Effects of three novel metalloproteinases from the venom of the West African saw-scaled viper, Echis ocellatus on blood coagulation and platelets

Two metalloproteinases, a 24-kDa P-I EoVMP1 and a 56-kDa P-III EoVMP2, have recently been isolated from the venom of the West African saw-scaled viper Echis ocellatus. We now reveal a new 65-kDa haemorrhagic group P-III metalloproteinase which we have designated EoVMP3. The aim of this study was to determine whether these three snake venom metalloproteinases (SVMPs) affect platelets and blood coagulation. EoVMP1 had no effect on the aggregation of washed human platelets, whereas EoVMP2 inhibited collagen-induced platelet aggregation. In contrast, EoVMP3 did not inhibit the aggregation of platelets by collagen but instead activated platelets in the absence of any additional co-factors. All three SVMPs were capable of activating prothrombin to varying degrees and can therefore be described as procoagulants. EoVMP1, EoVMP2 and EoVMP3 share sequence identity with other members of the reprolysin family, but differ greatly in their effects on some of the components that control haemostasis.     

Spatial organization of dual-species bacterial aggregates on leaf surfaces

The spatial organization of cells within bacterial aggregates on leaf surfaces was determined for pair-wise mixtures of three different bacterial species commonly found on leaves, Pseudomonas syringae, Pantoea agglomerans, and Pseudomonas fluorescens. Cells were coinoculated onto bean plants and allowed to grow under moist conditions, and the resulting aggregates were examined in situ by epifluorescence microscopy. Each bacterial strain could be localized because it expressed either the green or the cyan fluorescent protein constitutively, and the viability of individual cells was assessed by propidium iodide staining. Each pair of bacterial strains that was coinoculated onto leaves formed mixed aggregates. The degree of segregation of cells in mixed aggregates differed between the different coinoculated pairs of strains and was higher in mixtures of P. fluorescens A506 and P. agglomerans 299R and mixtures of P. syringae B728a and P. agglomerans 299R than in mixtures of two isogenic strains of P. agglomerans 299R. The fractions of the total cell population that were dead in mixed and monospecific aggregates of a gfp-marked strain of P. agglomerans 299R and a cfp-marked strain of P. agglomerans 299R, or of P. fluorescens A506 and P. agglomerans 299R, were similar. However, the proportion of dead cells in mixed aggregates of P. syringae B728a and P. agglomerans 299R was significantly higher (13.2% +/- 8.2%) than that in monospecific aggregates of these two strains (1.6% +/- 0.7%), and it increased over time. While dead cells in such mixed aggregates were preferentially found at the interface between clusters of cells of these strains, cells of these two strains located at the interface did not exhibit equal probabilities of mortality. After 9 days of incubation, about 77% of the P. agglomerans 299R cells located at the interface were dead, while only about 24% of the P. syringae B728a cells were dead. The relevance of our results to understanding bacterial interactions on leaf surfaces and the implications for biological control of pathogenic and other deleterious microorganisms is discussed.     

Fate of epoxiconazole and kresoxim-methyl in wheat according to time of application

During the 2000-2001 season a field trial was conducted with the aim of quantifying the distribution and persistence of epoxiconazole and kresoxlm-methyl in the different leaf layers of winter wheat plants. In the case of applications before flag leaf emergence, the redistribution of the two active ingredients in the newly formed leaves following the applications was also measured. Allegro (125 g/L epoxiconazole and 125 g/L kresoxim-methyl, SC) was applied at the manufacturer's recommended rate (1 L/ha) either in a single treatment at stages GS32, GS39 and GS59 or in 2, 3 or 4 split applications. Following spraying, leaf samples were collected over time, from each leaf layer, and the two active ingredients were quantified by gas chromatography with electron capture detection (GC-ECD). Fungicide distribution varies according to time of application. A descending gradient through the leaves was observed in the case of application at GS59. When sprayed at stage GS39, on the other hand, the second leaf intercepted more fungicide than the flag leaf. Kresoxim-methyl was found to degrade faster than epoxiconazole. With split treatments, the last spraying appears to be very significant in terms of final fungicide quantities. Redistribution appears possible, especially in the case of epoxiconazole, though in very small quantities.     

A new route to trihydroxamate-containing artificial siderophores and synthesis of a new fluorescent probe

A fluorescent labelled artificial siderophore 1 was synthesized by coupling a 7-nitrobenz-2-oxa-1,3-diazole (NBD) derivative to the terminal amino group of a new trihydroxamate-containing amine 2, a ferrichrome-type siderophore that was obtained from tris(hydroxymethyl)aminomethane. Compound 1 was shown to be a suitable tool for experiments on siderophore transport and uptake processes in various organisms cells and particularly in Candida albicans cells.     

Mapping quantitative trait loci associated with leaf and stem pubescence in cotton

Leaf pubescence in cotton have a potential for insect pest management. Varying degrees of leaf trichome density in Gossypium species and cultivars have been associated to a series of five genes, referred to as t(1)-t(5). We used two segregating interspecific G. hirsutum x G. barbadense backcross populations developed in our laboratory to assess qualitatively and quantitatively leaf and stem pubescence. QTL analyses were performed using simple and composite interval mapping. Based on both types of measurements and under both types of QTL analyses, nine QTLs met permutation-based thresholds. The nine QTLs mapped to four different chromosome regions. Highest LOD values corresponded to the QTLs detected on c6 (four colocalized QTLs) and on D03 (two QTLs) for which the higher pubescence in the progeny derived from the pubescent G. hirsutum parent alleles. Conversely, on c17 (one QTL) and A01 (two QTLs), the G. hirsutum parental alleles affected negatively pubescence. These results combined with another published study confirm (1) the location in a center region of chromosome 6 of the t(1) locus as a major locus/gene determining leaf pubescence, and (2) additional genes located on seven additional chromosomes have been shown to impart trichome density either positively or negatively. The existence of a high density of PCR-based loci in most of the regions identified as harboring leaf pubescence QTLs, particularly that on chromosome 6, will facilitate future efforts for map-based cloning.     

Ethanol impairs monosaccharide uptake and glycosylation in cultured rat astrocytes

Astrocyte and glial-neuron interactions have a critical role in brain development, which is partially mediated by glycoproteins, including adhesion molecules and growth factors. Ethanol affects the synthesis, intracellular transport, subcellular distribution and secretion of these glycoproteins, suggesting alterations in glycosylation. We analyzed the effect of long-term exposure to low doses of ethanol (30 mm) on glycosylation process in growing cultured astrocytes in vitro. Cells were incubated for short (5 min) and long (90 min) periods with several radioactively labeled carbohydrate precursors. The uptake, kinetics and metabolism of these precursors, as well as the radioactivity distribution in protein gels were analyzed. The levels of GLUT1 and mannosidase II were also determined. Ethanol increased the uptake of monosaccharides and the protein levels of GLUT1 but decreased those of mannosidase II. It altered the carbohydrate moiety of proteins and increased cell surface glycoproteins containing terminal non-reduced mannose. These results indicate that ethanol impairs glycosylation in rat astrocytes, thus disrupting brain development.     

Amphinema modernisme, a new Pandeid (Cnidaria: Anthomedusae) from the Southern Ocean

A new species of pandeid anthomedusa, Amphinema modernisme, is described from a single and complete specimen collected by a mid-water sediment trap placed at 500 m depth in open waters, over a 1066-m bottom in the Drake Passage near the South Shetland Islands. This new Amphinema is characterized by an egg-shaped apical chamber, partitioned by four narrow sacciform centrifugal prolongations of the radial canals, perradial gonads and cellular strands linking the radial canals to the exumbrella. Its morphological features are compared with those of the eight other species of the genus Amphinema and a new diagnosis for this genus is presented. Accepted: 19 July 1999     

Adaptive responses for seed and leaf phenology in natural populations of sessile oak along an altitudinal gradient

We assessed the adaptive potential of seed and leaf phenology in 10 natural populations of sessile oak (Quercus petraea) sampled along two altitudinal transects using common garden experiments. Population differentiation for both phenological traits was observed with high-altitude populations germinating and flushing later than low altitude ones. However, high genetic variation and heritability values were also maintained within populations, despite slightly decreasing for dates of leaf unfolding with increasing altitude. We suggest that biotic and abiotic fluctuating selection pressures within populations and high gene flow are the main mechanisms maintaining high genetic variation for these fitness related traits. Moreover, changes in selection intensity and/or selection pressures along the altitudinal gradient can explain the reduction in genetic variation observed for leaf phenology. We anticipate that the maintenance of high genetic variation will be a valuable resource for future adaptation of sessile oak populations undergoing an upslope shift caused by climate change.