Paulo M. Ruas

Reproductive isolation in the Aegean Ophrys omegaifera complex (Orchidaceae)

The orchid genus Ophrys operates a system of sexual deception by which high specificity of pollination is attained. Reproductive isolation in Ophrys mainly rests upon prezygotic isolation mechanisms. The level of genetic separateness of Ophrys taxa with different pollinators is therefore likely determined by the fidelity of pollinators. The present study employs genetic fingerprinting to investigate this in the east Aegean Ophrys omegaifera s.l. complex, also including O. dryis, a west Mediterranean species of this complex. Ophrys fleischmannii, O. basilissa, and the west Mediterranean O. dyris, are found to be well-separated genetic entities whereas O. omegaifera s.str. and the putative hybrid taxon, O. sitiaca, are found to be genetically inseparable across their entire range of co-occurrence. This suggests that specific pollinators have high enough fidelity to act as effective isolating factors in east Aegean O. omegaifera s.l. as a whole, but that the situation in the species pair of O. sitiaca and O. omegaifera is likely to be more complex.

Evidence for progenitor–derivative speciation in sexually deceptive orchids

BACKGROUND AND AIMSnSexually deceptive orchids of the genus Ophrys use mimicry of pollinator females to attract specific pollinators. Pollinator shifts may drive speciation in Ophrys, since novel pollinators may in principle act as isolating factors immediately. It is thus possible that evolution of novel species occurs rapidly and with a progenitor-derivative pattern. The aims of this study are to compare genetic structure and diversity among widespread and geographically restricted Ophrys taxa, to test whether genetic structure is associated with specific pollinators, and to investigate whether any widespread species may have acted as a progenitor for the evolution of more restricted taxa.nnnMETHODSnGenetic differentiation and diversity were investigated in O. leucadica and O. cinereophila, the two taxa of the Ophrys fusca sensu lato complex widespread in the Aegean, and three geographically restricted taxa from Rhodes, O. attaviria, O. parvula and O. persephonae, all differing in their specific pollinators. This was done using amplified fragment length polymorphism (AFLP) DNA fingerprinting, and sequencing of the low-copy nuclear gene LEAFY (LFY).nnnKEY RESULTSnAll taxa were found to be separate genetic entities, with O. leucadica forming two geographic groups from the west and east of the Aegean. Genetic structure was significantly shaped by pollinators and geography, and comparison of sequence and AFLP data revealed ancestral polymorphisms shared among several taxa. Among the sampled taxa, O. leucadica harbours the greatest genetic differentiation and geographic structure, and the highest genetic diversity. Part of the genome of O. parvula, endemic to Rhodes, may be derived from O. leucadica.nnnCONCLUSIONSnPollinators probably influence the genetic structure of the investigated Ophrys species. The genetic pattern identified is consistent with O. leucadica being the oldest of the sampled taxa, making O. leucadica a candidate progenitor species from which more restricted taxa such as O. parvula may have evolved.

Characterization, genomic organization and chromosomal distribution of Ty1-copia retrotransposons in species of Hypochaeris (Asteraceae)

This study aims to analyze the diversity of Ty1-copia retrotransposons in 18 taxa of Hypochaeris, including two Old World species H. maculata (2n=2x=10) and H. angustifolia (2n=2x=8), and representatives of the South American species (16 accessions of 15 species; all 2n=2x=8). Analysis of 380 PCR-amplified sequences, corresponding to a conserved domain of the subset of Ty1-copia reverse transcriptase (rt) gene amplifiable with degenerate standard primers, showed high levels of intra- and interspecific heterogeneity. Nucleotide diversity (Pi) of the copia fragments was high in all species and varied from 0.229 (H. angustifolia) to 0.412 (H. chillensis). Higher sequence heterogeneity correlates positively with larger genome size among analyzed species. Phylogenetic analyses of amplified fragments revealed different patterns of intraspecific heterogeneity within species, with most sequences forming one well-supported main clade while a few sequences fall into small clades or are left ungrouped. The combined analysis of all sequences revealed the presence of three main clades and showed that highly diverged species contain closely related Tyl-copia group retrotransposons. One of the main clades includes rt sequences of all South American species and three sequences of their putative ancestor, H. angustifolia, but no sequence of the Old World H. maculata. FISH with copia retrotransposons in four Hypochaeris species, including H. maculata and H. angustifolia and New World H. apargioides and H. spathulata, revealed differences in the chromosomal distribution between the two groups. In Old World species copia retroelements are distributed over the whole length of the chromosomes, excluding rDNA sites and some centromeres. In the South American species the two largest chromosome pairs are enriched in copia, while most of the long arms of the two small pairs of chromosomes are devoid of these elements. The patterns of heterogeneity and chromosomal distribution of Ty1-copia retrotransposons in Hypochaeris are discussed in the context of the origin, genome evolution and organization of the South American species.

Genetic Structure and Diversity of Brazilian Tree Species from Forest Fragments and Riparian Woods

Understanding the genetic structure of populations that occur in forest remnants is fundamentally necessary to establish efficient strategies for the re-composition, management, and conservation programs. For such, it is necessary not only to understand the genetic diversity of a species, but also, how this diversity is distributed within and between forest populations. Notably, a considerable part, if not the majority, of Brazilian Atlantic Forest fragments are linked to rivers or streams, once the policy applied in Brazil regarding conservation in agricultural areas favours the maintenance of legal reserves in proximity of water sources. The vegetation of river margins are subjected to flooding, a strong limiting factor which can lead to local adaptation. These ecological and landscape characteristics may have important outcomes to the genetic diversity of tree populations.

Characterization of 12 microsatellite loci for Hypochaeris chillensis (Asteraceae) and cross-amplification in related species

UNLABELLEDnnnnPREMISE OF THE STUDYnHypochaeris is considered a biological model to understand evolutionary processes in the vascular flora of South America, particularly from the temperate portion of the continent. We report the development and characterization of microsatellite markers for H. chillensis to assess the genetic variability and patterns of population structure of the species. •nnnMETHODS AND RESULTSnTwelve microsatellite primers were isolated using a CT- and GT-enriched genomic library. PCR amplification detected one to five alleles, with 2.91 alleles per locus on average. Tested for cross-amplification, all primer pairs were successfully amplified in 10 South American species and in the putative ancestor of the group, H. angustifolia. •nnnCONCLUSIONSnThe microsatellites can be used to assess genetic diversity and population structure of H. chillensis. Application in other species will focus on the elucidation of adaptive radiation of the genus in South America.

Genetic variability in Brazilian Capsicum baccatum germplasm collection assessed by morphological fruit traits and AFLP markers.

Capsicum baccatum is one of the main pepper species grown and consumed in South America. In Brazil, it is commonly cultivated by family farmers, using mostly the genotypes bishops hat genotypes (locally cambuci) and red chili pepper (dedo-de-moça). This study had the objective of characterizing 116 C. baccatum accessions from different regions of Brazil, based on morphological fruit descriptors and AFLP (Amplified Fragment Length Polymorphisms) markers. Broad phenotypic variability among the C. baccatum accessions was detected when using morphological fruit descriptors. The Ward modified location model (Ward-MLM) discriminated five groups, based mainly on fruit shape. Six combinations of AFLP primers detected polymorphism in 97.93% of the 2466 identified bands, indicating the high genetic variability in the accessions. The UPGMA coincided with the Bayesian clustering analysis and three large groups were formed, separating the wild variety C. baccatum var. praetermissum from the other accessions. There was no relation between genetic distance and geographical origin of the accessions, probably due to the intense exchange of fruits and seeds between farmers. Morphological descriptors used together with AFLP markers proved efficient in detecting the levels of genetic variability among the accessions maintained in the germplasm collections. These results can be used as an additional source of helpful information to be exploited in C. baccatum breeding programs.

Distinctness of Brazilian common bean cultivars with carioca and black grain by means of morphoagronomic and molecular descriptors

Common bean (Phaseolus vulgaris L.) is one of the most important legumes for human consumption and is a staple food in the diet of the population of some countries of Latin America, Africa and Asia. The distinction between cultivars is based predominantly on morphological descriptors, which proved inefficient for the differentiation of some cultivars. This study had the objective of describing, distinguishing and evaluating the agronomic potential of 39 common bean cultivars of the carioca and black grain groups registered for cultivation in Brazil, based on 49 morphoagronomic descriptors and microsatellite (simple sequence repeat -SSR) markers. The morphoagronomic traits of each commercial group were characterized in four environments. Thirty-seven SSR markers were used for the molecular description. The morphological data, analyzed by the Shanonon-Weaver index, detected low variability among cultivars for qualitative data. On the other hand, the estimates of variance analysis, relative importance of the traits and hierarchical grouping analysis applied to the quantitative variables showed that the descriptors related to plant morphology were the most important for the carioca group, and those related to seed morphology were determining for the black group. The genetic parameters estimated for SSR markers by hierarchical and Bayesian cluster analysis identified 116 alleles, with 33 and 30 polymorphic loci and 24 and 22 private alleles for the carioca and black groups, respectively. The combined use of morphoagronomic and molecular descriptors improves the distinguishability of cultivars, contributing in a more efficient way to breeding and to the protection of cultivars.