[Intoxication from accidental ingestion of cannabis: analysis of eight cases]. / À propos de 8 cas d'intoxication accidentelle au cannabis chez des nourrissons.

Consultations at pediatric emergency units for acute consciousness alterations is frequent. Miscellaneous causes include cranial trauma, meningoencephalitis, metabolic disorders, drugs, or other intoxications. We report here eight cases of infants who were brought to the emergency division due to acute consciousness failure after accidental ingestion of hashish, confirmed by urinary dosage of Δ9-tetrahydrocannabinol. This series of under 24-month-old infants only emphasizes the value of screening for cannabis in urine in cases of abnormal consciousness and/or abnormal behavior in an infant.

Expression of an evolved engineered variant of a bacterial glycine oxidase leads to glyphosate resistance in alfalfa.

The main strategy for resistance to the herbicide glyphosate in plants is the overexpression of an herbicide insensitive, bacterial 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). A glyphosate resistance strategy based on the ability to degrade the herbicide can be useful to reduce glyphosate phytotoxicity to the crops. Here we present the characterization of glyphosate resistance in transgenic alfalfa (Medicago sativa L.) expressing a plant-optimized variant of glycine oxidase (GO) from Bacillus subtilis, evolved in vitro by a protein engineering approach to efficiently degrade glyphosate. Two constructs were used, one with (GO(TP+)) and one without (GO(TP-)) the pea rbcS plastid transit peptide. Molecular and biochemical analyses confirmed the stable integration of the transgene and the correct localization of the plastid-imported GO protein. Transgenic alfalfa plants were tested for glyphosate resistance both in vitro and in vivo. Two GO(TP+) lines showed moderate resistance to the herbicide in both conditions. Optimization of expression of this GO variant may allow to attain sufficient field resistance to glyphosate herbicides, thus providing a resistance strategy based on herbicide degradation.

A point mutation in the Medicago sativa GSA gene provides a novel, efficient, selectable marker for plant genetic engineering.

Bacterial selectable marker genes (SMG) conferring antibiotic resistance are valuable tools in plant genetic engineering, but public concern and regulatory requirements have stimulated the development of alternative selection systems. We have previously demonstrated that a mutated Synechococcus elongatus HemL gene encoding glutamate 1-semialdehyde aminotransferase (GSA) is an efficient SMG in alfalfa. In fact, GSA is irreversibly inhibited by gabaculine (3-amino-2,3-dihydrobenzoic acid), but the mutated enzyme is gabaculine insensitive. With the aim to develop a plant derived SMG, we cloned and sequenced the Medicago sativa GSA cDNA and reproduced one of the two mutations associated with gabaculine resistance in Synechococcus, a transversion resulting in a methionine to isoleucine (M→I) substitution. This mutated gene was assessed as a SMG in tobacco and alfalfa Agrobacterium transformation, in comparison with the wild type gene. In tobacco, about 43% of the leaf explants produced green shoots, whereas in alfalfa 47% of the explants produced green embryos in the presence of 30 µM gabaculine when the M→I GSA was introduced. Escapes were absent in tobacco and only 6% in alfalfa. No effect on the plant phenotype was noticed. We propose this new SMG as a widely acceptable alternative to those currently used.

Interleukin-1 receptor antagonist (anakinra) treatment in patients with systemic-onset juvenile idiopathic arthritis or adult onset Still disease: preliminary experience in France.

BACKGROUND: Anakinra treatment has been reported to be effective in some patients with systemic-onset juvenile idiopathic arthritis (SoJIA) or adult-onset Still disease (AoSD). OBJECTIVES: To assess the efficacy and the safety of anakinra treatment in SoJIA and AoSD. METHODS: SoJIA and AoSD patients were treated with anakinra (1-2 mg/kg/day in children, 100 mg/day in adults); we analysed its effect on fever, erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) levels, numbers of swollen and tender joints, the assessment of disease activity (by physician and parent/patient) and pain (by parent/patient), and American College of Rheumatology (ACR) pediatric core set criteria for JIA activity. RESULTS: A total of 35 patients were included, 20 with SoJIA and 15 with AoSD. Their mean age (range) at the onset of treatment was 12.4 (3-23) and 38.1 (22-62) years, respectively; disease duration was 7.0 (1-16) and 7.8 (2-27) years, respectively. Active arthritis was present in all cases but one. Of the 20 SoJIA patients, 5 achieved ACR 50% improvement in symptoms (ACR50) response criteria at 6 months. Steroid dose had been decreased by 15% to 78% in 10 cases. A total of 11 of the 15 AoSD patients achieved at least a 50% improvement for all disease markers (mean follow-up: 17.5 (11-27) months). Steroids had been stopped in two cases and the dose was decreased by 45% to 95% in 12 patients. Two patients stopped anakinra due to severe skin reaction, and two patients due to infection: one visceral leishmaniasis and one varicella. CONCLUSION: Anakinra was effective in most AoSD patients, but less than half SoJIA patients achieved a marked and sustained improvement.

Kanamycin-resistant alfalfa has a point mutation in the 16S plastid rRNA.

Genes conferring resistance to kanamycin are frequently used to obtain transgenic plants as spontaneous resistance to kanamycin is not known to exist in higher plants. Nevertheless, mutations conferring kanamycin resistance have been identified in Chlamydomonas reinhardtii, raising the question as to why kanamycin-resistant mutants have not been found in higher plants. While attempting plastid transformation of alfalfa, we obtained non-transgenic but kanamycin-resistant somatic embryos following 2 months of culture in the presence of 50 mg l(-1) kanamycin. Sequencing of the plastid DNA region corresponding to the decoding site of the 16S rRNA in ten independent resistant events revealed an A to C transversion at position 1357 of the 16S plastid rDNA, the same site at which an A to G conversion confers kanamycin resistance to C. reinhardtii by reducing the ability of the antibiotic to bind to its target site. All plants derived from the resistant embryos through additional cycles of somatic embryogenesis in the absence of kanamycin retained the mutant phenotype, suggesting that the mutation was homoplastomic. Resistant plants produced 85% less biomass than controls; their leaves were chlorotic during early development and over time slowly turned green. The absence of kanamycin- resistant mutants in higher plants might be explained by the requirement for a regeneration system capable of resulting in homoplastomic individuals, or it may be the result of the detrimental effect of the mutation on the phenotype.

Inheritance and mapping of 2n-egg production in diploid alfalfa.

The production of eggs with the sporophytic chromosome number (2n eggs) in diploid alfalfa (Medicago spp.) is mainly associated with the absence of cytokinesis after restitutional meiosis. The formation of 2n eggs through diplosporic apomeiosis has also been documented in a diploid mutant of M. sativa subsp. falcata (L.) Arcang. (2n = 2x = 16), named PG-F9. Molecular tagging of 2n-egg formation appears to be an essential step towards marker-assisted breeding and map-based cloning strategies aimed at investigating and manipulating reproductive mutants of the M. sativa complex. We made controlled crosses between PG-F9 and three wild type plants of M. sativa subsp. coerulea (Less.) Schm. (2n = 2x = 16) and then hand-pollinated the F1 progenies with tetraploid plants of M. sativa subsp. sativa L. (2n = 4x = 32). As a triploid embryo block prevents the formation of 3x progenies in alfalfa because of endosperm imbalance, and owing to the negligible selfing rate, seed set in 2x-4x crosses was used to discriminate the genetic capacity for 2n-egg production. F1 plants that exhibited null or very low seed sets were classified as normal egg producers and plants with high seed sets as 2n-egg producers. A bulked segregant analysis (BSA) with RAPD (random amplified polymorphic DNA), ISSR (inter-simple sequence repeat), and AFLP (amplified fragment length polymorphism) markers was employed to identify a genetic linkage group related to the 2n-egg trait using one of the three F1 progenies. This approach enabled us to detect a paternal ISSR marker of 610 bp, generated by primer (CA)8-GC, located 9.8 cM from a putative gene (termed Tne1, two-n-eggs) that in its recessive form determines 2n eggs and a 30% recombination genomic window surrounding the target locus. Eight additional RAPD and AFLP markers, seven of maternal, and one of paternal origin, significantly co-segregated with the trait under investigation. The minimum number of quantitative trait loci (QTLs) controlling seed set in 2x-4x crosses was estimated by ANOVA and regression analysis. Four maternal and three paternal independent molecular markers significantly affected the trait. A paternal RAPD marker allele, mapped in the same linkage group of Tne1, explained 43% of the variation for seed set in 2x-4x crosses indicating the presence of a major QTL. A map of the PG-F9 chromosome regions carrying the minor genes that determine the expression level of 2n eggs was constructed using selected RAPD and AFLP markers. Two of these genes were linked to previously mapped RFLP loci belonging to groups 1 and 8. Molecular and genetic evidence support the involvement of at least five genes.

Genotoxicity of vanadium compounds in yeast and cultured mammalian cells.

The ability of vanadium compounds to induce genetic activity was investigated in D7 and D61M strains of Saccharomyces cerevisiae and in Chinese hamster V79 cell line. In our previous work, ammonium metavanadate (pentavalent form, V5) induced mitotic gene conversion and point reverse mutation in the D7 strain of yeast. The genotoxicity was reduced by the presence of S9 fraction, which probably reduced pentavalent vanadium to the tetravalent form. In the present study, vanadyl sulfate (tetravalent form, V4) induced no convertants and revertants in yeast cells harvested from stationary growth phase. With yeast cells from logarithmic growth phase, which contain high levels of cytochrome P-450, a significant increase in genetic effects was observed. Further experiments, performed by treating cells harvested from logarithmic growth phase in the presence of cytochrome P-450 inhibitors, indicated that the monooxygenase system influenced the genotoxicity of metavanadate while the genetic activity of vanadyl remained unaffected. Aneuploidy effect in the D61M strain of Saccharomyces cerevisiae was induced by either V5 or V4, confirming that vanadium compounds are potentially antitubulin agents in eukaryotic cells. Although these compounds are very toxic in V79 cells, no mutagenic effect was observed in the presence or in the absence of S9 fraction.

Tetrachloroethane, pentachloroethane, and hexachloroethane: genetic and biochemical studies.

Tetrachloroethane (TTCE), pentachloroethane (PCE), and hexachloroethane (HCE) were tested in diploid strain (D7) of the yeast Saccharomyces cerevisiae in suspension test with and without mammalian metabolic activation (S9). TTCE, PCE, and HCE gave positive results on cells harvested from logarithmic growth phase; only PCE induced a significant increase (P less than or equal to .01) of mitotic gene conversion and point reverse mutation on cells from stationary growth phase with metabolic activation (S9). The in vivo effects on cytochrome P450 content (cyt. P450), pentoxyresorufin O-dealkylase (P450-like, class IIB, PROD), and ethoxy-resorufin O-deethylase (P448-like, class IA, EROD) activities were examined in hepatic microsomes from mice 24 h after acute intoxication. All the halogenated hydrocarbons displayed a marked toxic effect as shown by the significant decrease in cyt. P450 levels (maximum of 76% decrease, with TTCE 753.2 mg/kg) and EROD (maximum of 69% decrease, with PCE 925.4 mg/kg), and to a lesser extent in PROD (maximum of 52.4% decrease, with HCE 3150 mg/kg). Although a general decrease of P450 functions was observed, the toxic effects of TTCE and PCE seem to be preferentially related to P448 forms.

A new method for the purification of the different stages of carrot embryoids.

An easy method is presented for the purification of the different stages of carrot embryoids. This is based on a synchronization of the regenerating culture and on a filtration through filters of various pore sizes. A differential sedimentation was used for removing undifferentiated cells. At the end of the process, the different stages: globular, heart- and torpedo-shaped were obtained with a degree of purity that always exceeded 90%. This method can be used for the separation of relatively large numbers of embryoids (from thousands to a million) of haploid and diploid carrot lines and is very gentle on embryoids in that it does not affect their viability or further development.

Induction of gene mutations and gene conversions by vinyl chloride metabolites in yeast.

Chloroethylene oxide and 2-chloroacetaldehyde, two metabolites of vinyl chloride, and 2-chloroethanol, a putative metabolic intermediate, were assayed for their genetic activity in the yeasts Schizosaccharomyces pombe and Saccharomyces cerevisiae. Chloroethylene oxide was found to be the most effective in inducing forward mutations in Sch. pombe and gene conversions in S. cerevisiae, increasing the mutation and conversion frequencies 340 and 50 times, respectively, over those of the controls. In either the presence or the absence of mouse liver microsomes, 2-chloroacetaldehyde showed only feeble genetic activity, and 2-chloroethanol was completely inactive in both yeast strains. In contrast to vinyl chloride, 2-chloroacetaldehyde did not induce forward mutations in Sch. pombe inthe host-mediated assay in mice. The results strongly support the hypothesis that chloroethylene oxide is one of the principal mutagenic agents formed from vinyl chloride in the presence of mouse liver enzymes.

Mutagenicity of industrial compounds: styrene and its possible metabolite styrene oxide.

Styrene and its presumed metabolite, styrene oxide, were tested for their mutagenic effect on a forward mutation system of yeast and of Chinese hamster cells, and on a gene-conversion system of yeast. Experiments with liver microsomal preparations and host-mediated assay with yeast were also carried out. Styrene oxide was mutagenic in all test systems. Styrene was mutagenic only in the host-mediated assay.