Alessandra Maria Moreira Reis

The complete genome sequence of Chromobacterium violaceum reveals remarkable and exploitable bacterial adaptability

Chromobacterium violaceum is one of millions of species of free-living microorganisms that populate the soil and water in the extant areas of tropical biodiversity around the world. Its complete genome sequence reveals (i) extensive alternative pathways for energy generation, (ii) ≈500 ORFs for transport-related proteins, (iii) complex and extensive systems for stress adaptation and motility, and (iv) widespread utilization of quorum sensing for control of inducible systems, all of which underpin the versatility and adaptability of the organism. The genome also contains extensive but incomplete arrays of ORFs coding for proteins associated with mammalian pathogenicity, possibly involved in the occasional but often fatal cases of human C. violaceum infection. There is, in addition, a series of previously unknown but important enzymes and secondary metabolites including paraquat-inducible proteins, drug and heavy-metal-resistance proteins, multiple chitinases, and proteins for the detoxification of xenobiotics that may have biotechnological applications.

Bacterial Community Associated with Healthy and Diseased Reef Coral Mussismilia hispida from Eastern Brazil

In order to characterize the bacterial community diversity associated to mucus of the coral Mussismilia hispida, four 16S rDNA libraries were constructed and 400 clones from each library were analyzed from two healthy colonies, one diseased colony and the surrounding water. Nine bacterial phyla were identified in healthy M. hispida, with a dominance of Proteobacteria, Actinobacteria, Acidobacteria, Lentisphaerae, and Nitrospira. The most commonly found species were related to the genera Azospirillum, Hirschia, Fabibacter, Blastochloris, Stella, Vibrio, Flavobacterium, Ochrobactrum, Terasakiella, Alkalibacter, Staphylococcus, Azospirillum, Propionibacterium, Arcobacter, and Paenibacillus. In contrast, diseased M. hispida had a predominance of one single species of Bacteroidetes, corresponding to more than 70% of the sequences. Rarefaction curves using evolutionary distance of 1% showed a greater decrease in bacterial diversity in the diseased M. hispida, with a reduction of almost 85% in OTUs in comparison to healthy colonies. ∫-Libshuff analyses show that significant p values obtained were <0.0001, demonstrating that the four libraries are significantly different. Furthermore, the sympatric corals M. hispida and Mussismilia braziliensis appear to have different bacterial community compositions according to Principal Component Analysis and Lineage-specific Analysis. Moreover, lineages that contribute to those differences were identified as α-Proteobacteria, Bacteroidetes, and Firmicutes. The results obtained in this study suggest host–microbe co-evolution in Mussismilia, and it was the first study on the diversity of the microbiota of the endemic and endangered of extinction Brazilian coral M. hispida from Abrolhos bank.

Bacterial diversity associated with the Brazilian endemic reef coral Mussismilia braziliensis

Aims:  We performed the first characterization of the microbiota associated with the reef coral Mussismilia braziliensis by means of a culture‐independent approach.

RAPD variation in a germplasm collection of Myracrodruon urundeuva (Anacardiaceae), an endangered tropical tree: Recommendations for conservation

RAPD markers were used to investigate the distribution of genetic variability in a germplasm collection of the dioecious tropical tree Aroeira (Myracrodruon urundeuva) composed of open pollinated maternal half-sib families collected from trees at least 300 meters apart and grouped in nine collection areas throughout its geographical range in Brazil. Open pollinated families from nearby trees from two natural populations were also included in the study. A Principal Coordinate Analysis (PCO) based on genetic similarities estimated with 83 RAPD markers showed no defined clustering among individuals from the same collection areas. However, a matrix correlation test showed low, but significant correlation between genetic and geographic distances (r = 0.17; p < 0.02). An AMOVA showed that 92% of the genetic variation is found within collection areas. Significant variation was found both between regions and between areas within regions although only 3.54 and 4.44% of the variation was found at these levels respectively. An AMOVA on the individuals from the two natural populations resulted in a pattern of distribution congruent to the one found for the collection areas, with 97% of variability within populations. An analysis of 10 open-pollinated half-sib families revealed that between 75% and 89% of the variability is contained within families, suggesting that seeds from one tree represent a significant proportion of the variability found in the population. These results indicate that, either for the establishment of in situ genetic reserves or for ex situ conservation, efforts should be directed to the conservation of several individual trees or collection of open pollinated seed families in a few distant sites. In addition, if resources are available to enlarge the genetic base of the existing collection, expeditions should prioritize collection sites located in areas at significant distances to those already sampled.

Development and characterization of microsatellite markers for the Brazil nut tree Bertholletia excelsa Humb. & Bonpl. (Lecythidaceae).

Twelve polymorphic microsatellite markers were developed for the Brazil nut (Bertholletia excelsa), one of the most valuable non‐timber forest products from the Amazon, based on enrichment protocol. Six to 18 (mean 10.4) alleles per locus were identified and the expected heterozygosity ranged from 0.663 to 0.923 based on a screen of 40 individuals from one population of B. excelsa. The combined probabilities of genetic identity (8.39 × 10−17) and paternity exclusion (0.999999) indicated that multilocus genotypes are likely to be unique allowing precise analyses of genetic structure, gene flow, and mating system of this economically important species.

Use of Targeted Exome Sequencing for Molecular Diagnosis of Skeletal Disorders.

Genetic disorders of the skeleton comprise a large group of more than 450 clinically distinct and genetically heterogeneous diseases associated with mutations in more than 300 genes. Achieving a definitive diagnosis is complicated due to the genetic heterogeneity of these disorders, their individual rarity and their diverse radiographic presentations. We used targeted exome sequencing and designed a 1.4Mb panel for simultaneous testing of more than 4,800 exons in 309 genes involved in skeletal disorders. DNA from 69 individuals from 66 families with a known or suspected clinical diagnosis of a skeletal disorder was analyzed. Of 36 cases with a specific clinical hypothesis with a known genetic basis, mutations were identified for eight cases (22%). Of 20 cases with a suspected skeletal disorder but without a specific diagnosis, four causative mutations were identified. Also included were 11 cases with a specific skeletal disorder but for which there was at the time no known associated gene. For these cases, one mutation was identified in a known skeletal disease genes, and re-evaluation of the clinical phenotype in this case changed the diagnoses from osteodysplasia syndrome to Apert syndrome. These results suggest that the NGS panel provides a fast, accurate and cost-effective molecular diagnostic tool for identifying mutations in a highly genetically heterogeneous set of disorders such as genetic skeletal disorders. The data also stress the importance of a thorough clinical evaluation before DNA sequencing. The strategy should be applicable to other groups of disorders in which the molecular basis is largely known.

Microsatellite markers for Caesalpinia echinata Lam. (Brazilwood), a tree that named a country

Caesalpinia echinata, commonly known as Pau-brasil (Brazilwood), the famous tree that named Brazil is native to the Atlantic forest. Men extensively exploited it ever since discovery and colonial times due to its value as a source of red dye. As a consequence, Brazilwood is a threatened species with populations reduced to small forest fragments. Ten polymorphic microsatellite loci were developed from an enriched genomic library. Using fluorescently-labeled primers, a total of 83 alleles were found after analyzing a sample of 44 trees. These high genetic information content markers should allow detailed investigations of mating systems, gene flow, population structure and paternity in natural populations.

Interspecific discovery and expression profiling of Eucalyptus micro RNAs by deep sequencing

Background Micro RNAs (miRNAs) are a class of small (~21 nucleotide) non-coding RNAs that recently gained much attention due to their perceived role as master regulators of gene expression in Eukaryotes, responsible for fine tuning gene expression regulation and, in plants, has been shown to be involved in a diverse range of biological processes such as plant development and architecture, flowering, cell differentiation and response to biotic and abiotic stresses [1]. The repertoire of expressed miRNAs differs among cell types, tissues, development, environmental condition, etc [2]. Notwithstanding, the exact function of thousands miRNAs sequences present in miRBase [http://www.mirbase.org/] is not elucidated. At this point, discovery and profiling of new and conserved miRNAs are critical in the attempt to understand their function and mechanism. Deep sequencing through next generation sequencing is the methodology of choice for this purpose as its ultra high throughput permits a comprehensive interrogation of the small RNA transcriptome, permitting de novo identification and relative quantification of different small RNA species [3]. Due to its economic importance the Eucalyptus grandis genome has been sequenced by JGI and the annotation of miRNAs is pivotal. In order to provide the first large scale experimental characterization of Eucalyptus miRNAs we performed an Illumina deep sequencing run that allowed us to discover and quantify the miRNA levels in two different tissues – xylem and leaves. Additionally, to get insights of the observed phenotypic differences in wood quality among Eucalyptus species, we characterized the xylem small RNA transcriptome of two different E. globulus individuals and