Genetic variability and demographic history of Heliothis virescens (Lepidoptera: Noctuidae) populations from Brazil inferred by mtDNA sequences

Intra-and inter-population genetic variability and the demographic history of Heliothis virescens (F.) populations were evaluated by using mtDNA markers (coxI, coxII and nad6) with samples from the major cotton-and soybean-producing regions in Brazil in the growing seasons 2007/08, 2008/09 and 2009/10. AMOVA indicated low and non-significant genetic structure, regardless of geographical scale, growing season or crop, with most of genetic variation occurring within populations. Clustering analyzes also indicated low genetic differentiation. The haplotype network obtained with combined datasets resulted in 35 haplotypes, with 28 exclusive occurrences, four of them sampled only from soybean fields. The minimum spanning network showed star-shaped structures typical of populations that underwent a recent demographic expansion. The recent expansion was supported by other demographic analyzes, such as the Bayesian skyline plot, the unimodal distribution of paired differences among mitochondrial sequences, and negative and significant values of neutrality tests for the Tajima's D and Fu's F-S parameters. In addition, high values of haplotype diversity ((H) over cap) and low values of nucleotide diversity (pi), combined with a high number of low frequency haplotypes and values of theta(pi)<theta(W), suggested a recent demographic expansion of H. virescens populations in Brazil. This demographic event could be responsible for the low genetic structure currently found; however, haplotypes present uniquely at the same geographic regions and from one specific host plant suggest an initial differentiation among H. virescens populations within Brazil.

Genetic Variability of Beauveria bassiana and a DNA Marker for Environmental Monitoring of a Highly Virulent Isolate Against Cosmopolites sordidus

The banana weevil Cosmopolites sordidus (Germar) is one of a number of pests that attack banana crops. The use of the entomopathogenic fungus Beauveria bassiana as a biological control agent for this pest may contribute towards reducing the application of chemical insecticides on banana crops. In this study, the genetic variability of a collection of Brazilian isolates of B. bassiana was evaluated. Samples were obtained from various geographic regions of Brazil, and from different hosts of the Curculionidae family. Based on the DNA fingerprints generated by RAPD and AFLP, we found that 92 and 88 % of the loci were polymorphic, respectively. The B. bassiana isolates were attributed to two genotypic clusters based on the RAPD data, and to three genotypic clusters, when analyzed with AFLP. The nucleotide sequences of nuclear ribosomal DNA intergenic spacers confirmed that all isolates are in fact B. bassiana. Analysis of molecular variance showed that variability among the isolates was not correlated with geographic origin or hosts. A RAPD-specific marker for isolate CG 1024, which is highly virulent to C. sordidus, was cloned and sequenced. Based on the sequences obtained, specific PCR primers BbasCG1024F (5'-TGC GGC TGA GGA GGA CT-3') and BbasCG1024R (5'-TGC GGC TGA GTG TAG AAC-3') were designed for detecting and monitoring this isolate in the field.

Virulence attributes and genetic variability of oral Candida albicans and Candida tropicalis isolates

The wide spectrum of candidiasis and its clinical importance encourage the research with the purpose of clarifying the mechanisms of pathogenicity and identification of virulence factors of Candida sp. Therefore, the aim of this study was to verify the adhesion capacity, protease activity and genotypic diversity of oral C. albicans and C. tropicalis isolates. The adhesion ability to the extracellular matrix glycoproteins laminin and fibronectin was evaluated using the ELISA technique. The research of proteases was carried out in agar plate containing bovine albumin and through a quantitative method in buffer solution containing haemoglobin. Intra and interspecies polymorphisms was verified through random amplified polymorphic DNA (RAPD) technique. All C. albicans and C. tropicalis isolates binded to immobilised laminin and fibronectin. Ca33 and Ct13 isolates had relative adhesion index significantly higher than the other isolates for both glycoproteins (P < 0.001). Protease activity was observed in all isolates of C. albicans using either the semi-quantitative or quantitative assay. The protease activity of C. tropicalis was better detected through the quantitative assay. The genotypic diversity by RAPD revealed a heterogeneous population in both species. Nevertheless, C. tropicalis presented higher genetic variability than C. albicans strains.

GENETIC VARIABILITY FOR JUVENILE CHARACTERS OF Cedrela fissilis VELL. PROGENIES: A SUBSIDY FOR DEFINITION OF SEED COLLECTION ZONE

The objective of this study was to evaluate the genetic differences among three matrix groups of Cedrela fissilis based on quantitative juvenile variables on a progeny test to define seed collecting zones and use of seeds of this species in the study region as well as to evaluate genetic variability of the sampled material. A progeny test was established in a nursery with seeds from 48 seed trees collected in the municipalities of Rio Negrinho, Mafra and Sao Bento do Sul, state of Santa Catarina, and in the municipalities of Lapa, Rio Negro, Campo do Tenente and Antonio Olinto, state of Parana. Of the collected seed trees, 33 sampled trees were distributed in three sites and 15 trees were dispersed in the studied region. It was used a complete random block design, with 8 replicates and 20 plants per plot. Evaluated data included: emergency rate; seedling base diameter and height (61, 102 and 145 days after the seeds were sowed); seedling survival; number of leaves per seedling; aerial section dry mass and root dry mass; and the foliar area of the third fully expanded leaf measured from the apical meristem. The Maximum Restricted Likelihood Method (REML) was used, using the software SELEGEN for analysis. It was found that the juvenile characters are strongly genetically controlled and they can be used to estimate genetic variability of population samples of Cedrela fissilis. The three groups of trees spatially limited did not significantly differ among each other, allowing to conclude that the three areas are part of the same tree seed transfer zone.

Genetic variability in mitochondrial and nuclear genes of Larus dominicanus (Charadriiformes, Laridae) from the Brazilian coast

Several phylogeographic studies of seabirds have documented low genetic diversity that has been attributed to bottleneck events or individual capacity for dispersal. Few studies have been done in seabirds on the Brazilian coast and all have shown low genetic differentiation on a wide geographic scale. The Kelp Gull is a common species with a wide distribution in the Southern Hemisphere. In this study, we used mitochondrial and nuclear markers to examine the genetic variability of Kelp Gull populations on the Brazilian coast and compared this variability with that of sub-Antarctic island populations of this species. Kelp Gulls showed extremely low genetic variability for nnitochondrial markers (cytb and ATPase) and high diversity for a nuclear locus (intron 7 of the beta-fibrinogen). The intraspecific evolutionary history of Kelp Gulls showed that the variability found in intron 7 of the beta-fibrinogen gene was compatible with the variability expected under neutral evolution but suggested an increase in population size during the last 10,000 years. However, none of the markers revealed evidence of a bottleneck population. These findings indicate that the recent origin of Kelp Gulls is the main explanation for their nuclear diversity, although selective pressure on the mtDNA of this species cannot be discarded.

GENETIC VARIABILITY OF SUGARCANE-ASSOCIATED DIAZOTROPHIC BACTERIA CAPABLE OF INORGANIC PHOSPHATE SOLUBILIZING

The sugarcane is a culture of great importance for the Brazilian agriculture. Every year this culture consumes great amounts of nitrogen and phosphate fertilizers. However, the use of plant growth-promoting bacteria can reduce the use of the chemical fertilizers, contributing to the economy and the environment conservation. So, the goal of this study was to select sugarcane-associated diazotrophic bacteria able to solubilize inorganic phosphate and to evaluate the genetic diversity of these bacteria. A total of 68 diazotrophic bacteria, leaf and root endophytic and rizoplane, of three sugarcane varieties. The selection of inorganic phosphate solubilizing diazotrophic bacteria was assayed by the solubilization index (SI) in solid medium containing insoluble phosphate. The genetic variability was analyzed by the BOX-PCR technique. The results showed that 74% of the diazotrophic strains were able to solubilize inorganic phosphate, presenting classes of different SI. The results showed that the vegetal tissue and the genotype plant influenced in the interaction between phosphate solubilizing diazotrophic bacteria and sugarcane plants. BOX-PCR revealed high genetic variability among the strains analyzed. So, sugarcane-associated diazotrophic bacteria express the capacity to solubilize inorganic phosphate and they present high genetic diversity.

Genetic variability of banana with ornamental potential

The Musa germplasm collection at Embrapa Cassava and Fruits detains accessions from different sections of the Musa genus. The objective of the present study was to identify and morphologically characterize banana accessions from the banana germplasm with ornamental potential, as well as to quantify their genetic variability; and identify possible progenitors to be used in breeding aiming to achieve ornamental crossbreeds. The accessions were evaluated with the use of 32 morphological descriptors. Then, they were the following grouped into categories: landscape plants, cut flower, potted plants, and male inflorescence minifruits. The pre-selected accessions presented great genetic variability and ornamental potential for different uses. The accessions of the Rhodochlamys and Callimusa sections were selected to be used as landscape plants, cut flowers, potted plants, male inflorescence and minifruits. Most of the diploids from the Eumusa section evaluated in this study are indicated for the production of ornamental minifruits, except for 'Lidi' and Cici, which can also be indicated as landscape plants. The BB diploids have great potential for the use of the male inflorescence in floral arrangements, and did not offer any other indication.

Morphological and genetic variability in Hippolyte obliquimanus dana, 1852 (Decapoda, Caridea, Hippolytidae) from Brazil and the caribbean sea

Hippolyte obliquimanus is a marine shrimp reported from the Caribbean Sea and Brazil. The literature provides indications for morphological variation between populations from those regions and the species has a troubled taxonomic history. The aims of this study were to analyse morphological and genetic variation in the populations of H. obliquimanus from Brazil and the Caribbean Sea and to verify if those might support separation of H. obliquimanus into two or more species. This hypothesis was tested with the analysis of morphological and genetic data (mitochondrial gene 16S and the barcode region Cytochrome Oxidase I). The material analysed was obtained from samples and from loans of zoological collections. The rostrum as well as pereiopods 3, 4, and 5 were the adult morphological characters that showed variation, but this occurred in samples from both regions, Brazil and the Caribbean Sea. The sequences of the 16S gene were identical among all specimens analysed. There was, however, variation among the sequences of the barcoding gene COI (<2.0%); this divergence separated the specimens into two groups (Brazil versus the Caribbean) and these groups did not share haplotypes. In conclusion, specimens from the regions analysed showed both morphological and genetic variation, but these did not support the separation of H. obliquimanus into two or more species.

Genetic characterization of fig tree mutants with molecular markers

The fig (Ficus carica L.) is a fruit tree of great world importance and, therefore, the genetic improvement becomes an important field of research for better crops, being necessary to gather information on this species, mainly regarding its genetic variability so that appropriate propagation projects and management are made. The improvement programs of fig trees using conventional procedures in order to obtain new cultivars are rare in many countries, such as Brazil, especially due to the little genetic variability and to the difficulties in obtaining plants from gamete fusion once the wasp Blastophaga psenes, responsible for the natural pollinating, is not found in Brazil. In this way, the mutagenic genetic improvement becomes a solution of it. For this reason, in an experiment conducted earlier, fig plants formed by cuttings treated with gamma ray were selected based on their agronomic characteristics of interest. We determined the genetic variability in these fig tree selections, using RAPD and AFLP molecular markers, comparing them to each other and to the Roxo-de-Valinhos, used as the standard. For the reactions of DNA amplification, 140 RAPD primers and 12 primer combinations for AFLP analysis were used. The selections did not differ genetically between themselves and between them and the Roxo-de-Valinhos cultivar. Techniques that can detect polymorphism between treatments, such as DNA sequencing, must be tested. The phenotypic variation of plants may be due to epigenetic variation, necessitating the use of techniques with methylation-sensitive restriction enzymes.

Genetic diversity of polysporic isolates of Moniliophthora perniciosa (Tricholomataceae)

The causal agent of witches' broom disease, Moniliophthora perniciosa is a hemibiotrophic and endemic fungus of the Amazon basin and the most important cocoa disease in Brazil. The purpose of this study was to analyze the genetic diversity of polysporic isolates of M. perniciosa to evaluate the adaptation of the pathogen from different Brazilian regions and its association with different hosts. Polysporic isolates obtained previously in potato dextrose agar cultures of M. perniciosa from different Brazilian states and different hosts (Theobroma cacao, Solanum cernuum, S. paniculatum, S. lycocarpum, Solanum sp, and others) were analyzed by somatic compatibility grouping where the mycelium interactions were distinguished after 4-8 weeks of confrontation between the different isolates of M. perniciosa based on the precipitation line in the transition zone and by protein electrophoresis through SDS-PAGE. The diversity of polysporic isolates of M. perniciosa was grouped according to geographic proximity and respective hosts. The great genetic diversity of M. perniciosa strains from different Brazilian states and hosts favored adaptation in unusual environments and dissemination at long distances generating new biotypes.

Genetic diversity of Stryphnodendron adstringens (Mart.) Coville determined by AFLP molecular markers

Bark extracts of Stryphnodendron adstringens (Mart) Coville a Leguminosae species, well known in Brazil as barbatimao, are popularly used as healing agent. The objective of this work was to determine the genetic diversity of S. adstringens populations and to correlate genetic distances to the production of tannins. S. adstringens accessions from populations found in Cerrado regions in the states of Goias, Minas Gerais and Sao Paulo were analyzed using the AFLP (Amplified Fragment Length Polymorphism) technique. A total of 236 polymorphic bands were scored and higher proportion of genetic diversity was found inter populations (70.9%), rather than intra populations (29.1%). F-ST value was found to be significantly greater than zero (0.2906), demonstrating the complex genetic structure of S. adstringens populations. Accessions collected in Cristalina, GO, showed higher percentage of polymorphic loci (87.3%) and the highest genetic diversity. The lowest genetic variability was detected among accessions from the population growing in Caldas Novas, GO. The genetic distance among populations was estimated using the Unweighted Pair Group Method with Arithmetic Mean (UPGMA), which grouped populations into 3 clusters. Moreover, chemotypes with tannin concentration above 40% showed higher genetic similarity. AFLP analysis proved to be an efficient gene mapping technique to determine the genetic diversity among remaining populations of S. adstringens. Obtained results may be employed to implement further strategies for the conservation of this medicinal plant. (C) 2011 Elsevier Ltd. All rights reserved.

Genetic diversity of Leishmania amazonensis strains isolated in northeastern Brazil as revealed by DNA sequencing, PCR-based analyses and molecular karyotyping

Abstract Background Leishmania (Leishmania) amazonensis infection in man results in a clinical spectrum of disease manifestations ranging from cutaneous to mucosal or visceral involvement. In the present study, we have investigated the genetic variability of 18 L. amazonensis strains isolated in northeastern Brazil from patients with different clinical manifestations of leishmaniasis. Parasite DNA was analyzed by sequencing of the ITS flanking the 5.8 S subunit of the ribosomal RNA genes, by RAPD and SSR-PCR and by PFGE followed by hybridization with gene-specific probes. Results ITS sequencing and PCR-based methods revealed genetic heterogeneity among the L. amazonensis isolates examined and molecular karyotyping also showed variation in the chromosome size of different isolates. Unrooted genetic trees separated strains into different groups. Conclusion These results indicate that L. amazonensis strains isolated from leishmaniasis patients from northeastern Brazil are genetically diverse, however, no correlation between genetic polymorphism and phenotype were found.

Diversity and genetic structure in natural populations of Hancornia speciosa var. speciosa Gomes in northeastern Brazil

Hancornia speciosa Gomes is a fruit tree native from Brazil that belongs to Apocinaceae family, and is popularly known as Mangabeira. Its fruits are widely consumed raw or processed as fruit jam, juices and ice creams, which have made it a target of intense exploitation. The extractive activities and intense human activity on the environment of natural occurrence of H. speciosa has caused genetic erosion in the species and little is known about the ecology or genetic structure of natural populations. The objective of this research was the evaluation of the genetic diversity and genetic structure of H. speciosa var. speciosa. The genetic variability was assessed using 11 allozyme loci with a sample of 164 individuals distributed in six natural populations located in the States of Pernambuco and Alagoas, Northeastern Brazil. The results showed a high level of genetic diversity within the species (e= 0.36) seeing that the most of the genetic variability of H. speciosa var. speciosa is within its natural populations with low difference among populations ( or = 0.081). The inbreeding values within ( = -0.555) and among populations ( =-0.428) were low showing lacking of endogamy and a surplus of heterozygotes. The estimated gene flow ( m ) was high, ranging from 2.20 to 13.18, indicating to be enough to prevent the effects of genetic drift and genetic differentiation among populations. The multivariate analyses indicated that there is a relationship between genetic and geographical distances, which was confirmed by a spatial pattern analysis using Mantel test (r = 0.3598; p = 0.0920) with 1000 random permutations. The high genetic diversity index in these populations indicates potential for in situ genetic conservation.

The potentially toxic diatom Pseudo-nitzschia H. Peragallo in the Parana and Santa Catarina States, Southern Brazil

A taxonomic survey of the diatom Pseudo-nitzschia H. Peragallo was carried out in the Parana and Santa Catarina coasts, Southern Brazil. Samples were taken in various localities along the coastline using vertical hauls from the bottom to the surface. Electron microscopy revealed five species: the potentially toxic P. australis Frenguelli, P calliantha Lundholm, Moestrup & Has le, P multiseries (Has le) Has le and P pungens (Grunow ex Cleve) Has le (plus P pungens var. cingulata Villac), and the non-toxic P linea Lundholm, Has le & G. A. Fryxell. Southern Brazilian strains of P. calliantha and P multiseries have previously been shown to be toxic, raising concerns about a potential contamination of mussels and oysters being commercially grounded in the region. High morphological variability was observed in valve characters of P calliantha and P pungens, in some cases confirmed in the literature. In P calliantha there was a conspicuous differentiation in two morphotypes separated from each other by the width and the shape of the valve, and the density of the poroids. P linea has not previously been found in Brazilian waters, and P pungens var. cingulata is a new record in Western Atlantic waters. Future investigations using molecular techniques will elucidate whether the genetic variability corresponds to the morphological variation and unveil the possible existence of semicryptic species of Pseudo-nitzschia inhabiting the South Brazilian coast.

Molecular epidemiology of the SH (small hydrophobic) gene of human respiratory syncytial virus (HRSV), over 2 consecutive years

Human respiratory syncytial virus (HRSV) strains were isolated from nasopharyngeal aspirates collected from 965 children between 2004 and 2005, yielding 424 positive samples. We sequenced the small hydrophobic protein (SH) gene of 117 strains and compared them with other viruses identified worldwide. Phylogenetic analysis showed a low genetic variability among the isolates but allowed us to classify the viruses into different genotypes for both groups, HRSVA and HRSVB. It is also shown that the novel BA-like genotype was well segregated from the others, indicating that the mutations are not limited to the G gene. (C) 2011 Elsevier B.V. All rights reserved.