Direct molecular profiling of minicircle signatures and lineages of Trypanosoma cruzi bloodstream populations causing congenital Chagas disease

Congenital transmission of Trypanosoma cruzi may occur in some or all the gestations from a T. cruzi-infected mother. Variable rates of congenital transmission have been reported in different geographical areas where different parasitic strains predominate, suggesting that parasitic genotypes might play a role in the risk of congenital transmission. Moreover, in cases of transmission it is unknown if the whole maternal T. cruzi population or certain clones are preferentially transmitted by the transplacental route. In this study, bloodstream T. cruzi lineages were identified in blood samples from congenitally infected children, transmitting and non-transmitting mothers and unrelated Chagas disease patients, using improved PCR strategies targeted to nuclear genomic markers. T. cruzi IId was the prevalent genotype among 36/38 PCR-positive congenitally infected infants, 5/5 mothers who transmitted congenital Chagas disease, 12/13 mothers who delivered non-infected children and 28/34 unrelated Chagas disease patients, all coming from endemic localities of Argentina and Bolivia. These figures indicate no association between a particular genotype and vertical transmission. Furthermore, minicircle signatures from the maternal and infants' bloodstream trypanosomes were profiled by restriction fragment length polymorphism of the 330-bp PCR-amplified variable regions in seven cases of mothers and congenitally infected infants. Minicircle signatures were nearly identical between each mother and her infant/s and unique to each mother-infant/s case, a feature that was also observed in twin deliveries. Moreover, allelic size polymorphism analysis of microsatellite loci from populations transmitted to twins showed that all clones from the maternal polyclonal population were equally infective to both siblings.     

Barcoding quarantine nematodes and their close relatives: an update on the QBOL-project

Identification of plant pests, in particular quarantine species, needs to be fast and accurate to enable timely plant protection measures. In addition, a false diagnosis can cause serious financial losses for trade and producers. It is now well established that genetically based diagnosis is a reliable alternative to the classical identification procedures generally based on morphological features, which usually require expert taxonomic skills. On the other hand, genetic diagnosis through the use of DNA-barcodes, i.e. stretches of DNA that contain taxon-specific information, can be performed by any skilled laboratory worker. The European Union 7th framework project QBOL aims to establish DNA-barcodes for all European quarantine organisms as well as their close relatives. The results and protocols will be disseminated in the publicly available and curated database Q-BANK. To enable genetically based identification requires knowledge of the genetic variation both within and between the species of interest as well as their close relatives. For the nematodes, several gene regions (the COI, COII, SSU, LSU and RNA polymerase subunit II) are being evaluated for their barcoding potential.     

Isolation, functional, and partial biochemical characterization of galatrox, an acidic lectin from Bothrops atrox snake venom

Snake venom lectins have been studied in regard to their chemical structure and biological functions. However, little is known about lectins isolated from Bothrops atrox snake venom. We report here the isolation and partial functional and biochemical characterization of an acidic glycan-binding protein called galatrox from this venom. This lectin was purified by affinity chromatography using a lactosyl-sepharose column, and its homogeneity and molecular mass were evaluated by high-performance liquid chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry. The purified galatrox was homogeneous and characterized as an acidic protein (pI 5.2) with a monomeric and dimeric molecular mass of 16.2 and 32.5 kDa, respectively. Alignment of N-terminal and internal amino acid sequences of galatrox indicated that this protein exhibits high homology to other C-type snake venom lectins. Galatrox showed optimal hemagglutinating activity at a concentration of 100 μg/ml and this effect was drastically inhibited by lactose, ethylenediaminetetraacetic acid, and heating, which confirmed galatrox's lectin activity. While galatrox failed to induce the same level of paw edema or mast cell degranulation as B. atrox crude venom, galatrox did alter cellular viability, which suggested that galatrox might contribute to venom toxicity by directly inducing cell death.     

Phylum-wide analysis of SSU rDNA reveals deep phylogenetic relationships among nematodes and accelerated evolution toward crown Clades

Inference of evolutionary relationships between nematodes is severely hampered by their conserved morphology, the high frequency of homoplasy, and the scarcity of phylum-wide molecular data. To study the origin of nematode radiation and to unravel the phylogenetic relationships between distantly related species, 339 nearly full-length small-subunit rDNA sequences were analyzed from a diverse range of nematodes. Bayesian inference revealed a backbone comprising 12 consecutive dichotomies that subdivided the phylum Nematoda into 12 clades. The most basal clade is dominated by the subclass Enoplia, and members of the order Triplonchida occupy positions most close to the common ancestor of the nematodes. Crown Clades 8-12, a group formerly indicated as "Secernentea" that includes Caenorhabditis elegans and virtually all major plant and animal parasites, show significantly higher nucleotide substitution rates than the more basal Clades 1-7. Accelerated substitution rates are associated with parasitic lifestyles (Clades 8 and 12) or short generation times (Clades 9-11). The relatively high substitution rates in the distal clades resulted in numerous autapomorphies that allow in most cases DNA barcode-based species identification. Teratocephalus, a genus comprising terrestrial bacterivores, was shown to be most close to the starting point of Secernentean radiation. Notably, fungal feeding nematodes were exclusively found basal to or as sister taxon next to the 3 groups of plant parasitic nematodes, namely, Trichodoridae, Longidoridae, and Tylenchomorpha. The exclusive common presence of fungivorous and plant parasitic nematodes supports a long-standing hypothesis that states that plant parasitic nematodes arose from fungivorous ancestors.     

Tree species composition and diversity gradients in white-water forests across the Amazon Basin

Aim Attention has increasingly been focused on the floristic variation within forests of the Amazon Basin. Variations in species composition and diversity are poorly understood, especially in Amazonian floodplain forests. We investigated tree species composition, richness and α diversity in the Amazonian white‐water (várzea) forest, looking particularly at: (1) the flood‐level gradient, (2) the successional stage (stand age), and (3) the geographical location of the forests. Location Eastern Amazonia, central Amazonia, equatorial western Amazonia and the southern part of western Amazonia. Methods The data originate from 16 permanent várzea forest plots in the central and western Brazilian Amazon and in the northern Bolivian Amazon. In addition, revised species lists of 28 várzea forest inventories from across the Amazon Basin were used. Most important families and species were determined using importance values. Floristic similarity between plots was calculated to detect similarity variations between forest types and over geographical distances. To check for spatial diversity gradients, α diversity (Fisher) of the plots was correlated with stand age, longitudinal and latitudinal plot location, and flood‐level gradient. Results More than 900 flood‐tolerant tree species were recorded, which indicates that Amazonian várzea forests are the most species‐rich floodplain forests worldwide. The most important plant families recorded also dominate most Neotropical upland forests, and c. 31% of the tree species listed also occur in the uplands. Species distribution and diversity varied: (1) on the flood‐level gradient, with a distinct separation between low‐várzea forests and high‐várzea forests, (2) in relation to natural forest succession, with species‐poor forests in early stages of succession and species‐rich forests in later stages, and (3) as a function of geographical distance between sites, indicating an increasing α diversity from eastern to western Amazonia, and simultaneously from the southern part of western Amazonia to equatorial western Amazonia. Main conclusions The east‐to‐west gradient of increasing species diversity in várzea forests reflects the diversity patterns also described for Amazonian terra firme. Despite the fine‐scale geomorphological heterogeneity of the floodplains, and despite high disturbance of the different forest types by sedimentation and erosion, várzea forests are dominated by a high proportion of generalistic, widely distributed tree species. In contrast to high‐várzea forests, where floristic dissimilarity increases significantly with increasing distance between the sites, low‐várzea forests can exhibit high floristic similarity over large geographical distances. The high várzea may be an important transitional zone for lateral immigration of terra firme species to the floodplains, thus contributing to comparatively high species richness. However, long‐distance dispersal of many low‐várzea trees contributes to comparatively low species richness in highly flooded low várzea.     

Cadinane sesquiterpenoids of Phomopsis cassiae, an endophytic fungus associated with Cassia spectabilis (Leguminosae)

Five cadinane sesquiterpenes derivatives were isolated by bioassay-guided fractionation from Phomopis cassiae, an endophytic fungus isolated from Cassia spectabilis. The structures of the two diastereoisomeric 3,9,12-trihydroxycalamenenes (1, 2); 3,12-dihydroxycalamenene (3); 3,12-dihydroxycadalene (4) and 3,11,12-trihydroxycadalene (5) were established on the basis of analyses of 1D and 2D NMR and HRTOFMS experiments. Antifungal activity of the isolates was evaluated against Cladosporium sphaerospermum and Cladosporium cladosporioides, revealing 5 as the most active compound.     

Nucleosome dynamics

In the 30 years since the discovery of the nucleosome, our picture of it has come into sharp focus. The recent high-resolution structures have provided a wealth of insight into the function of the nucleosome, but they are inherently static. Our current knowledge of how nucleosomes can be reconfigured dynamically is at a much earlier stage. Here, recent advances in the understanding of chromatin structure and dynamics are highlighted. The ways in which different modes of nucleosome reconfiguration are likely to influence each other are discussed, and some of the factors likely to regulate the dynamic properties of nucleosomes are considered.     

Osteogenesis versus chondrogenesis by BMP-2 and BMP-7 in adipose stem cells

Bone morphogenetic proteins (BMPs) initiate, promote, and maintain chondrogenesis and osteogenesis. We hypothesize that BMP-2 induces an osteogenic, and BMP-7 a chondrogenic phenotype in adipose tissue-derived mesenchymal stem cells (AT-MSCs). We compared the effects of a short 15min BMP-2 or BMP-7 (10ng/ml) treatment on osteogenic and chondrogenic differentiation of AT-MSCs. Gene expression was studied 4 and 14 days after BMP-treatment. At day 4 BMP-2, but not BMP-7, stimulated runx-2 and osteopontin gene expression, and at day 14 BMP-7 down-regulated expression of these genes. At day 4 BMP-2 and BMP-7 stimulated biglycan gene expression, which was down-regulated by BMP-7 at day 14. BMP-7 stimulated aggrecan gene expression at day 14. Our data indicate that BMP-2 treatment for 15min induces osteogenic differentiation, whereas BMP-7 stimulates a chondrogenic phenotype of AT-MSCs. Therefore, AT-MSCs triggered for only 15min with BMP-2 or BMP-7 provide a feasible tool for bone and cartilage tissue engineering.     

Spherical vs. granular immobilization support selection and performance on an optical flow cell immunosensor based on the fluorescence of Cyanine-5

A spherical porous glass support Trisoperl (TRISO) with four pore diameters (? 47.8; 55.9; 102.6, and 108.8 nm) was characterized and selected for application in an optical flow cell immunosensor, in comparison with controlled pore glass (CPG). The TRISO support was functionalized with aldehyde and isothiocyanate (-NCS) groups to attach bovine serum albumin and alkaline phosphatase (AP). The TRISO isothiocyanate pore diameter 47.8 nm (TRISO(-NCS) 47.8 nm) showed the better potential to be used in the immunosensor. It immobilized more protein (19.3 mg AP per g support) while presenting an optical performance comparable to the CPG. CPG(-NCS) and TRISO(-NCS) 47.8 nm were tested in the immunosensor model where the saturation of the Goat IgG immobilized in the supports with Monoclonal Anti-Goat IgG conjugated with Cyanine-5 was reached, followed by regeneration with the elution buffer modified PBS pH 2.0. The TRISO(-NCS) 47.8 nm presented lower fluorescence intensity at saturation (around 39 AU) than CPG(-NCS) (150 to 104 AU), but revealed a major advantage related to the uniform arrangement of the spherical particles in the flow cell, generating no significant fluorescence differences between gravity and flow package.