First report of a green toad (Bufo viridis sensu lato) in the Early Pleistocene of Spain: Palaeobiogeographical and palaeoecological implications

For the first time unequivocal fossil remains of a green toad (Bufo viridis s.l.) are described in the Iberian Peninsula. The fossils come from the Cueva Victoria site, a late Early Pleistocene (ca. 1.1–1.2 Ma) karstic filling in semi-arid southeastern Spain (Murcia region). By extension, other remains from two other Early Pleistocene Spanish localities, Barranco León D (ca. 1.3 Ma) and Almenara-Casablanca 3 (ca. 1.1 Ma), are cautiously attributed to the group B. viridis. The B. viridis group was previously reported with some uncertainty to the west of its current distribution area in Western Europe (Spain and France) in the Pliocene (Bufo cf. viridis) and less probably in the Early Miocene (Bufo aff. viridis). Since no osteological differences have been established between the recently described extant species of B. viridis s.l. (e.g. Bufo balearicus, Bufo siculus, Bufo boulengeri, B. viridis sensu stricto and Bufo variabilis) no precise palaeobiogeographical relationships can be drawn for the Spanish fossils. However, the occurrence of a third species of bufonid toad during the Pleistocene in the South of the Iberian Peninsula raises some interesting ecological questions in relation to the local disappearance of the green toad, which can be hypothetically linked to the intensification of the Pleistocene glacial/interglacial climate dynamic or to probable competition with another toad, Bufo calamita.     

The role of humans in the diversification of a threatened island raptor

Background  

Anthropogenic habitat modifications have led to the extinction of many species and have favoured the expansion of others. Nonetheless, the possible role of humans as a diversifying force in vertebrate evolution has rarely been considered, especially for species with long generation times. We examine the influence that humans have had on the colonization and phenotypic and genetic differentiation of an insular population of a long-lived raptor species, the Egyptian vulture (Neophron percnopterus).     

Conscious Brain-to-Brain Communication in Humans Using Non-Invasive Technologies

Human sensory and motor systems provide the natural means for the exchange of information between individuals, and, hence, the basis for human civilization. The recent development of brain-computer interfaces (BCI) has provided an important element for the creation of brain-to-brain communication systems, and precise brain stimulation techniques are now available for the realization of non-invasive computer-brain interfaces (CBI). These technologies, BCI and CBI, can be combined to realize the vision of non-invasive, computer-mediated brain-to-brain (B2B) communication between subjects (hyperinteraction). Here we demonstrate the conscious transmission of information between human brains through the intact scalp and without intervention of motor or peripheral sensory systems. Pseudo-random binary streams encoding words were transmitted between the minds of emitter and receiver subjects separated by great distances, representing the realization of the first human brain-to-brain interface. In a series of experiments, we established internet-mediated B2B communication by combining a BCI based on voluntary motor imagery-controlled electroencephalographic (EEG) changes with a CBI inducing the conscious perception of phosphenes (light flashes) through neuronavigated, robotized transcranial magnetic stimulation (TMS), with special care taken to block sensory (tactile, visual or auditory) cues. Our results provide a critical proof-of-principle demonstration for the development of conscious B2B communication technologies. More fully developed, related implementations will open new research venues in cognitive, social and clinical neuroscience and the scientific study of consciousness. We envision that hyperinteraction technologies will eventually have a profound impact on the social structure of our civilization and raise important ethical issues.     

Phylogeny, speciation and species turnover. The case of the Mediterranean gastropods of genus Cyclope Risso, 1826

The genus Cyclope Risso, 1826 (family Nassariidae) has appeared in the fossil record since the Pliocene. Although it is still found today, the teleoconch morphology has never undergone modification, despite the fact that the protoconch morphologies of fossils (multispiral) and living forms (paucispiral) are different. They vary in their embryological and larval development and, hence, are two different species: C. migliorinii (Bevilacqua, 1928), the fossil species, and C. neritea (Linnaeus, 1758), the living species. We discuss the morphologic modifications in the evolution of this genus: the speciation that leads to its appearance and the speciation driving the Pliocene species to the living one. The order and the direction of these changes are based on phylogenetic analysis. No intermediate forms have been found showing a gradual morphological change that could have been worked by natural selection. Our analysis takes as the origin of the morphological novelties the genetic modifications in the ontogenetic processes which resulted in rapid and important phenotypic changes. Both speciation processes are sympatric cladogenetic. The changes that determine the appearance of the genus affect only the teleoconch, not the larval development. The modifications that lead from one species to the other, within the genus Cycope, affect the larval development exclusively. This points to a certain disconnection between the development of the embryo-larval phase and the young-adult formation, such that evolutionary processes could have occurred independently in different ontogenetic stages. The influence of larval ecology in relation to extinction of the ancestor and persistence of the derived species is also analysed. We hypothesize that climatic fluctuations may have affected the planktonic larvae of the fossil species, driving it to extinction. The living species, developing without the planktonic phase, would have resisted these climatic changes. We consider that the mechanisms described as drivers of the evolution of this genus can be of more general validity in prosobranch gastropods.     

Immobilized cells: behaviour of carrageenan entrapped yeast during continuous ethanol fermentation

Summary Data of cell concentration, viability and microscopic observation of cell distribution inside carrageenan immobilized yeast beads are reported. Results were obtained from a continuous packed-bed reactor performing alcoholic fermentation and the main observations made on cell activity are in agreement with the fermentation profiles inside the fermenter.     

C-terminal labelling of beta-casein

This paper is the first to report specific labelling of a native protein at its C-terminal end by carboxypeptidase Y-catalyzed transpeptidation between beta-casein and tritiated Phe amide. A tryptic digest of the radiolabelled protein was resolved by reversed-phase HPLC and a single labelled peptide was isolated therefrom. Sequence determination and FAB mass spectrometry showed that the last 2 residues (Val-209, Ile-208) of beta-casein had been deleted and Ile 207 substituted by Phe, deamidation presumably occurring after transpeptidation. Identical results were obtained by transpeptidating the isolated C-terminal tryptic heptapeptide (203-209) of native beta-casein.     

Study of the causes of direct-acting mutagenicity in coffee and tea using the Ara test in Salmonella typhimurium

The mutagenic activities of 6 of the chemicals identified in coffee solutions were assayed with the Salmonella Ara test, under experimental conditions optimized for coffee mutagenicity. Caffeine was the only non-mutagenic compound. Among the other 5 chemicals, hydrogen peroxide was the strongest mutagen and chlorogenic acid the weakest; methylglyoxal, glyoxal and caffeic acid exhibited intermediate mutagenicities. The minimal mutagenic doses of these components correlated negatively with their relative concentrations in coffee. It was concluded that chlorogenic acid, caffeic acid, glyoxal and methylglyoxal cannot contribute alone to the mutagenicity of coffee in the Ara test, since their minimal mutagenic concentrations were much higher than their respective levels in the coffee samples assayed. By contrast, 40-60% of the mutagenic activity in coffee and also in tea could be attributed to their H2O2 contents. Catalase abolished more than 95% of the mutagenic activity of coffee, as detected by the Ara test. A similar sensitivity to catalase has been reported by other authors in relation to the coffee mutagenicity identified by the Salmonella His test. Nevertheless, the results presented in this paper suggest that the Ara forward and the His reverse mutation tests are sensitive to the mutagenicity of different constituents in coffee solutions. We propose that the His test, sensitive at high coffee doses, mainly recognizes the mutagenicity of methylglyoxal, whilst the Ara test, sensitive at low coffee doses, mainly detects the mutagenic activity of hydrogen peroxide. The data reported also suggest that the direct-acting mutagenicity(ies) detected by the Ara test in tea solutions is (are) based on similar, if not identical, mechanisms.     

Functional groups and quaternary structure of chicken liver xanthine dehydrogenase

Xanthine dehydrogenase from chicken liver is a dimeric enzyme, each hemimolecule containing one FAD and two Fe/S groups. Determination of sulfhydryl groups with 5,5-dithiobis(2-nitrobenzoic acid) (DTNB) andp-hydroxymercuribenzoic acid (PMB) showed a variable number of sulfhydryl groups depending onpH, ionic strength, and nature of the reaction medium and buffer. The number of disulfide bonds was determined with DTNB and reducing conditions. Amino groups were determined with 2,4,6,-trinitrobencensulfonic acid (TNBS). At constant temperature andpH the reaction of DTNB and TNBS with native xanthine dehydrogenase showed an exponential dependence on time. From the obtained parameters the number of available sulfhydryl and amino groups at infinite concentration of enzyme and the rate constant of the equation were determined. The absorption spectrum of the enzyme changed with time when a chaotropic agent (1 M sodium nitrate) was added to the medium. This difference was detected by measuring the absorbance in the range 450–550 nm. The absorption spectrum (between 350 and 600 nm) also changed when a denaturating agent (sodium dodecyl sulfate) was added. This modification increased with time and depended on the medium.     

Kinetics of action of chymosin (rennin) on some peptide bonds of bovine beta-casein

The first steps of proteolysis of bovine beta-casein by chymosin were studied quantitatively by using reverse-phase high-performance liquid chromatography (RP-HPLC). Although chymosin has a broad specificity, it has been possible to selectively study the hydrolysis of two bonds (Ala-189-Phe-190 and Leu-192-Tyr-193) by choosing appropriate conditions. The disappearance of the substrate and the appearance of the reaction products as a function of time were followed at 220 nm by RP-HPLC. For concentrations where beta-casein was in a micellar form, the Michaelian parameters corresponding to the cleavage of bond 192-193 were determined by measuring initial rates of reaction at different substrate concentrations in a time period for which splitting of bond 189-190 was negligible. The following results were obtained; k1cat = 1.54 s-1, K1m = 0.075 mM, and k1cat/K1m = 20.6 mM-1 s-1. Under conditions where the protein was in a monomeric state, the following parameters were determined for the splitting of bond 192-193 by integrating the Michaelis equation: k2cat = 0.056 s-1, K2m = 0.007 mM, and k2cat/K2m = 79.7 mM-1 s-1. Under the latter conditions the four enzymic reactions involved in the cleavage of bonds 189-190 and 192-193 were first-order reactions. The four corresponding apparent rate constants were calculated by using a computer program. Excellent agreement was obtained between concentrations of four molecular species measured during the reaction period and those calculated by using the four apparent rate constants.