Bacillus subtilis genome project: cloning and sequencing of the 97 kb region from 325° to 333deg;

In the framework of the European project aimed at the sequencing of the Bacillus subtilis genome the DNA region located between gerB (314°) and sacXV (333°) was assigned to the Institut Pasteur. In this paper we describe the cloning and sequencing of a segment of 97 kb of contiguous DNA. Ninety-two open reading frames were predicted to encode putative proteins among which only forty-two were found to display significant similarities to known proteins present in databanks, e.g. amino acid permeases, proteins involved in cell wall or antibiotic biosynthesis, various regulatory proteins, proteins of several dehydrogenase families and enzymes II of the phosphotransferase system involved in sugar transport. Additional experiments led to the identification of the products of new B. subtilis genes, e.g. galactokinase and an operon involved in thiamine biosynthesis.     

Bioguided Fractionation and Isolation of an Antiplasmodial Saponin from the Roots of Nauclea xanthoxylon (A.Chev.) Aubrév. (Rubiaceae)

The root extract of Nauclea xanthoxylon (A.Chev.) Aubrév. displayed significant 50 % inhibition concentration (IC₅₀s) of 0.57 and 1.26 μg/mL against chloroquine resistant and sensitive Plasmodium falciparum (Pf) Dd2 and 3D7 strains, respectively. Bio-guided fractionation led to an ethyl acetate fraction with IC₅₀s of 2.68 and 1.85 μg/mL and subsequently, to the new quinovic acid saponin named xanthoxyloside ( 1 ) with IC₅₀s of 0.33 and 1.30 μM, respectively against the tested strains. Further compounds obtained from ethyl acetate and hexane fractions were the known clethric acid ( 2 ), ursolic acid ( 3 ), quafrinoic acid ( 4 ), quinovic acid ( 5 ), quinovic acid 3-O-β-D-fucopyranoside ( 6 ), oleanolic acid ( 7 ), oleanolic acid 3-acetate ( 8 ), friedelin ( 9 ), β-sitosterol ( 10a ), stigmasterol ( 10b ) and stigmasterol 3-O-β-D-glucopyranoside ( 11 ). Their structures were characterised with the aid of comprehensive spectroscopic methods (1 and 2D NMR, Mass). Bio-assays were performed using nucleic acid gel stain (SYBR green I)-based fluorescence assay with chloroquine as reference. Extracts and compounds exhibited good selectivity indices (SIs) of >10. Significant antiplasmodial activities measured for the crude extract, the ethyl acetate fraction and xanthoxyloside ( 1 ) from that fraction can justify the use of the root of N. xanthoxylon in ethnomedicine to treat malaria.     

The electrophoretic polymorphism of bacterial esterases

Abstract: The specific activities of esterases and certain other molecular properties including immunospecificity indicate that the electrophoretic variations of these enzymes in bacterial populations are the result of allelic variations at specific gene loci. The esterase polymorphism of Enterobacteriaceae and some other species isolated from man or animals demonstrates that esterases can distinguish between bacteria at the species or subspecies level, both by their biochemical properties and by their electrophoretic differences. The esterase data complement DNA hybridization studies and agree with ribosomal DNA polymorphism, especially for delineating a phylogenetically distinct group of highly pathogenic strains in Escherichia coli . A two-dimensional electrophoretic profile obtained by establishing a direct correspondence between homologous esterase bands resolved by independent runs of isoelectric focusing and standard electrophoresis considerably improves the detection of allelic variations, whereas protein titration curves (electrophoresis in pH gradient) can be used to demonstrate the real electrophoretic homogeneity of allozymes or evalue their molecular relationship in terms of apparent amino acid substitutions. This overview establishes that esterases, by their significant electrophoretic polymorphism, are reliable molecular markers for systematics and epidemiology, and are suitable enzyme systems for studying population genetics and phylogeny.     

Seasonal incidence and ecology of the tick Ixodes ricinus (Acari: Ixodidae) on grazing pastures in Western France

A longitudinal survey was carried out during a 2 year period in Western France to assess the infestation level of grazing pastures byIxodes ricinus ticks. Four farms were visited once a month and each of the grazing pastures was sampled in the centre and at the border using the blanket dragging method. A total of 3562I. ricinus (34 adults, 900 nymphs and 2628 larvae) were collected and the infestation was significantly higher during the first year (p<0.0001). The infestation level byI. ricinus varied between grazing pastures and farms. Grazing pastures in the vicinity of forest were more infested than the others, all through the study. The seasonal distribution of ticks showed peaks, with low fluctuations between farms, years and stages. Tick abundance could not be related to vegetation, but only to the vicinity of woods.     

The effect of temperature on eicosane substrate uptake modes by a marine bacterium Pseudomonas nautica strain 617: relationship with the biochemical content of cells and supernatants

Three hydrocarbon uptake modes (adherence, emulsification and solubilization) were identified and quantified in cells and supernatants of a mesophilic marine bacterium Pseudomonas nautica strain 617 grown on eicosane. The adherence capacity was related to the enrichment of cells with wax esters and glycolipids. The emulsifying activity was related to the presence of extracellular biosurfactants composed of proteins, carbohydrates and lipids (35:63:2). The intensity of substrate uptake modes was sensitive to temperatures currently found in the original environment of P. nautica (16°C, 20°C and 32°C). When temperature decreased, a significant increase in adherence and emulsifying activity was observed in relation to biochemical changes, whereas solubilizing activity decreased. The marine bacterium was able to degrade 53–59% eicosane at the end of exponential growth after 13, 5 and 3 days incubation at 16°C, 20°C and 32°C respectively.     

Transfection of Reaggregating Embryonic Chicken Retinal Cells with an Antisense 5'-DNA Butyrylcholinesterase Expression Vector Inhibits Proliferation and Alters Morphogenesis

Abstract: The function of the enzyme butyrylcholinesterase (BChE) in the developing and mature brain is still unclear. We have inserted 577 bp of the 5' upstream region plus 106 bp of the exon 1 of the rabbit BChE gene in reverse orientation under control of an SV40 early promoter derivative in an expression vector. This vector was introduced by calcium phosphate-mediated transfection into embryonic chicken retina cells during the first days of reaggregation culture. Depending on the retinal origin, the transfected cell population forms histotypic retina-like spheres, so-called rosetted or stratified retinospheroids. We show that antisense 5'-BChE gene expression decreased the steady-state mRNA level of BChE and the translation of the BChE protein, inhibited proliferation, and accelerated histogenesis in both cellular systems. The pronounced effects of antisense 5'-BChE transfection of spheroids document a key role of BChE during the early reaggregation process of retinal cells, most likely by regulating their growth and differentiation.