Phylogeography of isolated freshwater three-spined stickleback Gasterosteus aculeatus populations in the Adriatic Sea basin.

Analyses of mitochondrial (mt) DNA and microsatellite variation were carried out to examine the relationships between 10 freshwater populations of three-spined sticklebacks Gasterosteus aculeatus along the eastern coast of the Adriatic Sea. Partial sequences of the mtDNA control region and cytochrome b gene, in addition to 15 microsatellite loci, were used to analyse populations from four isolated river catchments. Results uncovered an Adriatic lineage that was clearly divergent from the European lineage, and confirmed that the most divergent and ancient populations are located within the Adriatic lineage as compared with other European populations. Two northern Adriatic populations formed independent clades within the European mitochondrial lineage, suggesting different colonization histories of the different Adriatic populations. Nuclear marker analyses also indicated deep divergence between Adriatic and European populations, albeit with some discordance between the mtDNA phylogeny of the northern Adriatic populations, further highlighting the strong differentiation among the Adriatic populations. The southern populations within the Adriatic lineage were further organized into distinct clades corresponding to respective river catchments and sub-clades corresponding to river tributaries, reflecting a high degree of population structuring within a small geographic region, concurrent with suggestions of existence of several microrefugia within the Balkan Peninsula. The highly divergent clades and haplotypes unique to the southern Adriatic populations further suggest, in accordance with an earlier, more limited survey, that southern Adriatic populations represent an important reservoir for ancient genetic diversity of G. aculeatus.

A new endemic cyprinid species from the Danube drainage.

Populations of endemic Croatian dace were found to belong to two different species, one of which is first described in this study. Telestes karsticus sp. nov. differed morphologically from Telestes polylepis in the total count of lateral line scales, number of gill rakers and the shape of the posterior margin of the anal fin. Morphological differences were corroborated with mtDNA analyses (with p-distance between T. polylepis and T. karsticus sp. nov. ranging between 3·2 and 4·1%; and the number of substitutions between 37 and 47). The newly described species is geographically very localized. It has been recorded from only four localities around Velika Kapela and Mala Kapela mountains in Croatia.

Albumin-based nanoparticles as magnetic resonance contrast agents: II. Physicochemical characterisation of purified and standardised nanoparticles.

We are developing a nanoparticulate histochemical reagent designed for histochemistry in living animals (molecular imaging), which should finally be useful in clinical imaging applications. The iterative development procedure employed involves conceptual design of the reagent, synthesis and testing of the reagent, then redesign based on data from the testing; each cycle of testing and development generates a new generation of nanoparticles, and this report describes the synthesis and testing of the third generation. The nanoparticles are based on human serum albumin and the imaging modality selected is magnetic resonance imaging (MRI). Testing the second particle generation with newly introduced techniques revealed the presence of impurities in the final product, therefore we replaced dialysis with diafiltration. We introduced further testing methods including thin layer chromatography, arsenazo III as chromogenic assay for gadolinium, and several versions of polyacrylamide gel electrophoresis, for physicochemical characterisation of the nanoparticles and intermediate synthesis compounds. The high grade of chemical purity achieved by combined application of these methodologies allowed standardised particle sizes to be achieved (low dispersities), and accurate measurement of critical physicochemical parameters influencing particle size and imaging properties. Regression plots confirmed the high purity and standardisation. The good degree of quantitative physicochemical characterisation aided our understanding of the nanoparticles and allowed a conceptual model of them to be prepared. Toxicological screening demonstrated the extremely low toxicity of the particles. The high magnetic resonance relaxivities and enhanced mechanical stability of the particles make them an excellent platform for the further development of MRI molecular imaging.

Occurrence of four-spined Gasterosteus aculeatus in an isolated Croatian river population.

An isolated population of the three-spined stickleback Gasterosteus aculeatus in Croatia was found to have a high incidence of specimens either having a fourth dorsal spine or showing remnants of a fourth spine. Juvenile individuals showed a 9.4% incidence of a fourth spine. The population was examined for asymmetry of the skeletal defensive complex in order to determine whether the additional spine could be the result of developmental instability, a response to predation or environmental conditions.