Four new species of Cayaponia (Cucurbitaceae) from Brazil and Bolivia

Four new species ofCayaponia are described and illustrated: three from Brazil (C. cogniauxiana, C. nitida andC. rugosa) and one from Brazil and Bolivia (C. ferruginea).     

Validation of DNA methylation profiling in formalin-fixed paraffin-embedded samples using the Infinium HumanMethylation450 Microarray

A formalin-fixed paraffin-embedded (FFPE) sample usually yields highly degraded DNA, which limits the use of techniques requiring high-quality DNA, such as Infinium Methylation microarrays. To overcome this restriction, we have applied an FFPE restoration procedure consisting of DNA repair and ligation processes in a set of paired fresh-frozen (FF) and FFPE samples. We validated the FFPE results in comparison with matched FF samples, enabling us to use FFPE samples on the Infinium HumanMethylation450 Methylation array.     

Micropropagation of Rollinia mucosa (JACQ.) baill

Summary A protocol for in vitro propagation of Rollinia mucosa, an important medicinal plant, was developed. The presence of 500 mg l⁻¹ polyvinylpyrrolidone (PVP) during explant excision was important to avoid browning. Axillary buds, adventitious buds, and shoot cluster proliferation were achieved from epicotyl and hypocotyl explants from nursery-grown seedlings. The highest direct organogenesis percentage from hypocotyl explants was obtained upon culture of explants on Murashige and Skoog medium supplemented with 2.2 μM benzyladenine (BA) plus 2.32 μM kinetin. Epicotyl explants display highest regeneration frequency on a medium containing 8.8 μM BA and 0.54 μM naphthaleneacetic acid. Gibberellic acid was necessary for shoot elongation. Root induction was observed when shoots were pretreated with activated charcoal for 7 d in the dark before culture on Woody Plant Medium supplemented with 49.21 μM indolebutyric acid for 10 d. Root development was observed when 20 g l⁻¹ sucrose was used. Rooted plantlets were acclimatized and grown in the greenhouse.     

Biogeography of Iberian freshwater fishes revisited: the roles of historical versus contemporary constraints

Aim The question of how much of the shared geographical distribution of biota is due to environmental vs. historical constraints remains unanswered. The aim of this paper is to disentangle the contribution of historical vs. contemporary factors to the distribution of freshwater fish species. In addition, it illustrates how quantifying the contribution of each type of factor improves the classification of biogeographical provinces. Location Iberian Peninsula, south‐western Europe (c. 581,000 km²). Methods We used the most comprehensive data on native fish distributions for the Iberian Peninsula, compiled from Portuguese and Spanish sources on a 20‐km grid‐cell resolution. Overall, 58 species were analysed after being categorized into three groups according to their ability to disperse through saltwater: (1) species strictly intolerant of saltwater (primary species); (2) species partially tolerant of saltwater, making limited incursions into saltwaters (secondary species); and (3) saltwater‐tolerant species that migrate back and forth from sea to freshwaters or have invaded freshwaters recently (peripheral species). Distance‐based multivariate analyses were used to test the role of historical (basin formation) vs. contemporary environmental (climate) conditions in explaining current patterns of native fish assemblage composition. Cluster analyses were performed to explore species co‐occurrence patterns and redefine biogeographical provinces based on the distributions of fishes. Results River basin boundaries were better at segregating species composition for all species groups than contemporary climate variables. This historical signal was especially evident for primary and secondary freshwater fishes. Eleven biogeographical provinces were delineated. Basins flowing to the Atlantic Ocean north of the Tagus Basin and those flowing to the Mediterranean Sea north of the Mijares Basin were the most dissimilar group. Primary and secondary freshwater species had higher province fidelity than peripheral species. Main conclusions The results support the hypothesis that historical factors exert greater constraints on native freshwater fish assemblages in the Iberian Peninsula than do current environmental factors. After examining patterns of assemblage variation across space, as evidenced by the biogeographical provinces, we discuss the likely dispersal and speciation events that underlie these patterns.     

Determination of trace levels of fatty acid metal salts by high-performance liquid chromatography with fluorescence prelabeling

A simple method is described for picomole determinations of fatty acid metal salts. Fatty acid salts are directly labeled with 4-bromomethyl-7-methoxycoumarin in the presence of excess ethylenediaminetetraacetic acid tripotassium salt without any solvent extractions. The fluorescence derivatives of fatty acids are separated by reverse-phase high-performance liquid chromatography followed by fluorometric detection. The response of each fatty acid (C8-C18) calcium salt is linear from 1 to 50 micrograms/ml of samples. The detection limit is about 7 pmol. Good recoveries are obtained for the calcium salts of myrystic acid and soap (C8-C18, C18:1,2). The new method is successfully applied to the study on biodegradation of fatty acids in river water.     

Phylogeography of the cactophilic species Drosophila gouveai: demographic events and divergence timing in dry vegetation enclaves in eastern Brazil

Aim  The aim of this study was to assess the causal mechanisms underlying populational subdivision in Drosophila gouveai , a cactophilic species associated with xeric vegetation enclaves in eastern Brazil. A secondary aim was to investigate the genetic effects of Pleistocene climatic fluctuations on these environments.
Location  Dry vegetation enclaves within the limits of the Cerrado domain in eastern Brazil.
Methods  We determined the mitochondrial DNA haplotypes of 55 individuals (representing 12 populations) based on sequence data of a 483-bp fragment from the cytochrome c oxidase subunit II (COII) gene. Phylogenetic and coalescent analyses were used to test for the occurrence of demographic events and to infer the time of divergence amongst genetically independent groups.
Results  Our analyses revealed the existence of two divergent subclades (G1 and G2) plus an introgressed clade restricted to the southernmost range of D. gouveai . Subclades G1 and G2 displayed genetic footprints of range expansion and segregated geographical distributions in south-eastern and some central highland regions, east and west of the Paraná River valley. Molecular dating indicated that the main demographic and diversification events occurred in the late to middle Pleistocene.
Main conclusions  The phylogeographical and genetic patterns observed for D. gouveai in this study are consistent with changes in the distribution of dry vegetation in eastern Brazil. All of the estimates obtained by molecular dating indicate that range expansion and isolation pre-dated the Last Glacial Maximum, occurring during the late to middle Pleistocene, and were probably triggered by climatic changes during the Pleistocene. The current patchy geographical distribution and population subdivision in D. gouveai is apparently closely linked to these past events.     

Protein binding and stability of norepinephrine in human blood plasma. Involvement of prealbumin, alpha 1-acid glycoprotein and albumin

The binding of norepinephrine (NE) to plasma proteins of fresh human blood obtained from healthy volunteers was studied by ultrafiltration at different NE concentrations and incubation times at 37 degrees C. At 1.7 nM L-[3H]-NE binding was approximately 25%. The binding was rapid and was not influenced by the incubation time. [3H]-NE could be dissociated from its binding sites by acid precipitation and, after HPLC, showed to be unchanged NE. No difference in NE binding was found between plasma collected in EGTA-GSH or heparin solution. There was no degradation of NE when incubated in plasma at 37 degrees C for 10 h, even without the addition of antioxidants. Therefore, in the present study, binding represented interaction of unchanged NE with plasma proteins. The whole plasma binding was saturable over the range of 0.66 nM to 0.59 mM of NE. Scatchard plot of specific binding revealed high-affinity sites with a Kd of 5.4 nM and a Bmax of 3.9 fmoles.mg-1 protein, and low-affinity sites with a Kd of 2.7 microM and a Bmax of 3.3 pmoles.mg-1 protein. Electrophoretic characterization of NE-binding proteins showed that about 60% of bound NE was associated to albumin, and 20% to prealbumin. NE binding to pure human plasma proteins was also studied using ultrafiltration. Scatchard analyses revealed a single class of very high-affinity binding sites for prealbumin (Kd 4.9 nM), a single class of binding sites for alpha 1-acid glycoprotein (Kd 54 microM) and two classes of binding sites for albumin with high (Kd 1.7 microM) and low (Kd 0.8 mM) affinities respectively. The main results obtained in this study - a) reversibility of NE binding, b) stability of free and bound NE in plasma, c) involvement of the prealbumin as a specific binding protein - point out to a specific transport for NE in human blood plasma.