Artificial reproduction in 1-year-old tench (Tinca tinca L.)

Although it is generally accepted that sexual maturity of tench (Tinca tinca L.) is not reached before 2–3 years of age, a successful experiment on artificial reproduction in 1‐year‐old tench is described. It was possible to obtain semen and eggs from 48% of males and 27.7% females kept in a small concrete tank at a final density of 3 kg m⁻³ and fed on a commercial dry diet. Two hormonal treatments were applied: the first with 1354 degree‐days and the second with 2353 degree‐days. In each experiment, the positive response in males and females was around 80%. Total egg production exceeded 11% of body weight, fertilization rate was above 90% and hatching rates around 50%. These results differ little from those reported for older breeders.     

A novel yeast mutation, rad52-L89F, causes a specific defect in Rad51-independent recombination that correlates with a reduced ability of Rad52-L89F to interact with Rad59

We isolated a novel rad52 mutation, rad52-L89F, which specifically impairs recombination in rad51Delta cells. rad52-L89F displays phenotypes similar to rad59Delta and encodes a mutant protein impaired in its ability to interact with Rad59. These results support the idea that Rad59 acts in homologous recombination via physical interaction with Rad52.     

A molecular approach to the characterization of the eukaryotic communities of an extreme acidic environment: methods for DNA extraction and denaturing gradient gel electrophoresis analysis

The diversity of the phytobenthonic community present in six acidophilic microbial mats from Río Tinto (Iberian Pyritic Belt, SW Spain) was analysed by optical microscopy and two molecular techniques, denaturing gradient gel electrophoresis (DGGE) and sequence analysis of 18S rDNA cloned gene fragments. Sixteen DNA isolation protocols as well as two commercial DNA extraction kits were tested and their efficiency compared. Purified DNA extracts were amplified by PCR using universal eukaryotic primers and the PCR products analysed by DGGE. Bead-mill homogenization was found to be superior to the other cell lysis methodologies assayed (sonication or freeze-thawing cycles) as it allowed efficiencies of cell disruption of over 95%. The methods combining bead-mill homogenization in the presence of SDS, treatment with chemical extractants (hexadecylmethylammonium bromide or guanidine isothiocyanate) and phenol extraction resulted in DNA preparations that amplified the same number of bands when analysed by DGGE as the two commercial kits assayed. The phylogenetic affiliations of the DGGE bands were determined by a BLAST search, and nine different species related to the Chlorophyta, Ciliophora, Kinetoplastida, Ascomycota, Streptophyta and Colcochaetales taxonomical groups were identified. Similar levels of diversity were found using cloning procedures. Although not all the species observed under the microscope were detected using molecular techniques, e.g. euglenas, heliozoan, or amoebae, DGGE fingerprints showed rather well the level of diversity present in the samples analysed, with limitations similar to cloning techniques.     

Activities of the protein kinases STK, PI3K, MEK, and ERK are required for the development of the head organizer in Hydra magnipapillata

The development of the hydra's head and its hypostome has been studied at the molecular level. Many genes have been cloned from hydra as potential candidates that control the development of its head. Much work was performed on the mechanisms controlling expression of these genes in the position-dependent manner. Moreover, there have been data to support the involvement of three main signaling pathways that involve PKC, SRC, and PI3K kinases in the regulation of the head formation and in the expression of several head-specific genes. In this report, we present data supporting the participation of these three signaling pathways on the development of the hypostome. We used grafting experiments and inhibitors of the specific kinases to show the participation of these enzymes in hypostome formation. From our results, we postulate that these signal transduction pathways regulate the very early stages of the head development, most likely at the point when the cells start to differentiate to form the head organizer.     

Production of a Metal-Binding Exopolysaccharide by Paenibacillus jamilae Using Two-Phase Olive-Mill Waste as Fermentation Substrate

The present study investigated the use of two-phase olive mill waste (TPOMW) as substrate for the production of exopolysaccharide (EPS) by the endospore-forming bacilli Paenibacillus jamilae. This microorganism was able to grow and produce EPS in aqueous extracts of TPOMW as a unique source of carbon. The effects of substrate concentration and the addition of inorganic nutrients were investigated. Maximal polymer yield in 100-ml batch-culture experiments (2 g l⁻¹) was obtained in cultures prepared with an aqueous extract of 20% TPOMW (w/v). An inhibitory effect was observed on growth and EPS production when TPOMW concentration was increased. Nutrient supplementation (nitrate, phosphate, and other inorganic nutrients) did not increase yield. Finally, an adsorption experiment of Pb (II), Cd (II), Cu (II), Zn (II), Co (II), and Ni (II) by EPS is reported. Lead was preferentially complexed by the polymer, with a maximal uptake of 230 mg/g EPS.     

Genetic structure of Triatoma venosa (Hemiptera: Reduviidae): molecular and morphometric evidence

Triatoma venosa presents a restricted geographical distribution in America and is considered as a secondary vector of Chagas disease in Colombia and Ecuador. A total of 120 adult insects were collected in domestic and peridomestic habitats in an endemic area of the department of Boyacá, Colombia, in order to determine their genetic structure through morphometric and molecular techniques. The head and wings of each specimen were used for the analyses of size, shape, and sexual dimorphism. A significant sexual dimorphism was found, although no differences in size among the studied groups were detected. Differences were found in the analyzed structures except for male heads. DNA was extracted from the legs in order to carry out the internal transcriber space-2 (ITS-2) amplification and the randon amplified polymorphic DNA (RAPD) analyses. Length polymorphisms were not detected in the ITS-2. Fst and Nm values were estimated (0.047 and 3.4, respectively). The high genetic flow found among the insects captured in the domicile and peridomiciliary environment does not permit a genetic differentiation, thus establishing the peridomicile as an important place for epidemiological surveillance.     

Eukaryotic community distribution and its relationship to water physicochemical parameters in an extreme acidic environment, Rio Tinto (southwestern Spain)

The correlation between water physicochemical parameters and eukaryotic benthic composition was examined in Río Tinto. Principal component analysis showed a high inverse relationship between pH and most of the heavy metals analyzed as well as Dunaliella sp., while Chlamydomonas sp. abundance was positively related. Zn, Cu, and Ni clustered together and showed a strong inverse correlation with the diversity coefficient and most of the species analyzed. These eukaryotic communities seem to be more influenced by the presence of heavy metals than by the pH.     

Synaptic proteins associate with a sub-set of lipid rafts when isolated from nerve endings at physiological temperature

Although the high presence of cholesterol in nerve terminals is well documented, specific roles of this lipid in transmitter release have remained elusive. Since cholesterol is a highly enriched component in the membrane microdomains known as lipid rafts, it is probable that these domains are very important in synaptic function. The extraction of lipid rafts using Brij 98 at 37 degrees C avoids the formation of nonspecific membrane aggregates at low temperature, allowing the isolation of more physiologically relevant lipid rafts. In the present work, we examine, by means of buoyancy analysis in sucrose gradients after solubilization of the membranes with Brij 98 or with Lubrol WX, the presence of proteins involved in exocytosis in detergent-resistant membranes (DRM) using rat brain synaptosomes as a neurological model. Significant proportions of the proteins tested in the present work, which are involved in neurotransmitter release, are found in Brij 98 raft fractions, demonstrating that significant pools of synaptic proteins are segregated in specific parts of the membrane at physiological temperature. On the other hand, Lubrol WX is unable to solubilize the major fraction of the proteins tested. Treatment of synaptosomes with methyl-beta-cyclodextrin (mbetaCD) causes alteration in the buoyancy properties of proteins initially present in Brij- as well as in Lubrol-resistant membranes, indicating the cholesterol-dependency of both kinds of microdomains. Finally, we detect the depolarization-induced enhancement of the cholesterol-dependent association of syntaxin 1 with Brij 98-rafts, under the same conditions in which prolonged neurotransmitter release is stimulated.