Gustavo M. Mori

Multiple-geographic-scale genetic structure of two mangrove tree species: the roles of mating system, hybridization, limited dispersal and extrinsic factors.

Mangrove plants comprise a unique group of organisms that grow within the intertidal zones of tropical and subtropical regions and whose distributions are influenced by both biotic and abiotic factors. To understand how these extrinsic and intrinsic processes influence a more fundamental level of the biological hierarchy of mangroves, we studied the genetic diversity of two Neotropical mangrove trees, Avicenniagerminans and A. schaueriana, using microsatellites markers. As reported for other sea-dispersed species, there was a strong differentiation between A. germinans and A. schaueriana populations sampled north and south of the northeastern extremity of South America, likely due to the influence of marine superficial currents. Moreover, we observed fine-scale genetic structures even when no obvious physical barriers were present, indicating pollen and propagule dispersal limitation, which could be explained by isolation-by-distance coupled with mating system differences. We report the first evidence of ongoing hybridization between Avicennia species and that these hybrids are fertile, although this interspecific crossing has not contributed to an increase in the genetic diversity the populations where A. germinans and A. schaueriana hybridize. These findings highlight the complex interplay between intrinsic and extrinsic factors that shape the distribution of the genetic diversity in these sea-dispersed colonizer species.

Development and characterization of microsatellite markers for the wild South American Passiflora cincinnata (Passifloraceae)

PREMISE OF THE STUDY We developed the first set of microsatellite markers for the passion fruit, Passiflora cincinnata, to provide tools for further study of its genetic diversity and to support current conservation and genetic studies. METHODS AND RESULTS We used a microsatellite-enriched library approach to isolate and characterize 25 new molecular markers. Seven primer pairs successfully amplified polymorphic microsatellite regions, with an average of five alleles per locus. The mean values of expected and observed levels of heterozygosity were 0.516 and 0.525, respectively. CONCLUSIONS The microsatellite markers identified in this study may be valuable tools for population genetic studies, and this set of markers also may be useful in the design of a genetic conservation strategy, mating system, and hybridization studies.

New Microsatellite Markers for Wild and Commercial Species of Passiflora (Passifloraceae) and Cross-Amplification

Premise of the study: We developed the first microsatellites for Passiflora setacea and characterized new sets of markers for P. edulis and P. cincinnata, enabling further genetic diversity studies to support the conservation and breeding of passion fruit species. Methods and Results: We developed 69 microsatellite markers and, in conjunction with assessments of cross-amplification using primers available from the literature, present 43 new polymorphic microsatellite loci for three species of Passiflora. The mean number of alleles per locus was 3.1, and the mean values of the expected and observed levels of heterozygosity were 0.406 and 0.322, respectively. Conclusions: These microsatellite markers will be valuable tools for investigating the genetic diversity and population structure of wild and commercial species of passion fruit (Passiflora spp.) and may be useful for developing conservation and improvement strategies by contributing to the understanding of the mating system and hybridization within the genus.

Microsatellites for the mangrove tree Avicennia germinans (Acanthaceae): Tools for hybridization and mating system studies

UNLABELLED PREMISE OF THE STUDY We developed a new set of microsatellite markers for the black mangrove Avicennia germinans, to provide new informative tools for further studies of the mating system, interspecific hybridization, and population genetics. • METHODS AND RESULTS We used the microsatellite-enriched library approach to isolate and characterize 25 new primer pairs. Sixteen of them are polymorphic, showing a variable degree of variation in A. germinans, while nine were monomorphic in the samples examined. Eight exhibited private alleles in A. schaueriana. • CONCLUSIONS These results indicate that these new microsatellite markers will be useful molecular tools for further studies of A. germinans and A. schaueriana population genetics, mating systems, and hybridization.

Species distribution and introgressive hybridization of two Avicennia species from the Western Hemisphere unveiled by phylogeographic patterns

BackgroundMangrove plants grow in the intertidal zone in tropical and subtropical regions worldwide. The global latitudinal distribution of the mangrove is mainly influenced by climatic and oceanographic features. Because of current climate changes, poleward range expansions have been reported for the major biogeographic regions of mangrove forests in the Western and Eastern Hemispheres. There is evidence that mangrove forests also responded similarly after the last glaciation by expanding their ranges. In this context, the use of genetic tools is an informative approach for understanding how historical processes and factors impact the distribution of mangrove species. We investigated the phylogeographic patterns of two Avicennia species, A. germinans and A. schaueriana, from the Western Hemisphere using nuclear and chloroplast DNA markers.ResultsOur results indicate that, although Avicennia bicolor, A. germinans and A. schaueriana are independent lineages, hybridization between A. schaueriana and A. germinans is a relevant evolutionary process. Our findings also reinforce the role of long-distance dispersal in widespread mangrove species such as A. germinans, for which we observed signs of transatlantic dispersal, a process that has, most likely, contributed to the breadth of the distribution of A. germinans. However, along the southern coast of South America, A. schaueriana is the only representative of the genus. The distribution patterns of A. germinans and A. schaueriana are explained by their different responses to past climate changes and by the unequal historical effectiveness of relative gene flow by propagules and pollen.ConclusionsWe observed that A. bicolor, A. germinans and A. schaueriana are three evolutionary lineages that present historical and ongoing hybridization on the American continent. We also inferred a new evidence of transatlantic dispersal for A. germinans, which may have contributed to its widespread distribution. Despite the generally wider distribution of A. germinans, only A. schaueriana is found in southern South America, which may be explained by the different demographic histories of these two species and the larger proportion of gene flow produced by propagules rather than pollen in A. schaueriana. These results highlight that these species responded in different ways to past events, indicating that such differences may also occur in the currently changing world.

Molecular Genetic Variability of Commercial and Wild Accessions of Passion Fruit (Passiflora spp.) Targeting ex Situ Conservation and Breeding

Passiflora species are distributed throughout Latin America, and Brazil and Colombia serve as the centers of diversity for this genus. We performed cross-species amplification to evaluate 109 microsatellite loci in 14 Passiflora species and estimated the diversity and genetic structure of Passiflora cincinnata, Passiflora setaceae and Passiflora edulis. A total of 127 accessions, including 85 accessions of P. edulis, a commercial species, and 42 accessions of 13 wild species, were examined. The cross-species amplification was effective for obtaining microsatellite loci (average cross-amplification of 70%). The average number of alleles per locus (five) was relatively low, and the average diversity ranged from 0.52 in P. cincinnata to 0.32 in P. setacea. The Bayesian analyses indicated that the P. cincinnata and P. setacea accessions were distributed into two groups, and the P. edulis accessions were distributed into five groups. Private alleles were identified, and suggestions for core collections are presented. Further collections are necessary, and the information generated may be useful for breeding and conservation.

Can environmental pollution by metals change genetic diversity? Ucides cordatus (Linnaeus, 1763) as a study case in Southeastern Brazilian mangroves

Industrial areas on estuarine systems are commonly affected by heavy metals, affecting all local biota. Random Amplified Polymorphic DNA (RAPD) was used to evaluate genetic diversity of Ucides cordatus at mangroves in southeastern Brazil (Juréia, J; São Vicente, SV; and Cubatão, C), with distinct pollution levels by metals. The genetic diversity of this species was compared with concentrations of metals (Cd, Pb, Cu, Cr and Hg) in the environment. A pollution gradient was confirmed (SV>C>J), with low levels detected in water, except for mercury in SV. All metals in the sediment samples were below Threshold Effect Level (TEL), without an apparent biological risk to the biota. Genetic distance was very similar between J and C, with SV occurring as an out-group. RAPD was a powerful tool to investigate the effect of metal pollution on genetic diversity of this mangrove crab, and to evaluate the conservation status of the mangrove ecosystem.

New polymorphic microsatellite loci for Theobroma cacao: isolation and characterization of microsatellites from enriched genomic libraries

Seventeen polymorphic microsatellite markers were isolated from enriched genomic libraries for Theobroma cacao, providing additional tools for studying the genetic diversity and map saturation of this species. These markers were characterized in 32 accessions of the T. cacao germplasm collection from the Centro de Pesquisas do Cacau. The number of alleles at each locus varied from 2 to 8, with an average of 4.41 alleles per locus. The polymorphism information content varied from 0.060 to 0.695, with an average of 0.333. The markers characterized in this study will be employed in map saturation studies and diversity assessments of cacao genotypes.

Isolation and characterization of microsatellites from Scaptotrigona xanthotricha (Apidae, Meliponini): a stingless bee in the Brazilian Atlantic rainforest

Scaptotrigona / population genetics / polymorphism / Hymenoptera Scaptotrigona bees are efficient pollinators as they use odors in marking trails to distant food sources. They build their nests in cavities of large living trees making them susceptible to anthropic actions, such as deforestation. Due to the reduction in population size as a result of fragmentation and destruction of natural habitat, many species are threatened with extinction (Freitas et al. 2009). Scaptotrigona comprises 30 Neotropical species, out of which 18 have been recorded in Brazil (Moure et al. 2007). This genus still harbors many undescribed species and cryptic species complexes (Silveira et al. 2002). Scaptotrigona xanthotricha shows a wide distribution in the Atlantic rainforest ranging from southern to the northeastern Brazil (Silveira et al. 2002). Duarte et al. (2009) reported a remarkable geographical variation in the number and location of the Nucleolus Organizer Regions in this species. This finding lent support to the existence of interpopulational divergence or even additional taxa, thus reinforcing the need for more detailed assessment of this group using molecular markers. Studies on the genetic diversity and population structure of stingless bees have become frequent due to the development of microsatellite markers for several species such as Melipona mondury (Lopes et al. 2010), Melipona bicolor (Peters et al. 1998), and Scaptotrigona postica (Paxton et al. 1999), among others. However, these studies are still scarce when compared to the high diversity within this group. The present study aimed at the identification and characterization of microsatellite loci for S. xanthotricha. An enriched genomic library was constructed from a pool of DNA from 30 individuals of the same colony following Billotte et al. (1999). DNA extraction followed Han and McPheron (1997). Five micrograms of DNA (300 ng/μl) was digested with RsaI and linked to RsaI adapters. The library was enriched for (CT) 8 and (GT) 8 repeats using biotinylated probes attached to streptavidin-labeled magnetic beads as described by the manufacturer (MagneSphere Paramagnetic Particles , Promega). Selected fragments amplified by polymerase chain reaction (PCR) using primers with sequences complementary to the adapters and inserted into the plasmid pGEM-T (Promega) were transformed into competent XL-1 Blue Escherichia coli cells. Once most of the colonies were transformed, 96 were selected for long-term storage at −80°C into a micro-plate. Forty-eight positive clones were sequenced on an automated sequencer ABI PRISM 377 (Applied Biosystems) using SP6 and T7 primers and the Big Dye terminator kit …

Characterization of microsatellite loci in Himatanthus drasticus (Apocynaceae), a medicinal plant from the Brazilian savanna

UNLABELLED PREMISE OF THE STUDY We developed a new set of microsatellite markers for studying the genome of the janaguba tree, Himatanthus drasticus (Mart.) Plumel, which is used in folk medicine in northeastern Brazil. These novel markers are being used to evaluate the effect of harvesting on the genetic structure and diversity of natural populations of this species. • METHODS AND RESULTS Microsatellite loci were isolated from an enriched H. drasticus genomic library. Nine primer pairs successfully amplified polymorphic microsatellite regions, with an average of 8.5 alleles per locus. The average values of observed and expected heterozygosity were 0.456 and 0.601, respectively. • CONCLUSIONS The microsatellite markers described here are valuable tools for population genetics studies of H. drasticus. The majority of the primers also amplified sequences in the genome of another species of the same genus. This new set of markers may be useful in designing a genetic conservation strategy and a sustainable management plan for the species.