MULTIPLE ROUTES OF SEXUALITY IN ALEXANDRIUM TAYLORI (DINOPHYCEAE) IN CULTURE1

Alexandrium taylori Balech is a cyst‐forming dinoflagellate species responsible for recurrent blooms in Mediterranean coastal waters. The nuclear development of the cells during the sexual cycle and the effect of different external nitrate and phosphate levels were studied. Nuclear fusion of gametes occurred 6–12 h after the complete cytoplasmic fusion. The U‐shaped nuclei fused through the end of one nucleus and the mid‐area of the other. The mobile and biflagellated zygote had a large, U‐shaped nucleus and may follow three different fates: direct division, short‐term encystment (ecdysal), and long‐term encystment (resting). Ecdysal cysts may divide in >24–96 h into two, four, six, or eight cells before germinating. Meiosis presumably occurred in three locations: in the planozygote, within the ecdysal cyst, and in the planomeiocyte (germling) liberated either from ecdysal or resting cysts. The effects of nutrients on these routes were studied in individually isolated sexual stages. (1) Direct divisions occurred mainly under replete conditions (L1), whereas no direct planozygote divisions were recorded in media with no phosphate added (L‐P). (2) Short‐term encystment was larger in media lacking phosphate (L‐P and L/30) than in medium with no nitrate added (L‐N) or under replete conditions (L1). (3) Long‐term encystment was only observed in medium with no nitrate added (L‐N). The long‐lived resting cyst, not previously described for this species, had a clear double wall, an irregular shape, a flat morphology, and a middle orange spot. No cysts germinated in 1–2 months, whereas 86% of the cysts germinated 2–3 months after being formed. A flow cytometry analysis showed that sexual induction and zygote formation were very fast and highly common processes, zygotes being nearly half of the population at days 3 and 5 after the induction of sexuality in the cultures.     

Palmitate acutely raises glycogen synthesis in rat soleus muscle by a mechanism that requires its metabolization (Randle cycle)

The acute effect of palmitate on glucose metabolism in rat skeletal muscle was examined. Soleus muscles from Wistar male rats were incubated in Krebs-Ringer bicarbonate buffer, for 1 h, in the absence or presence of 10 mU/ml insulin and 0, 50 or 100 microM palmitate. Palmitate increased the insulin-stimulated [(14)C]glycogen synthesis, decreased lactate production, and did not alter D-[U-(14)C]glucose decarboxylation and 2-deoxy-D-[2,6-(3)H]glucose uptake. This fatty acid decreased the conversion of pyruvate to lactate and [1-(14)C]pyruvate decarboxylation and increased (14)CO(2) produced from [2-(14)C]pyruvate. Palmitate reduced insulin-stimulated phosphorylation of insulin receptor substrate-1/2, Akt, and p44/42 mitogen-activated protein kinases. Bromopalmitate, a non-metabolizable analogue of palmitate, reduced [(14)C]glycogen synthesis. A strong correlation was found between [U-(14)C]palmitate decarboxylation and [(14)C]glycogen synthesis (r=0.99). Also, palmitate increased intracellular content of glucose 6-phosphate in the presence of insulin. These results led us to postulate that palmitate acutely potentiates insulin-stimulated glycogen synthesis by a mechanism that requires its metabolization (Randle cycle). The inhibitory effect of palmitate on insulin-stimulated protein phosphorylation might play an important role for the development of insulin resistance in conditions of chronic exposure to high levels of fatty acids.     

The green thorns of Ulex europaeus play both defensive and photosynthetic roles: consequences for predictions of the enemy release hypothesis

Biological Invasions - The widespread invasive success of Ulex europaeus, a thorny shrub native to NW Europe, remains to be understood from a functional perspective. According to the Enemy Release...     

Development of a primer system to study abundance and diversity of the gene coding for alanine aminopeptidase pepN gene in Gram-negative soil bacteria

A new set of primers was developed allowing the specific detection of the pepN gene (coding for alanine aminopeptidase) from Gram-negative bacteria. The primers were designed in silico by sequence alignments based on available DNA sequence data. The PCR assay was validated using DNA from selected pure cultures. The analysis of gene libraries from extracted DNA from different soil samples revealed a high diversity of pepN related sequences mainly related to α-Proteobacteria. Most sequences obtained from clone libraries were closely related to already published sequences (<80% homology on amino acid level), which may be related to the conserved character of the amplified region of pepN. By linking the diversity data obtained by the clone library studies to potential enzymatic activities of alanine aminopeptidase, lowest diversity of pepN was found in those soil samples which displayed lowest activity levels, which confirms the importance of diversity for the ecosystem function mainly when transformation processes of complex molecules are studied.     

Population Genetics of Streptococcus dysgalactiae Subspecies equisimilis Reveals Widely Dispersed Clones and Extensive Recombination

Background

Streptococcus dysgalactiae subspecies equisimilis (SDSE) is an emerging global pathogen that can colonize and infect humans. Although most SDSE isolates possess the Lancefield group G carbohydrate, a significant minority have the group C carbohydrate. Isolates are further sub-typed on the basis of differences within the emm gene. To gain a better understanding of their molecular epidemiology and evolutionary relationships, multilocus sequence typing (MLST) analysis was performed on SDSE isolates collected from Australia, Europe and North America.

Methodology/Principal Findings

The 178 SDSE isolates, representing 37 emm types, segregate into 80 distinct sequence types (STs) that form 17 clonal complexes (CCs). Eight STs recovered from all three continents account for >50% of the isolates. Thus, a small number of STs are highly prevalent and have a wide geographic distribution. Both ST and CC strongly correlate with group carbohydrate. In contrast, eleven STs were associated with >1 emm type, suggestive of recombinational replacements involving the emm gene; furthermore, 35% of the emm types are associated with genetically distant STs. Data also reveal a history of extensive inter- and intra-species recombination involving the housekeeping genes used for MLST. Sequence analysis of single locus variants identified through goeBURST indicates that genetic change mediated by recombination occurred ∼4.4 times more frequently than by point mutation.

Conclusions/Significance

A few genetic lineages with an intercontinental distribution dominate among SDSE causing infections in humans. The distinction between group C and G isolates reflects recent evolution, and no long-term genetic isolation between them was found. Lateral gene transfer and recombination involving housekeeping genes and the emm gene are important mechanisms driving genetic variability in the SDSE population.     

Human host genetic factors in mycobacterial and Salmonella infection: lessons from single gene disorders in IL-12/IL-23-dependent signaling that affect innate and adaptive immunity

Interleukin (IL)-12/IL-23 signal transduction-deficient individuals with genetic defects in IL12RB1 or IL12B often suffer from unusual mycobacterial and Salmonella infections. Here we discuss recent questions that have arisen from clinical observations that cast doubt on the necessity of IL-12/IL-23 signaling in controlling infections with intracellular bacteria. Alternative IL-12/IL-23-dependent, interferon-gamma-independent pathways of immunity to intracellular bacteria are also discussed.     

Recognition of pathogenic Yersinia enterocolitica by crystal violet binding and polymerase chain reaction

Cefsulodin-Irgasan-Novobiocin (CIN) agar is used for the selective isolation and enumeration of Yersinia enterocolitica from clinical specimens and food. The medium contains crystal violet and about 1 mmol l^-1 calcium and can be used for the phenotypic characterization of strains that carry a virulence plasmid. At 32°C, irrespective of pathogenicity, colonies are translucent with a pale pink centre surrounded by a transparent border ('bullseye'), while at 37°C pathogenic strains grow as calcium-dependent microcolonies which, because of crystal violet binding, are intensely coloured. These results were confirmed by the polymerase chain reaction with primers directed at the vir F gene, which is present only in pathogenic strains of Y. enterocolitica. Pathogenic strains of Y. enterocolitica can be recognized by growth at 37°C on Yersinia selective agar.