TESTING MITOCHONDRIAL CAPTURE AND DEEP COALESCENCE IN AMAZONIAN CICHLID FISHES (CICHLIDAE: CICHLA)

Hybridization and introgression have important consequences in evolution, such as increasing the genetic diversity and adaptive potential of a species. One of their most conspicuous footprints is discordance among gene trees or between genes and phenotypes. However, most studies that report introgression fail to disprove the null hypothesis that genetic incongruence may result from stochastic sorting of ancestral allelic polymorphisms. In the case of ancient introgression, these two processes may be especially difficult to distinguish topologically, but they make different predictions about the patterns of coalescence among loci. Here we apply three methods, molecular dating, multispecies coalescent models, and gene tree simulation under coalescence, to compare these two hypotheses that explain the polyphyletic mtDNA of the butterfly peacock bass, Cichla orinocensis. In comparison with a species tree based on 20 unlinked nuclear loci, we determined that mtDNA divergences were too recent to be explained by ancestral polymorphism. Similarly, coalescent species tree branches were significantly shorter when putative introgressed mtDNA was incorporated, and simulations showed the mtDNA topology to be unlikely under lineage sorting only. We conclude that introgression approximately 1.5 million years ago resulted in capture by C. orinocensis of an mtDNA lineage ancestral to the modern subspecies C. oc. monoculus.     

Halilectin 1 (H‐1) and Halilectin 2 (H‐2): two new lectins isolated from the marine sponge Haliclona caerulea

Two new lectins named Halilectin 1 (H‐1) and Halilectin 2 (H‐2) were isolated from the marine sponge Haliclona caerulea using a combination of affinity chromatography on stroma fixed onto Sephadex G‐25 and cation and anion exchange chromatography. H‐1 is a monomeric protein with a molecular mass of 40 kDa estimated using sodium dodecyl sulfate polyacrylamide gel electrophoresis and 15 kDa estimated using a TSK gel. Conversely, H‐2 is a homodimeric protein with 15 kDa monomers linked via weak interactions. H‐1 more effectively agglutinates trypsinized rabbit erythrocytes, whereas H‐2 more effectively agglutinates native rabbit erythrocytes. The hemagglutinating activity of H‐1 could be not inhibited by any tested sugars, but H‐2 was inhibited by orosomucoid and porcine stomach mucin. Neither lectin was dependent on divalent ions. H‐1 was stable at basic pH range and temperatures up to 50 °C, whereas H‐2 was stable at acid pH range and temperatures up to 80 °C. The H. caerulea lectins exhibited dose‐dependent toxicity against Artemia nauplii. Additionally, 76% of the primary structure of H‐2 was determined using tandem mass spectrometry to contain a unique amino acid sequence with no similarity to any members of the animal lectin family. Copyright © 2012 John Wiley & Sons, Ltd.     

Afro-derived Amazonian populations: inferring continental ancestry and population substructure

A panel of Ancestry Informative Markers (AIMs) was used to identify population substructure and estimate individual and overall interethnic admixture in 294 individuals from seven African-derived communities of the Brazilian Amazon. A panel of 48 biallelic markers, representing the insertion (IN) or the deletion (DEL) of small DNA fragments, was employed for this purpose. Overall interethnic admixture estimates showed high miscegenation with other ethnic groups in all populations (between 46% and 64%). The proportion of ancestral genes varied significantly among individuals of the sample: the contribution of African genes varied between 12% and 75%; of European genes between 10% and 73%; and of Amerindians genes between 8% and 66%. The obtained data reveal a high contribution of Amerindian genes in these communities, unlike in other African-derived communities of the Northeast and the South of Brazil. In addition, the majority of the Amerindian contribution may result from the preferential inclusion of indigenous women in the African descent groups. High heterogeneity of the proportion of interethnic admixture among analyzed individuals was found when the proportion of ancestral genes of each individual of the sample was estimated. This heterogeneity is reflected in the fact that four populations can be considered as substructured and that the global African descent sample is possibly formed by two subpopulations.     

Morphophysiological in vitro performance of Brazilian ginseng (Pfaffia glomerata (Spreng.) Pedersen) based on culture medium formulations

Pfaffia glomerata has potential pharmacological and medicinal properties due to the production of a secondary metabolite known as the phytoecdysteroid 20-hydroxyecdysone (20E). There have been increasing efforts for massive in vitro propagation of Pfaffia plants due to high extractivism and overharvesting of this species. Research on the species has shown that photoautotrophic cultivation can improve the production of 20E. In addition, other abiotic factors such as the formulations of culture media can influence the morphophysiological behavior of the plants in vitro. Therefore, the objective of this study was to analyze the morphological and physiological performances of P. glomerata plants in different formulations of culture media, under photoautotrophic and photomixotrophic propagation conditions. Six medium formulations, the Driver and Kuniyuki medium (DKW), Correia et al. medium (JADS), Murashige and Skoog medium (MS), Quoirin and Lepoivre medium (QL), Rugini medium (OM), and Woody Plant medium (WPM), all supplemented with DKW vitamins, 100 mg L⁻¹ myo-inositol, 6.5 g L⁻¹ agar, and with or without 3% (w/v) sucrose, were evaluated. Cultures were maintained at 25 ± 2°C, with a 16 h-photoperiod under 60 μmol m⁻² s⁻¹ of irradiance under a fluorescent lamp for 50 d. Results showed that the presence or absence of sucrose, and the different nutritional formulations influenced growth, photosynthetic pigment content, endogenous levels of sugars, leaf morphology, levels of 20E, and transport of water and minerals in P. glomerata. Notably, OM, DKW, QL, and WPM media promoted higher production of 20E under photomixotrophic growth conditions.

     

Paenibacillus piscarius sp. nov., a novel nitrogen-fixing species isolated from the gut of the armored catfish Parotocinclus maculicauda

A Gram-positive, nitrogen-fixing and endospore-forming strain, designated P121^T, was isolated from the gut of the armored catfish (Parotocinclus maculicauda) and identified as a member of the genus Paenibacillus based on the sequences of the 16S rRNA encoding gene, rpoB, gyrB and nifH genes and phenotypic analyses. The most closely related species to strain P121^T were Paenibacillus rhizoplanae DSM 103993^T, Paenibacillus silagei DSM 101953^T and Paenibacillus borealis DSM 13188^T, with similarity values of 98.9, 98.3 and 97.6%, respectively, based on 16S rRNA gene sequences. Genome sequencing revealed a genome size of 7,513,698 bp, DNA G?+?C content of 53.9 mol% and the presence of the structural nitrogenase encoding genes (nifK, nifD and nifH) and of other nif genes necessary for nitrogen fixation. Digital DNA-DNA hybridization (dDDH) experiments and average nucleotide identity (ANI) analyses between strain P121^T and the type strains of the closest species demonstrated that the highest values were below the thresholds of 70% dDDH (42.3% with P. borealis) and 95% ANI (84.28% with P. silagei) for bacterial species delineation, indicating that strain P121^T represents a distinct species. Its major cellular fatty acid was anteiso-C_15:0 (42.4%), and the major isoprenoid quinone was MK-7. Based on physiological, genomic, biochemical and chemotaxonomic characteristics, we propose that strain P121^T represents a novel species for which the name Paenibacillus piscarius sp. nov. is proposed (type strain?=?DSM 25072?=?LFB-Fiocruz 1636).

     

Evaluation of the novel folate receptor ligand [18F]fluoro-PEG-folate for macrophage targeting in a rat model of arthritis

Introduction

Detection of (subclinical) synovitis is relevant for both early diagnosis and monitoring of therapy of rheumatoid arthritis (RA). Previously, the potential of imaging (sub)clinical arthritis was demonstrated by targeting the translocator protein in activated macrophages using (R)-[¹¹C]PK11195 and positron emission tomography (PET). Images, however, also showed significant peri-articular background activity. The folate receptor (FR)-β is a potential alternative target for imaging activated macrophages. Therefore, the PET tracer [¹⁸F]fluoro-PEG-folate was synthesized and evaluated in both in vitro and ex vivo studies using a methylated BSA induced arthritis model.

Methods

[¹⁸F]fluoro-PEG-folate was synthesized in a two-step procedure. Relative binding affinities of non-radioactive fluoro-PEG-folate, folic acid and naturally circulating 5-methyltetrahydrofolate (5-Me-THF) to FR were determined using KB cells with high expression of FR. Both in vivo [¹⁸F]fluoro-PEG-folate PET and ex vivo tissue distribution studies were performed in arthritic and normal rats and results were compared with those of the established macrophage tracer (R)-[¹¹C]PK11195.

Results

[¹⁸F]fluoro-PEG-folate was synthesized with a purity >97%, a yield of 300 to 1,700 MBq and a specific activity between 40 and 70 GBq/µmol. Relative in vitro binding affinity for FR of F-PEG-folate was 1.8-fold lower than that of folic acid, but 3-fold higher than that of 5-Me-THF. In the rat model, [¹⁸F]fluoro-PEG-folate uptake in arthritic knees was increased compared with both contralateral knees and knees of normal rats. Uptake in arthritic knees could be blocked by an excess of glucosamine-folate, consistent with [¹⁸F]fluoro-PEG-folate being specifically bound to FR. Arthritic knee-to-bone and arthritic knee-to-blood ratios of [¹⁸F]fluoro-PEG-folate were increased compared with those of (R)-[¹¹C]PK11195. Reduction of 5-Me-THF levels in rat plasma to those mimicking human levels increased absolute [¹⁸F]fluoro-PEG-folate uptake in arthritic joints, but without improving target-to-background ratios.

Conclusions

The novel PET tracer [¹⁸F]fluoro-PEG-folate, designed to target FR on activated macrophages provided improved contrast in a rat model of arthritis compared with the accepted macrophage tracer (R)-[¹¹C]PK11195. These results warrant further exploration of [¹⁸F]fluoro-PEG-folate as a putative PET tracer for imaging (sub)clinical arthritis in RA patients.     

A reassessment of the in vitro RBC haemolysis assay with defibrinated sheep blood for the determination of the ocular irritation potential of cosmetic products: comparison with the in vivo Draize rabbit test

We examined the correlation between results obtained from the in vivo Draize test for ocular irritation and in vitro results obtained from the sheep red blood cell (RBC) haemolytic assay, which assesses haemolysis and protein denaturation in erythrocytes, induced by cosmetic products. We sought to validate the haemolytic assay as a preliminary test for identifying highly-irritative products, and also to evaluate the in vitro test as alternative assay for replacement of the in vivo test. In vitro and in vivo analyses were carried out on 19 cosmetic products, in order to correlate the lesions in the ocular structures with three in vitro parameters: (i) the extent of haemolysis (H50); (ii) the protein denaturation index (DI); and (iii) the H50/DI ratio, which reflects the irritation potential (IP). There was significant correlation between maximum average scores (MAS) and the parameters determined in vitro (r = 0.752-0.764). These results indicate that the RBC assay is a useful and rapid test for use as a screening method to assess the IP of cosmetic products, and for predicting the IP value with a high level of concordance (94.7%). The assay showed high sensitivity and specificity rates of 91.6% and 100%, respectively.     

Fast projection matching for cryo-electron microscopy image reconstruction

A new FFT-accelerated projection matching method is presented and tested. The electron microscopy images are represented by their Fourier-Bessel transforms and the 3D model by its expansion in spherical harmonics, or more specifically in terms of symmetry-adapted functions. The rotational and translational properties of these representations are used to quickly access all the possible 2D projections of the 3D model, which allow an exhaustive inspection of the whole five-dimensional domain of parameters associated to each particle.     

A preponderantly 4-sulfated, 3-linked galactan from the green alga Codium isthmocladum

The green algae of the genus Codium have recently been demonstrated to be an important source of sulfated galactans from the marine environment. Here, a sulfated galactan was isolated from the species Codium isthmocladum and its structure was studied by a combination of chemical analyses and NMR spectroscopy. Two fractions (SG 1, approximately 14 kDa, and SG 2, approximately 20 kDa) were derived from this highly polydisperse and heterogeneous polysaccharide. Both exhibited similar structures in (1)H 1D NMR spectra. The structural features of SG 2 and its desulfated derivative were analyzed by COSY, TOCSY, DEPT-HSQC, HSQC, and HMBC. This sulfated galactan is composed preponderantly of 4-sulfated, 3-linked beta-D-galactopyranosyl units. In minor amounts, it is sulfated and glycosylated at C-6. Pyruvate groups are also found, forming five-membered cyclic ketals as 3,4-O-(1'carboxy)-ethylidene-beta-D-galactose residues. A comparison of sulfated galactans from different marine taxonomic groups revealed similar backbones of 3-beta-D-Galp-1.