrinae than to those in the narbonensis complex. In addition, the position of two new species, V. kalakhensis and V. eristaloides, as members of the narbonensis complex was supported by the molecular data, which also allowed a preliminary classification for recently collected Vicia accessions.     

Phylogenetic relationships betweenVicia faba (Fabaceae) and related species inferred from chloroplasttrnL sequences

The chloroplasttrnL intron from 46 differentVicia accessions, representing five of the nine sections of the genusVicia subg.Vicia sensuMaxted (1991a) were amplified by the Polymerase Chain Reaction (PCR) using oligonucleotide primers homologous to conserved regions intrnL. The products fell into two distinct groups; those of approximately 250 nt and those of around 450 nt in length. Of these, products from 17 differentVicia species were cloned and their nucleotide sequences determined. Multiple alignments were assembled and phylogenetic trees constructed by the weighted least-squares distance method. ALathyrus latifolius trnL intron sequence was used as an outgroup. The resulting trees clearly group and separate the sectt.Narbonensis, Bithynica andFaba species but were less able to distinguish species from sectt.Hypechusa andPeregrinae. Based on these sequence data,V. faba appears to be more distant from sect.Narbonensis than sectt.Hypechusa andPeregrinae. The results are in general agreement with a recent treatment ofVicia subg.Vicia (Maxted 1993) and lend further support to placingV. faba in the monospecific sect.Faba.     

Cladistic and phenetic analysis of relationships in Vicia subgenus Vicia (Fabaceae) by morphology and isozymes

 Variation of 80 multistate morphological characters and isozymes encoded by 13 loci among 23 vetch species of the type subgenus of the genus Vicia in comparison with V. dumetorum, V. pisiformis and V. sylvatica of the subgenus Cracca is described and analyzed with cladistic parsimony and phenetic neighbour-joining methods by using two different ways of coding. Morphological analyses showed the subgenus Vicia monophyletic and revealed subgroups in a general agreement with traditionally recognized sections, except showing V. faba nested within section Narbonensis and ambiguity in the position of V. lathyroides and V. bithynica. Parsimony analysis of orthozymes as presence/absence characters revealed in the subgenus two basic monophyletic clades: 1) V. faba and three species of the section Peregrinae, V. michauxii, V. aintabensis and V. peregrina, in one subclade linked with species of the Narbonensis and Hyperchusa sections together with V. pisiformis of subgenus Cracca in a second subclade; 2) species belonging to sections Vicia, Sepium, Pseudolathyrus and Lathyroides together with V. sylvatica of the subgenus Cracca. Neighbour-joining analysis of orthozymes revealed the same two basic groups, differing only in the relative position of some species in subclusters. Both isozyme analyses showed paraphyly of the subgenera Vicia and Cracca. Parsimony analysis of orthozymes as character states of isozymes yielded a largely unresolved strict consensus cladogram of 209 most parsimonious trees, and reweighting of characters failed to produce a stable tree. Phylogenetic congruence and discordance among morphological and isozyme analyses, coding ways, homoplasy and weighting of characters are discussed. Received November 20, 2001 Accepted January 31, 2002     

Genetic diversity and relationships in some <Emphasis Type="Italic">Vicia</Emphasis> species as determined by SDS-PAGE of seed proteins

To evaluate the genetic diversity of some Vicia species, seed proteins of 160 accessions (30 of Vicia faba, 15 of V. narbonensis, 82 of V. sativa and 25 of V. ervilia and 8 accessions of other Vicia species) were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The dendrogram showed that the two outcrossing species V. faba and V. villosa were the most distant among all species (average percent disagreement value PDV 0.47 and 0.45, respectively). The tree was divided into small clusters of two species each. V. narbonensis fell in one cluster with V. michausai (at PDV = 0.35) and V. lutea (var. hirta) fell in one cluster with V. serococorpes (at PDV = 0.32) whereas, V. ervilia fell in one cluster with V. sativa (at PDV = 0.27). The V. sativa subspecies, however, were closely related (PDV < 0.1). In general, this study did not prove any relationship between the studied storage proteins and the geographical distribution or ecological needs of the studied accessions.     

Nuclear DNA contents, rDNAs, and karyotype evolution in Vicia subgenus Vicia: II. Section Peregrinae

Summary. Nuclear DNA contents, automated karyotype analyses, and sequences of rDNA spacers have been determined for the species of Vicia belonging to sect. Peregrinae, as well as for V. mollis. The phylogenetic data generated from the comparison of rDNA sequences and karyomorphological results would both indicate that Vicia mollis is a sister group to sect. Peregrinae. The relationships among the species belonging to the Peregrinae section and species enclosed in sections Faba, Narbonensis, and Bithynicae have been also investigated: a clade including V. mollis and sect. Peregrinae is a sister group to a clade including V. bithynica and sect. Narbonensis. With our choice of outgroup, Vicia faba (including subsp. paucijuga) is external to the above mentioned inclusive group. Correspondence and reprints: Dipartimento di Agrobiologia ed Agrochimica, Università della Tuscia, via San Camillo de Lellis, 01100 Viterbo, Italy.     

Field evaluation of resistance to black bean aphid, Aphis fabae, in close relatives of the Faba bean, Vicia faba

A field assessment of 26 accessions of Vicia narbonensis and three of V. johannis confirmed previous laboratory studies demonstrating higher levels of resistance to Aphis fabae in these two wild species compared to the closely related crop, Vicia faba. Accessions of V. johannis were significantly more resistant than most accessions of V. narbonensis for all resistance indices measured except survival of aphid nymphs. Plant growth stage significantly affected levels of resistance in both Vicia species, resistance being moderate at pre-bud stage, decreasing on flowering and rising again at pod fill and onset of leaf senescence. Significant intraspecific variability in aphid resistance was found only within the 26 accessions of V. narbonensis, var. serratifolia being more resistant than var. narbonensis. Possible resistance factors and the agronomic potential of these two wild relatives of Faba bean are considered.     

Molecular taxonomy of Vicia ser. Vicia based on Amplified Fragment Length Polymorphisms

 Amplified Fragment Length Polymorphisms (AFLPs) were used to study the taxonomic relationships in Vicia ser. Vicia. 109 accessions representing the taxa and distribution in the series were evaluated using two primer combinations in an attempt to clarify its taxonomy. The study concludes that four species exist in the series. Vicia incisa should be considered as a separate taxon and does not belong to the Vicia sativa aggregate. Within the series Vicia pyrenaica appears to be the most closely related to the Vicia sativa aggregate. Within the aggregate 6 taxa can be recognised. They are very closely related and for none of the taxa in the aggregate is there a taxon-specific absence or presence of AFLP bands. We consider these taxa to be subspecies. Several accessions belonging to the aggregate were found which could not be placed within one of the taxa and are most likely the products of mutations or hybridisation between taxa. Received March 12, 2001 Accepted July 12, 2001     

Microarray-based survey of repetitive genomic sequences in Vicia spp.

A modified DNA microarray-based technique was devised for preliminary screening of short fragment genomic DNA libraries from three Vicia species (V. melanops, V. narbonensis, and V. sativa) to isolate representative highly abundant DNA sequences that show different distribution patterns among related legume species. The microarrays were sequentially hybridized with labeled genomic DNAs of thirteen Vicia and seven other Fabaceae species and scored for hybridization signals of individual clones. The clones were then assigned to one of the following groups characterized by hybridization to: (1) all tested species, (2) most of the Vicia and Pisum species, (3) only a few Vicia species, and (4) preferentially a single Vicia species. Several clones from each group, 65 in total, were sequenced. All Group I clones were identified as rDNA genes or fragments of chloroplast genome, whereas the majority of Group II clones showed significant homologies to retroelement sequences. Clones in Groups III and IV contained novel dispersed repeats with copy numbers 10²–10⁶/1C and two genus-specific tandem repeats. One of these belongs to the VicTR-B repeat family, and the other clone (S12) contains an amplified portion of the rDNA intergenic spacer. In situ hybridization using V. sativa metaphase chromosomes revealed the presence of the S12 sequences not only within rDNA genes, but also at several additional loci. The newly identified repeats, as well as the retroelement-like sequences, were characterized with respect to their abundance within individual genomes. Correlations between the repeat distributions and the current taxonomic classification of these species are discussed.     

Variation of pollen morphology, and its implications in the phylogeny of Clematis (Ranunculaceae)

Clematis s.l. (including Archiclematis and Naravelia) is a genus of approximately 300 species with cosmopolitan distribution. The diversity of its pollen was surveyed in 162 taxa belonging to all infrageneric groups of Clematis s.l. Pollen morphology was investigated by use of scanning electron microscopy to identify useful characters, test taxonomic and systematic hypotheses, and elucidate pollen character evolution on the basis of the molecular phylogeny. Clematis pollen is small to medium (14.8–32.1?μm?×?14.2–28.7?μm), oblate to prolate (P/E?=?0.9–1.4) in shape. The apertures may be tricolpate and pantoporate sometimes with 4-zonocolpate and pantocolpate pollen grains as transitional forms. The tricolpate pollen grains are predominant and occur in all the sections of the genus, whereas pantoporate pollen grains can be found in sect. Tubulosae, sect. Viorna, sect. Viticella, and Naravelia only. Phylogenetic mapping of aperture types reveals that the pantoporate pollen type may be the apomorphy in the genus and evolved several times. The surface ornamentation in all taxa studied is similar and characterized by microechinae evenly distributed on the microperforate tectum. The size and density of spinules on the tectum vary greatly but successive in the whole genus. According to the character syndromes of the ornamentation, separating sect. Brachiata from sect. Meclatis is supported. Though pollen morphology may contribute to investigation of problematic taxa, the taxonomic value of pollen morphology is limited at the species level.     

Vicia johannis and wild relatives of the faba bean: a taxonomic study

Preliminary taxonomic and agronomic investigation was made of close relatives of the faba bean (Vicia faba) to aid future plant breeding. Cluster analysis of 2 yr’s morphometric data showed thatV. johannis is clearly separable from other members of theV. narbonensis complex. AlsoV. faba was much more distinct from eitherV. narbonensis orV. johannis than they are from each other. Petal colours at anthesis provided a rapid method of field identification forV. faba, V. narbonensis, andV. johannis. The 2 varieties ofV. johannis can also be distinguished by their petal colours: var. procumbens shows a marked colour change at anthesis and var. johannis does not. The geographical distribution of the plant material studied suggests thatV. johannis is found in Turkey and Europe despite being omitted from the major published floras. BothV. johannis andV. narbonensis possess agronomically useful characters of importance to faba bean breeders, including increased frost tolerance and resistance to bothAphis fabae andBotrytis fabae.     

Isozyme evidence on the specific distinctness and phylogenetic position of <Emphasis Type="Italic">Vicia incisa</Emphasis> (Fabaceae)

Vicia incisa is a taxonomically controversial species that has been also treated as a subspecies of V. sativa because of a great morphological similarity. The phylogenetic position of V. incisa is uncertain because various DNA markers have provided contradictory results. Isozymes of V. incisa encoded by 15 loci and resolved with the use of polyacrylamide gel electrophoresis (PAGE) are described and compared with those of seven related species belonging to sections Vicia, Sepium, Lathyroides and Pseudolathyrus in order to get new evidence about its taxonomic rank and phylogenetic position. Phylogenetic relationships are analyzed with maximum parsimony and neighbour joining methods. Vicia incisa is shown to differ from all three subspecies of V. sativa including, sativa, cordata and nigra, by alternate variants of ten isozymes out of 15 analysed. Instead, V. incisa has much more similarity to V. grandiflora and V. sepium by sharing eight isozyme variants which differ from the subspecies of V. sativa. The most parsimony and neighbour joining analyses of the isozyme variation placed V. incisa as basally linked to the V. grandiflora and V. sepium species couple in the clade of section Sepium (= sect. Atossa), while the subspecies of V. sativa together with V. lathyroides formed a separate clade of section Vicia. The isozyme data provide further support to the species status of V. incisa.     

Electrophoretic seed albumin patterns inVicia species of sectt.Hypechusa andPeregrinae (Fabaceae)

This work is a continuation of electrophoretic investigations aimed at revealing a wild relative ofVicia faba. Electrophoretic analysis (PAGE) of seed albumins covered 52 accessions representing eightVicia species of sect.Hypechusa and two species of sect.Peregrinae. Most of the examined species showed an intraspecific variation due to differences between accessions and/or individual variation within accessions. In spite of the intraspecific variation, marked interspecific differences were recorded. However, none of the investigated species displayed electrophoretic seed albumin patterns similar to those reported earlier forV. faba. Contribution of the obtained results to characterization of the examined taxa is discussed.     

Cytological and molecular characterization of Vicia esdraelonensis Warb. & Eig: a rare taxon

Summary. Vicia esdraelonensis, a rare taxon belonging to section Hypechusa of subgenus Vicia, was recovered and analyzed by cytological, karyological, and molecular methods, with the aim of both characterizing this species and furthering our knowledge of the phylogeny of subgenus Vicia. Automated karyotype analysis, nuclear DNA content, and chromatin organization were determined by the Feulgen reaction, as well as chromosome banding after double staining with 4′,6-diamidino-2-phenylindole (DAPI) and chromomycin A3. The chromosome number and the nuclear DNA content were in agreement with the values of the species of section Hypechusa. The GC- and AT-rich preferential sites were determined by chromomycin A3 and DAPI staining. Karyomorphological parameters indicated that V. esdraelonensis is in an intermediate position in the spatial representation of the species of section Hypechusa on the basis of symmetry indices, as well as in the dendrogram of linkage distance constructed on 37 chromosome parameters. Molecular data based on internal transcribed spacer sequences show that V. esdraelonensis can doubtlessly be included in section Hypechusa and document its closeness to V. noeana. A cladistic analysis combining the molecular data set with karyological characters is also reported. Karyological, cytological, and molecular data allow characterization of the V. esdraelonensis genome and provide information about the phylogenetic position of this species within the Hyrcanicae series of section Hypechusa. Correspondence and reprints: Dipartimento di Biologia, Università di Pisa, Via L. Ghini 5, 56126 Pisa, Italy.     

Analisi citologica di alcune specie del genere Vicia

Abstract

Karyology of some species of Vicia. The species studied are: Vicia articulata, Vicia monantha, Vicia narbonensis and Vicia ervilia.

The chromosome number 2n = 14 counted for the first three species agrees with that reported in literature; that of Vicia monantha (2n=14) agrees with that quoted by Heitz, while Senn finds 2n=12.

Karyotypes of V. narbonensis and V. ervilia show appreciable differences from those reported in literature.     

Electrophoretic seed albumin patterns and species relationships inVicia sect.Faba (Fabaceae)

Electrophoretic analysis of seed albumins (PAGE) covered 173 accessions representing nine species ofVicia sect.Faba. The number of albumin bands recorded in particular species varied from three inV. eristaloides to 23 inV. faba; in total, 38 bands were distinguished in the investigated material. The examined species, exceptV. eristalioides, showed intraspecific variation with respect to the number and relative staining intensity of albumin bands; individual variation was especially marked inV. faba and inV. narbonensis. Hierarchical clustering of the investigated taxa was based onBhattacharyya distances calculated from the electrophoretic data. The taxa grouped in three main clusters.Vicia faba and the rather remotely relatedV. kalakhensis formed one cluster. The second cluster was composed ofV. narbonensis distantly related toV. hyaeniscyamus. The third cluster comprised three subgroups: 1.V. johannis, V. galilaea andV. serratifolia, 2.V. eristalioides, and 3.V. bithynica. The obtained results are discussed with reference to taxonomic relationships inVicia sect.Faba.