Macronutrients requirements of the dinoflagellate Protoceratium reticulatum

The basal L1 medium was found to be unsatisfactory for culturing the red tide dinoflagellate Protoceratium reticulatum at a high growth rate and biomass yield. The L1 medium enhanced with phosphate to a total concentration of 217 μM supported the highest attainable growth rate and biomass yield. Once the phosphate concentration exceeded 6× L1, phosphate inhibited the dinoflagellate growth and negatively affected cell viability. At the optimal phosphate concentration of 217 μM, an increase in nitrate concentration over the range of 882–8824 μM, did not affect cell growth and yield. Nitrate did not inhibit growth at any of the concentrations used. Clearly, the basal nitrate level in L1 is sufficient for effectively culturing P. reticulatum. At the ranges of phosphate and nitrate concentrations tested, cell volume was not sensitive to the concentration of nutrients but the concentration of phosphate affected both the specific cell number and cell volume growth rates. Elevated levels of nutrients supported their intracellular accumulation. Cell-specific production of yessotoxin was not influenced by concentration of phosphate in the culture medium, but elevated (>1764 μM) nitrate concentration did enhance the yessotoxin level. Phosphate concentration that maximized biomass yield also maximized volumetric production of yessotoxin in the culture broth.     

Robust and highly informative microsatellite-based genetic identity kit for potato

The fingerprinting of 742 potato landraces with 51 simple sequence repeat (SSR, or microsatellite) markers resulted in improving a previously constructed potato genetic identity kit. All SSR marker loci were assayed with a collection of highly diverse landraces of all species of cultivated potato with ploidies ranging from diploid to pentaploid. Loci number, amplification reproducibility, and polymorphic information content were recorded. Out of 148 SSR markers of which 30 are new, we identified 58 new SSR marker locations on at least one of three potato genetic linkage maps. These results permitted the selection of a new potato genetic identity kit based on 24 SSR markers with two per chromosome separated by at least 10 cM, single locus, high polymorphic information content, and high quality of amplicons as determined by clarity and reproducibility. The comparison of a similarity matrix of 742 landraces obtained with the 24 SSR markers of the new kit and with the entire dataset of 51 SSR markers showed a high correlation (r = 0.94) by a Mantel test and even higher correlations (r = 0.99) regarding topological comparisons of major branches of a neighbor joining tree. This new potato genetic identity kit is able to discriminate 93.5% of the 742 landraces compared to 98.8% with 51 SSR markers. In addition, we made a marker-specific set of allele size standards that conveniently and unambiguously provide accurate sizing of all alleles of the 24 SSR markers across laboratories and platforms. The new potato genetic identity kit will be of particular utility to standardize the choice and allele sizing of microsatellites in potato and aid in collaborative projects by allowing cumulative analysis of independently generated data. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Dispersal and establishment both limit colonization during primary succession on a glacier foreland

Plant colonization can be limited by lack of seeds or by factors that reduce establishment. The role of seed limitation in community assembly is being increasingly recognized, but in early primary succession, establishment failure is still considered more important. We studied the factors limiting colonization on the foreland of Coleman Glacier, Washington, USA, to determine the importance of seed and establishment limitation during primary succession. We also evaluated the effects of seed predation, drought, and existing vegetation on establishment. We planted seeds of seven species into plots of four different ages and found evidence that both seed and establishment limitation are strong in early succession. We also found that seed and establishment limitation both remained high in later stages of succession. Seed predation reduced establishment for most species and some evidence suggested that drought and existing vegetation also limit establishment. Because both dispersal and establishment failure restrict colonization in recently exposed habitat, late-seral forest species may have a difficult time migrating upward in response to global climate change. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Early phases of a successful invasion: mitochondrial phylogeography of the common genet (Genetta genetta) within the Mediterranean Basin

The Mediterranean Basin, connected by cultural exchanges since prehistoric times, provides an outstanding framework to study species translocations. We address here the early phases of the successful invasion of the common genet (Genetta genetta), a small carnivoran supposedly introduced from Africa to Europe during historical times, by assessing mitochondrial nucleotide variability in 134 individuals from its native and invasive ranges. We identify four lineages within the native species range [northern Algeria, Peninsular Arabia, southern Africa and western Africa + Maghreb (including northern Algeria)], in contradiction with morphological taxonomy. We propose that the co-occurrence in Maghreb of two divergent lineages (autochthonous and western African) is due to secondary contact through intermittent permeability of the Saharan belt during the Plio-Pleistocene. Estimates of coalescence time and genetic diversity, in concert with other available evidences in the literature, indicate that the origin of European populations of common genets is in Maghreb, possibly restricted to northern Algeria. The autochthonous mitochondrial lineage of Maghreb was the only contributor to the European pool, suggesting that translocations were associated to a cultural constraint such as a local use of the species, which might have artificially excluded the western African lineage. Haplotype network and nested clade analysis (NCA) provide evidence for independent events of introductions throughout Spain (Andalucia, Cataluña, and the Balearic Isl.)—and, to a lesser extent, Portugal—acting as a ‘translocation hotspot’. Due to the reduced number of northern Algerian individuals belonging to the autochthonous mitochondrial lineage of Maghreb, it remains impossible to test hypotheses of historical translocations, although a main contribution of the Moors is likely. Our demographic analyses support a scenario of very recent introduction of a reduced number of individuals in Europe followed by rapid population expansion. We suggest that an exceptional combination of factors including multiple translocations, human-driven propagation across natural barriers, and natural processes of colonization allowed by a wide ecological tolerance, promoted the successful spread of the common genet into Europe.     

Clinical Comparison of Two Mexican Coccidioidins

Coccidioidin, an extract from the saprophytic mycelial form of Coccidioides spp., has been a very useful antigen preparation both for skin and serological tests for coccidioidomycosis. Unfortunately, coccidioidin is not currently available for skin testing in the United States. Coccidioidin has been produced commercially in Mexico by a vaccine and reagents laboratory of the Mexican Federal Government. It also has been produced at the Microbiology Laboratory of the Faculty of Medicine, Universidad Nacional Autónoma de México exclusively as an antigen for research projects. The objective of the study was to compare both coccidioidins in their reactivity and safety when applied in humans. One hundred and eighty-four volunteers were tested; median age was 33 (range 14–82). When the cutoff point is set in 5 mm, 88 subjects (47.8%) had a positive test for the commercial coccidioidin and 76 (41.3%; CI_95% 0.50, 1.15; P = 0.20) were positive with the research antigen. Seventy-five subjects were positive for both antigens and 96 were negative for both. Fifty-nine subjects (31.3%) reported an adverse reaction after the application of the antigen; they were mostly very mild local reactions. Mexican research coccidioidin is a safe and reliable antigen that can be used for the detection of coccidioidomycosis infection in mammals.     

Differential expression of P-type ATPases in intestinal epithelial cells: Identification of putative new atp1a1 splice-variant

P-type ATPases are membrane proteins that couple ATP hydrolysis with cation transport across the membrane. Ten different subtypes have been described. In mammalia, 15 genes of P-type ATPases from subtypes II-A, II-B and II-C, that transport low-atomic-weight cations (Ca²⁺, Na⁺, K⁺ and H⁺), have been reported. They include reticulum and plasma-membrane Ca²⁺-ATPases, Na⁺/K⁺-ATPase and H⁺/K⁺-ATPases. Enterocytes and colonocytes show functional differences, which seem to be partially due to the differential expression of P-type ATPases. These enzymes have 9 structural motifs, being the phosphorylation (E) and the Mg²⁺ATP-binding (H) motifs the most preserved. These structural characteristics permitted developing a Multiplex-Nested-PCR (MN-PCR) for the simultaneous identification of different P-type ATPases. Thus, using MN-PCR, seven different cDNAs were cloned from enterocytes and colonocytes, including SERCA3, SERCA2, Na⁺/K⁺-ATPase α1-isoform, H⁺/K⁺-ATPase α2-isoform, PMCA1, PMCA4 and a cDNA-fragment that seems to be a new cassette-type splice-variant of the atp1a1 gen. PMCA4 in enterocytes and H⁺/K⁺-ATPase α2-isoform in colonocytes were differentially expressed. This cell-specific expression pattern is related with the distinctive enterocyte and colonocyte functions.     

A Single Mutation at the Sheet Switch Region Results in Conformational Changes Favoring λ6 Light-Chain Fibrillogenesis

Systemic amyloid light-chain (LC) amyloidosis is a disease process characterized by the pathological deposition of monoclonal LCs in tissue. All LC subtypes are capable of fibril formation although λ chains, particularly those belonging to the λ6 type, are overrepresented. Here, we report the thermodynamic and in vitro fibrillogenic properties of several mutants of the λ6 protein 6aJL2 in which Pro7 and/or His8 was substituted by Ser or Pro. The H8P and H8S mutants were almost as stable as the wild-type protein and were poorly fibrillogenic. In contrast, the P7S mutation decreased the thermodynamic stability of 6aJL2 and greatly enhanced its capacity to form amyloid-like fibrils in vitro. The crystal structure of the P7S mutant showed that the substitution induced both local and long-distance effects, such as the rearrangement of the V_L (variable region of the light chain)-V_L interface. This mutant crystallized in two orthorhombic polymorphs, P2₁2₁2₁ and C222₁. In the latter, a monomer that was not arranged in the typical Bence-Jones dimer was observed for the first time. Crystal-packing analysis of the C222₁ lattice showed the establishment of intermolecular β-β interactions that involved the N-terminus and β-strand B and that these could be relevant in the mechanism of LC fibril formation. Our results strongly suggest that Pro7 is a key residue in the conformation of the N-terminal sheet switch motif and, through long-distance interactions, is also critically involved in the contacts that stabilized the V_L interface in λ6 LCs.