Three complete chloroplast genomes from two north American Rhus species and phylogenomics of Anacardiaceae.

BACKGROUND: The suamc genus Rhus (sensu stricto) includes two subgenera, Lobadium (ca. 25 spp.) and Rhus (ca. 10 spp.). Their members, R. glabra and R. typhina (Rosanae: Sapindales: Anacardiaceae), are two economic important species. Chloroplast genome information is of great significance for the study of plant phylogeny and taxonomy. RESULTS: The three complete chloroplast genomes from two Rhus glabra and one R. typhina accessions were obtained with a total of each about 159k bp in length including a large single-copy region (LSC, about 88k bp), a small single-copy regions (SSC, about 19k bp) and a pair of inverted repeats regions (IRa/IRb, about 26k bp), to form a canonical quadripartite structure. Each genome contained 88 protein-coding genes, 37 transfer RNA genes, eight ribosomal RNA genes and two pseudogenes. The overall GC content of the three genomes all were same (37.8%), and RSCU values showed that they all had the same codon prefers, i.e., to use codon ended with A/U (93%) except termination codon. Three variable hotspots, i.e., ycf4-cemA, ndhF-rpl32-trnL and ccsA-ndhD, and a total of 152-156 simple sequence repeats (SSR) were identified. The nonsynonymous (Ka)/synonymous (Ks) ratio was calculated, and cemA and ycf2 genes are important indicators of gene evolution. The phylogenetic analyses of the family Anacardiaceae showed that the eight genera were grouped into three clusters, and supported the monophyly of the subfamilies and all the genera. The accessions of five Rhus species formed four clusters, while, one individual of R. typhina grouped with the R. glabra accessions instead of clustering into the two other individuals of R. typhina in the subgenus Rhus, which showed a paraphyletic relationship. CONCLUSIONS: Comparing the complete chloroplast genomes of the Rhus species, it was found that most SSRs were A/T rich and located in the intergenic spacer, and the nucleotide divergence exhibited higher levels in the non-coding region than in the coding region. The Ka/Ks ratio of cemA gene was > 1 for species collected in America, while it was < 1 for other species in China, which dedicated that the Rhus species from North America and East Asia have different evolutionary pressure. The phylogenetic analysis of the complete chloroplast genome clarified the Rhus placement and relationship. The results obtained in this study are expected to provide valuable genetic resources to perform species identification, molecular breeding, and intraspecific diversity of the Rhus species.

Genetic diversity, asexual reproduction and conservation of the edible fruit tree Spondias purpurea L. (Anacardiaceae) in the Costa Rican tropical dry forest.

The term circa situm has been used to describe different conservation strategies within agricultural landscapes. Circa situm conserves planted or remnant species in farmlands, where natural vegetation has been modified through anthropogenic intervention. It has been proposed that trees planted or retained under circa situm conditions may contribute to maintaining genetic diversity, however information on the role of this strategy in preserving genetic diversity is scarce. The aim of this study was to determine the levels of genetic diversity and structure, and mating patterns in planted and unmanaged stands of the tropical fruit tree Spondias purpurea L. in north western Costa Rica. In three localities, we used seven polymorphic microsatellite loci and genotyped 201 adults and 648 seeds from planted and wild stands. We found no differences in genetic diversity among planted and wild stands. Genetic structure analysis revealed that gene flow occurs among planted and wild stands within localities. Clones were present and their diversity and evenness were both high and similar between planted and wild stands. The number of pollen donors per progeny array was low (Nep = 1.01) which resulted in high levels of correlated paternity (rp = 0.9). Asexual seeds were found in 4.6% of the progeny arrays, which had multilocus genotypes that were identical to the maternal trees. Our results show that although planted stands under circa situm conditions can maintain similar levels of genetic diversity than wild stands, the low number of sires and asexual seed formation could threaten the long term persistence of populations.

Genetic and chemodiversity in native populations of Schinus terebinthifolia Raddi along the Brazilian Atlantic forest.

Schinus terebinthifolia is a species native to different ecoregions in the Brazilian Atlantic Forest. The plant is listed on the National Relation of Medicinal Plants and recommended as phytomedicine, however while extractive exploitation prevails as the main route of raw material a significant variation of compounds will be detected. To assure the expansion of productive chain it is important to start by studying population diversity and chemical variations. We used SSR markers for studies of genetic structure among populations from dense ombrophilous forest (ES); the deciduous seasonal forest (SM); the savanna (DOU) and the sandbanks (ITA and MSP), and compared the results to their chemical profiles of essential oil. Genetic structure revealed differences among populations and significant fixation rates. Pairwise studies and Bayesian analysis showed similarities between ITA and SM and between DOU and MSP, proving that the patterns of distribution for the species do not follow the isolation by distance or similarity by environmental conditions. The comparison between PCA of genotypes and chemodiversity reinforces the unique profile for each population despite the environmental similarity observed and genetic analysis. The most divergent genotype and chemical group was found at the ombrophilous forest, strong evidence that we should undertake conservation efforts to prevent losses of biodiversity in that area.

Spondias mombin (Anarcadiaceae): molecular characterization and conservation / Spondias mombin (Anarcadiaceae): caracterización molecular y conservación

Abstract Introduction: The fruit of the yellow mombin (Spondias mombin L.) is notable due to its sensory and functional qualities. However, there is little knowledge regarding the genetic diversity of this species, and this would aid the implantation of the cultivation of the fruit as a crop, since current production is based on extractivism. Objective: Evaluate the diversity and genetic structure of natural populations of S. mombin in the state of Mato Grosso, Brazil, through microsatellite molecular markers in order to assist in the implementation of conservation strategies and the collection of genetic resources. Methods: A total of 139 S. mombin individuals were sampled in ten natural populations. PCR amplifications were performed with seven fluorescence-marked microsatellite primers. Genetic diversity was evaluated by the number of alleles, expected (He) and observed heterozygosity (Ho), polymorphic information content (PIC), fixation index (ƒ), rare and exclusive alleles. The genetic structure was evaluated using AMOVA, UPGMA dendrogram and Bayesian statistical analysis. Results: 46 alleles were amplified, which had an average of 6.6 alleles per locus. He was higher than Ho and f was positive, indicating the presence of inbreeding. The PIC ranged from 0.048 to 0.700, and only two loci were poorly informative. We found 27 rare alleles and 16 unique alleles. The largest component of variation was intrapopulational (90.64 %). The estimated gene flow was 1.99, which indicates that there is no genetic isolation between populations, and justifies the FST value (0.0963). The ten populations were grouped into two groups, and two populations constituted an isolated group. The Mantel test demonstrated that the genetic structure is not related to the geographic distance between populations. Conclusion: There is genetic diversity in the populations of S. mombin, which must be conserved in situ or ex situ, due to the diversity they present and because they are promising sources for collection of germplasm.

Resumen Introducción: El fruto amarillo del jobo o yuplón (Spondias mombin L.) destaca por sus cualidades sensoriales y funcionales. Sin embargo, existe poco conocimiento sobre la diversidad genética de esta especie, lo que ayudaría a la implantación del cultivo del fruto como cultivo, ya que la producción actual se basa en el extractivismo. Objetivo: Evaluar la diversidad y estructura genética de poblaciones naturales de S. mombin en el estado de Mato Grosso, Brasil, a través de marcadores moleculares microsatélites para ayudar en la implementación de estrategias de conservación y recolección de recursos genéticos. Métodos: Se muestrearon un total de 139 individuos de S. mombin en diez poblaciones naturales. Las amplificaciones por PCR se realizaron con siete cebadores de microsatélites marcados con fluorescencia. La diversidad genética se evaluó por el número de alelos, heterocigosidad esperada (He) y observada (Ho), contenido de información polimórfica (PIC), índice de fijación (ƒ), alelos raros y exclusivos. La estructura genética se evaluó mediante AMOVA, dendrograma UPGMA y análisis estadístico bayesiano. Resultados: Se amplificaron 46 alelos, los cuales tenían un promedio de 6.6 alelos por locus. Fue más alto que Ho y f positivo, lo que indica la presencia de endogamia. El PIC osciló entre 0.048 y 0.700, y solo dos loci fueron poco informativos. Encontramos 27 alelos raros y 16 alelos únicos. El mayor componente de variación fue intrapoblacional (90.64 %). El flujo de genes estimado fue de 1.99, lo que indica que no hay aislamiento genético entre poblaciones y justifica el valor de FST (0.0963). Las diez poblaciones se agruparon en dos grupos y dos poblaciones constituyeron un grupo aislado. La prueba de Mantel demostró que la estructura genética no está relacionada con la distancia geográfica entre poblaciones. Conclusión: Existe diversidad genética en las poblaciones de S. mombin, la cual debe ser conservada in situ o ex situ, por la diversidad que presentan y porque son fuentes promisorias para la recolección de germoplasma.

Diversity and genetic structure of Spondias tuberosa (Anacardiaceae) accessions based on microsatellite loci / Diversidad y estructura genética de "accesiones" de Spondias tuberosa (Anacardiaceae) basadas en loci de microsatélites

Abstract Introduction: Spondias tuberosa is a tree endemic to the semiarid region of Brazil with fruticulture potential. Objective: To estimate the diversity and genetic structure of S. tuberosa accessions from four areas of the semiarid region of Brazil, in order to facilitate conservation genetic resources studies in this species. Methods: DNA was extracted, using the CTAB 2x method, from leaf samples of 24 accessions of S. tuberosa available in the germplasm bank at Embrapa Semiárido, Brazil. Ten microsatellite loci were used in this study. Results: The UPGMA dendrogram, generated with a Jaccard coefficient similarity matrix, contains four groups at a 0.44 cutoff point. The similarity coefficient ranged from 0.30 to 0.84, indicating great divergence among the accessions. A Bayesian analysis conducted with the software Structure suggests there are two subpopulations, one formed by accessions from the Januária region and another by accessions from the Juazeiro, Uauá and Petrolina regions. The ΦST value of 0.12 for the analysis of molecular variance indicates moderate genetic differentiation among the four populations, suggesting that the genetic variability is moderately structured in function of region. Conclusions: Together, the analyses indicate that the genetic diversity of S. tuberosa is not uniformly distributed in the studied regions. Thus, germplasm from a greater number of populations should be collected to increase the germplasm bank genetic diversity of the species.

Resumen Introducción: Spondias tuberosa es un árbol endémico de la región semiárida de Brasil con potencial frutícola. Objetivo: Estimar la diversidad y caracterizar la estructura genética de accesiones de S. tuberosa en cuatro áreas del semiárido brasileño, para así facilitar estudios de conservación de recursos genéticos de esta especie. Metodología: El ADN fue extraído utilizando el método CTAB 2x a partir de muestras de hojas de 24 accesiones de S. tuberosa disponibles en el banco de germoplasma de Embrapa Semiárido, Brasil. Diez loci de microsatélites fueron usados en este estudio. Resultados: El dendrograma UPGMA generado con una matriz de similitud de coeficientes de Jaccard, formó cuatro grupos con punto de corte en 0.44. El coeficiente de similitud osciló entre 0.30 y 0.84, indicando una gran divergencia entre las accesiones. El análisis Bayesiano realizado en el software Structure sugiere la existencia de dos subpoblaciones, una formada por las accesiones de la región de Januária y otra derivada de las regiones de Juazeiro, Uauá y Petrolina. El valor de ΦST de 0.12 derivado del análisis molecular de la varianza indica moderada variación genética entre las cuatro poblaciones, sugiriendo que la variabilidad genética se estructura moderadamente en función de la región. Conclusiones: Los análisis en conjunto indican que la diversidad genética de S. tuberosa no se encuentra distribuida uniformemente en las regiones estudiadas. Por lo tanto, se debe recolectar germoplasma de un mayor número de poblaciones para aumentar la diversidad genética del banco actual de la especie.

Pretreatment of Mango (Mangifera indica L. Anacardiaceae) Seed Husk for Bioethanol Production by Dilute Acid Treatment and Enzymatic Hydrolysis.

One of the targets of the Sustainable Development Goals is clean and affordable energy. This is also the aim of the Biofuels Act of 2007 in the Philippines. However, this law is confronted with challenges such as the limitation of lignocellulosic feedstock, specifically available for bioethanol production. The present study sought to address the issue by exploring the potential of mango seed husk (MSH), a by-product of the mango industry, in bioethanol production. MSH is considered a waste material and its utilization also permit value-addition as this can serve as an alternative and affordable source of feedstock in energy production. Two pretreatment strategies are employed to exploit the cellulose and hemicellulose content of MSH, namely, dilute acid treatment and enzymatic hydrolysis. Results show that the %H2SO4 resulting in the highest glucose concentration and yield is 4% v/v at 95 °C hydrolysis temperature, 1:10 (w/v) solid-to-solvent ratio, and 60-min hydrolysis time. For enzymatic hydrolysis using a commercial enzyme preparation, the reaction time up to 72 h did not affect glucose concentration and yield at the following conditions: 50 °C hydrolysis temperature, 150 rpm, pH 5.0, 10% solids loading, and 4% enzyme loading. This could be attributed to the lignin and non-structural compounds present in MSHs. However, a combined process strategy of dilute acid pretreatment followed by enzymatic hydrolysis in the pretreatment of MSH contributes to an increased concentration and yield of sugars in the hydrolysates, which is advantageous for bioethanol production. Graphical Abstract.

Isolation and characterization of microsatellite loci in Astroniumgraveolens (Anacardiaceae) and cross amplification in related species.

Microsatellite markers are a useful genetic tool to answer ecological and conservation genetics questions. Microsatellite primers were developed and characterized to evaluate forest fragmentation effects on genetic structure, diversity and gene flow patterns in the dioecious tropical tree Astroniumgraveolens (Anacardiaceae). Using genomic library enrichment, sixteen microsatellite loci were developed for A.graveolens. Polymorphism was evaluated in 80 individuals from four localities in Mexico. The overall number of alleles detected was 63, average alleles per locus 3.9 with a range from one to 11 per locus. Cross amplification trails on related species in the Anacardiaceae family: Spondiaspurpurea and Amphipterygiumadstringens, achieved successful amplification for all primers. Microsatellite markers described here are the first to be characterized for A.graveolens. These genetic markers will be a useful tool to assess the genetic consequences of habitat fragmentation and selective logging on A.graveolens populations. Cross amplification success on S.purpureaandA.adstringens suggest that they may be used for population genetic studies in other species within the family.

Janzen-Connell effects shape gene flow patterns and realized fitness in the tropical dioecious tree Spondias purpurea (ANACARDIACEAE).

Pollination and seed dispersal patterns determine gene flow within plant populations. In tropical forests, a high proportion of trees are dioecious, insect pollinated and dispersed by vertebrates. Dispersal vectors and density dependent factors may modulate realized gene flow and influence the magnitude of Fine Scale Genetic Structure (FSGS), affecting individual fitness. Spondias purpurea is a vertebrate-dispersed, insect-pollinated dioecious tropical tree. We assessed the influence of sex ratio, effective and realized gene flow on genetic diversity, FSGS and individual fitness within a 30 ha plot in the tropical dry forest reserve of Chamela-Cuixmala, Mexico. All individuals within the plot were tagged, geo-referenced and sampled for genetic analysis. We measured dbh and monitored sex expression during two reproductive seasons for all individuals. We collected seeds directly from maternal trees for effective pollen dispersal analysis, and analyzed established seedlings to assess realized pollen and seed dispersal. Nine microsatellite loci were used to describe genetic diversity parameters, FSGS and gene flow patterns among different size classes. A total of 354 individuals were located and classified into three size classes based on their dbh (<10, 10-20, and >20 cm). Population sex ratios were male biased and diametric size distributions differed among sexes, these differences may be the result of precocious male reproduction at early stages. Autocorrelation analyses indicate low FSGS (Fj <0.07) across all size classes. Long realized pollen and seed dispersal and differences among effective and realized gene flow were detected. In our study site low FSGS is associated with high gene flow levels. Effective and realized gene flow indicate a population recruitment curve indicating Janzen-Connell effects and suggesting fitness advantages for long-distance pollen and seed dispersal events.

Complete mitochondrial genomes of the Spondias tuberosa Arr. Cam and Spondias mombin L. reveal highly repetitive DNA sequences.

Mitogenomes in plants are well-known as exhibiting high diversity in genome size architecture and repetitive DNA sequences. In this research study, we report on the complete mitochondrial genomes of S. tuberosa and S. mombin using Illumina paired-end and mate-pair end reads. These genomes were obtained by a combination of methods of de novo assembly and contig extension. The mitogenomes of S. tuberosa and S. mombin showed 779,106 bp and 685,788 bp in length, with a total of 35 genes. Genome comparisons showed many rearrangements that were mediated by repetitive DNA, and also high incorporation of DNA from chloroplast. In summary, we demonstrate: (1) first complete mitochondrial genomes for the genus Spondias; (2) the synteny between S. tuberosa and S. mombin showed rearrangements, mediated by repetitive DNA; (3) that gene content in Spondias mitogenomes is highly conserved; and (4) the high incorporation DNA from chloroplast genome, (5) the mitogenome size is due intergenic spacers and (6) the non-tandem repeats contributes for giant intergenic spacers.

Isolation and characterization of microsatellites loci in Spondias purpurea (Anacardiaceae) and cross amplification in congeneric species.

Microsatellite markers provide high polymorphism levels, useful to study genetic diversity and gene flow patterns in plant populations. Here we develop and characterize microsatellite primers to evaluate patterns of genetic structure and diversity, and gene flow levels in the dioecious tropical tree Spondias purpurea (Anacardiaceae). Twenty-four microsatellite primers were developed for Spondias purpurea. Polymorphism was evaluated in 139 individuals from three localities in Mexico. Ten loci were polymorphic. The number of alleles ranged between two and 21, the average number of alleles was 5.88. Cross-amplification trials on S. mombin, S. radlkoferi, Astronium graveolens and Amphipterygium adstringens achieved successful amplification for only six microsatellites in S. mombin and S. radlkoferi. Microsatellites developed for S. purpurea will be a useful tool to estimate genetic diversity within and among populations, as well as to assess the consequences of habitat fragmentation on gene flow patterns of this species.

Genome survey and development of 18 microsatellite markers to assess genetic diversity in Spondias tuberosa Arruda Câmara (Anacardiaceae) and cross-amplification in congeneric species.

Spondias tuberosa Arr. Cam belongs to the Anacardiaceae family, an economically important family of plants whose fruits are consumed by humans and animals. The aim of this study was to develop microsatellite markers using sequences from high-throughput sequencing and a magnetic bead enrichment method. The sequences were used to obtain contigs with a minimum of 500 nucleotides using Ray software and the mining of the simple sequence repeats (SSR) was performed with Phobos software, while the primers were designed by Primer3. We developed 18 polymorphic nuclear microsatellite markers and successfully cross-amplified them to three Spondias species. In S. tuberosa, the alleles ranged from 2 to 5 for each locus and Hardy-Weinberg equilibrium was found for 16 loci, with an expected and observed heterozygosity at 0.095-0.755 and 0.1-0.75, respectively. Cross-transferability was obtained for all loci in S. bahiensis, S. dulcis and S. purpurea. We concluded that the microsatellite markers developed in this study are useful in genetic population and conservation studies, as well as for investigating the hybrid origins of Spondias species.

Development of microsatellite markers for Myracrodruon urundeuva (F.F. & M.F. Allemão), a highly endangered species from tropical forest based on next-generation sequencing.

Myracrodruon urundeuva is a tree species of high economic importance due the strength and durability of its wood. Threatened of extinction in Brazil, it is present only in a few forest remnants, mostly in conservation units. Currently, there is little information on the genetic diversity of natural populations in Brazil and even less information about the genome of this species. Here, new species-specific microsatellite loci were developed based on next-generation sequencing (Illumina). More than 100,000 loci were identified in the run, with di- to hexanucleotides motifs. Of these, 20 loci were selected for validation in 30 individuals, with 15 successfully polymorphic loci detected. The number of alleles ranged among loci from 3 to 16, with an average of 7.73, expected (H e ) and observed (H o ) heterozygosity ranged from 0.246 to 0.902 and from 0.103 to 0.867, respectively. These results point out that these new set of markers has a great potential for use in population genetic studies for genetic conservation of the species.

Structure and genetic diversity of Anacardium humile (Anacardiaceae): a tropical shrub.

Anacardium humile Saint Hilaire is a tropical shrub native to the Cerrado biome. It is a fruiting species with biological, medicinal, and socioeconomic significance. Thus, knowing how the genetic variability of natural populations is organized allows for the establishment of strategies for conservation and the sustainable use of the species and its biome. Six microsatellite loci previously developed from Anacardium occidentale were used to investigate the spatial genetic structure and genetic diversity of eight natural A. humile populations based on analyses of 242 adult plants. The results obtained indicate that these populations show a high level of genetic diversity (expected heterozygosity = 0.710). The endogamy coefficient was positive and significant for most populations, with a mean of 0.142 (P = 0.001). The genetic differentiation between populations was low (θ = 0.075 and GST = 0.066) but significant (P = 0.0001). The genotypes of five of the eight populations were non-randomly distributed with clusters of related plants for which the coancestry values were positive and significant. These populations exhibited high and significant endogamy indices. The results obtained for A. humile populations show that genetic conservation programs should be implemented to maintain this species.

Landscape genomics reveal that ecological character determines adaptation: a case study in smoke tree (Cotinus coggygria Scop.).

BACKGROUND: The adaptive evolution of species response to environment are the key issues in molecular ecology and evolutionary biology. The direction of adaptive differentiation of species in regions lacking strong selection pressure is usually diverse. However, the driving mechanism of the diverse adaptive differentiation for regional species is still undetermined to date. In this study, we used landscape genomics modelling to infer the adaptive evolution of Cotinus coggygria in China's warm-temperate zone. RESULTS: Using fifteen natural populations and nine start codon targeted (SCoT) markers, a total of 1131 unambiguous loci were yielded. Our results showed two genetic groups existed in the fifteen natural populations of C. coggygria, which is due to the divergent selection driven by six environmental factors. Environmental association analyses revealed the environmental variables related to precipitation were associated with high numbers of environment-associated loci. CONCLUSIONS: Our results indicated that the ecological characters of C. coggygria, i.e. avoiding wetness and tolerating drought, determine its adaptive evolution. This study provides a reference that ecological character determines the adaptive evolution of species in regions lacking strong selection pressure.

Restoration over time and sustainability of Schinus terebinthifolius Raddi.

The success of recovery programs on degraded areas is dependent on the genetic material to be used, which should present heterozygosity and genetic diversity in native and recovered populations. This study was carried out to evaluate the model efficiency to enable the recovery of a degraded area of the Lower São Francisco, Sergipe, Brazil. The target species for this study was Schinus terebinthifolius Raddi. Three populations were analyzed, the recovered area, seed-tree source population, and native tree population border established to the recovered area. The random amplified polymorphic DNA (RAPD) markers were used for diversity analysis. Genetic structure was estimated to evaluate the level of genetic variability existent in each population. There was no correlation between the spatial distribution and the genetic distances for all trees of the recovered area. The heterozygosity present in the recovered population was higher than the native tree population. The seed-tree source population presents genetic bottlenecks. Three clusters were suggested (ΔK = 3) with non-genetic structure. High intra-population genetic variability and inter-population differentiation are present. However, gene flow may also introduce potentially adaptive alleles in the populations of the recovered area, and the native population is necessary to ensure the sustainability and maintenance of the populations by allelic exchange.

A set of novel microsatellite markers developed for an economically important tree, Dracontomelon duperreanum, in China.

Dracontomelon duperreanum, the most representative species of the family Anacardiaceae, is an important multipurpose tree in China and Vietnam. However, no genetic diversity studies have been reported on this species. In this study, we identified 11 microsatellite markers for D. duperreanum by using the restriction-site-associated DNA sequencing (RAD-seq) method and examined their polymorphisms in 22 samples obtained from the South China Botanical Garden, South China. We could detect only two or three alleles for each microsatellite marker. The mean observed and expected heterozygosities were 0.41 and 0.39, respectively, which were lower than those reported for the species with similar life history forms. These relatively low genetic diversities in this common plant species are unexpected and might have resulted from its extensive cultivation. To our knowledge, this is the first report of microsatellite markers in the genus Dracontomelon. These microsatellite markers will be valuable for studying the genetic diversities and structures in D. duperreanum and other Dracontomelon species.

Comparison of two DNA extraction protocols from leave samples of Cotinus coggygria, Citrus sinensis and Genus juglans.

High quality DNA is essential for molecular research. Secondary metabolites can affect the quantity and quality DNA. In current research two DNA isolation methods including CTAB and Delaporta (protocols 1 & 2 respectively) were applied in three leave samples from Cotinus coggygria, Citrus sinensis and Genus juglans that their leaves are rich of secondary metabolites. We successfully isolated DNA from C. coggygria, C. sinensis and Genus Juglans using the two protocols described above. Good quality DNA was isolated from C. coggygria, C. sinensis and Genus Juglans using protocol 1, while protocol 2 failed to produce usable DNA from these sources. The highest amount of DNA (1.3-1.6) was obtained from them using protocol 1. As we discovered, procedure 1 may work better for plants with secondary metabolites.

Genetic diversity of Cajazeira (Spondias mombin L.) in three geographic regions.

Cajazeira (Spondias mombin L.), of the family Anacardiaceae, is a species of fruit tree found in the Amazon region with fruits that have excellent prospects for commercial use. We aimed to evaluate the genetic diversity within and among natural populations of S. mombin, with natural occurrence in northern Mato Grosso State, by using inter-simple sequence repeat (ISSR) markers. Overall, 126 individuals were evaluated from three populations located Alta Floresta (AFL) 42, Marcelândia (MAR) 41, and Nova Bandeirantes (NBA) 43. The individuals were genotyped with 14 ISSR primers, which amplified 99 fragments. All markers, with the exception of DiGA3'A, presented a polymorphic information content above 0.25, and thus, are recommended for diversity analyses in S. mombin. Genetic diversity of the AFL [Nei's diversity (H) = 0.2430 and Shannon index (I) = 0.3547] and MAR (H = 0.2062 and I = 0.2993) populations was higher when compared to the NBA population, which presented the lowest genetic diversity (H = 0.2002 and I = 0.2957). Analysis of molecular variance showed that 77.38% of the total genetic variation is found within populations while 22.62% is found among populations. AFL and NBA are genetically the most similar populations and also the closest "Structure" revealed genetic diversity among the genotypes of each population. As there is genetic variability in both populations, and there are no genetically identical individuals, both populations can be a source of genotypes for germplasm banks and for future commercial fruitful plantations S. mombin.

Pressurized liquid extraction of anthocyanins and biflavonoids from Schinus terebinthifolius Raddi: A multivariate optimization.

Response surface methodology was employed to investigate the effects of pressurized liquid extraction (PLE) parameters on the recovery of phenolic compounds (anthocyanins, biflavonoids) from Brazilian pepper (Schinus terebinthifolius Raddi) fruits. The effects of temperature, static time, and ethanol as well as acid concentration on the polyphenol yield were described well by quadratic models (p<0.0001). A significant influence of the ethanol concentration (p<0.0001) and several interactions (p<0.05) were identified. Identification of the biflavonoid I3',II8-binaringenin in drupes of S. terebinthifolius was achieved by UHPLC-MS(2). Interestingly, at high extraction temperatures (>75°C), an artifact occurred and was tentatively identified as a diastereomer of I3',II8-binaringenin. Multivariate optimization led to high yields of phenolic compounds from the exocarp/drupes at 100/75°C, 10/10min, 54.5/54.2% ethanol, and 5/0.03% acetic acid. This study demonstrates that PLE is well suited for the extraction of phenolic compounds from S. terebinthifolius and can efficiently be optimized by response surface methodology.

DNA barcode for the identification of the sand fly Lutzomyia longipalpis plant feeding preferences in a tropical urban environment.

Little is known about the feeding behavior of hematophagous insects that require plant sugar to complete their life cycles. We studied plant feeding of Lutzomyia longipalpis sand flies, known vectors of Leishmania infantum/chagasi parasites, in a Brazilian city endemic with visceral leishmaniasis. The DNA barcode technique was applied to identify plant food source of wild-caught L. longipalpis using specific primers for a locus from the chloroplast genome, ribulose diphosphate carboxylase. DNA from all trees or shrubs within a 100-meter radius from the trap were collected to build a barcode reference library. While plants from the Anacardiaceae and Meliaceae families were the most abundant at the sampling site (25.4% and 12.7% of the local plant population, respectively), DNA from these plant families was found in few flies; in contrast, despite its low abundance (2.9%), DNA from the Fabaceae family was detected in 94.7% of the sand flies. The proportion of sand flies testing positive for DNA from a given plant family was not significantly associated with abundance, distance from the trap, or average crown expansion of plants from that family. The data suggest that there may indeed be a feeding preference of L. longipalpis for plants in the Fabaceae family.