Spastin mutations are frequent in sporadic spastic paraparesis and their spectrum is different from that observed in familial cases.

BACKGROUND: SPG4 encodes spastin, a member of the AAA protein family, and is the major gene responsible for autosomal dominant spastic paraplegia. It accounts for 10-40% of families with pure (or eventually complicated) hereditary spastic paraparesis (HSP). OBJECTIVE: To assess the frequency of SPG4 mutation in patients with spastic paraplegia but without family histories. METHODS: 146 mostly European probands with progressive spastic paraplegia were studied (103 with pure spastic paraplegia and 43 with additional features). Major neurological causes of paraplegia were excluded. None had a family history of paraplegia. DNA was screened by DHPLC for mutations in the 17 coding exons of the SPG4 gene. Sequence variants were characterised by direct sequencing. A panel of 600 control chromosomes was used to rule out polymorphisms. RESULTS: The overall rate of mutations was 12%; 19 different mutations were identified in 18 patients, 13 of which were novel. In one family, where both parents were examined and found to be normal, the mutation was transmitted by the asymptomatic mother, indicating reduced penetrance. The parents of other patients were not available for analysis but were reported to be normal. There was no evidence for de novo mutations. The mutations found in these apparently isolated patients were mostly of the missense type and tended to be associated with a less severe phenotype than previously described in patients with inherited mutations. CONCLUSIONS: The unexpected presence of SPG4 gene mutations in patients with sporadic spastic paraplegia suggests that gene testing should be done in individuals with pure or complicated spastic paraplegia without family histories.

Burkholderia tropica sp. nov., a novel nitrogen-fixing, plant-associated bacterium.

In an ecological survey of nitrogen-fixing bacteria isolated from the rhizosphere and as endophytes of sugarcane, maize and teosinte plants in Brazil, Mexico and South Africa, a new phylogenetically homogeneous group of N(2)-fixing bacteria was identified within the genus Burkholderia. This polyphasic taxonomic study included microscopic and colony morphology, API 20NE tests and growth on different culture media at different pH and temperatures, as well as carbon source assimilation tests and whole-cell protein pattern analysis. Analysis of 16S rRNA gene sequences showed 99.2-99.9 % similarity within the novel species and 97.2 % similarity to the closest related species, Burkholderia sacchari. The novel species was composed of four distinct amplified 16S rDNA restriction analysis groups. The DNA-DNA reassociation values within the novel species were greater than 70 % and less than 42 % for the closest related species, B. sacchari. Based on these results and on many phenotypic characteristics, a novel N(2)-fixing species is proposed for the genus Burkholderia, Burkholderia tropica sp. nov., with the type strain Ppe8(T) (=ATCC BAA-831(T)=LMG 22274(T)=DSM 15359(T)). B. tropica was isolated from plants grown in geographical regions with climates ranging from temperate subhumid to hot humid.

Expression, purification, and characterization of recombinant purine nucleoside phosphorylase from Escherichia coli.

Recombinant purine nucleoside phosphorylase (PNPase) from Escherichia coli was prepared in high yield in order to facilitate its use in coupled assays to measure the kinetics of phosphate-liberating enzymes. The E. coli enzyme was overexpressed in E. coli by inserting the genomic fragment containing the deoD gene downstream of the isopropyl beta-d-thiogalactoside-inducible promotor of pSE380 expression vector. The recombinant protein was purified to approximately 90% homogeneity and with a yield of approximately 9000 units of activity/L of culture, using an efficient one-column procedure. A continuous spectrophotometric assay coupling P(i) release to the phosphorolysis of the nucleoside analogue 7-methylinosine (m(7)Ino) was recently described. Here, we report the steady-state kinetic parameters of the recombinant E. coli PNPase catalyzed reaction with m(7)Ino and P(i) as substrates and compare these parameters with those of a bacterial PNPase commercially available for use in coupled assays. Under the assay conditions described, the recombinant E. coli protein is active at higher pH values and is stable up to a temperature of approximately 55 degrees C and following multiple freeze-thaw cycles. It is activated by high ionic strength but loses some activity following dialysis or concentration under pressure. Finally, a new procedure for the synthesis of m(7)Ino from inosine is described which is safe and cost effective, making the use of this methylated nucleoside in PNPase-coupled P(i) assays more attractive.

[Food consumption of infants and young children in France in 1997]. / Consommation alimentaire des nourrissons et des enfants en bas âge en France en 1997.

BACKGROUND: In 1981 and 1989, two French nationwide food consumption surveys reported the dietary intakes of infants and toddlers. In 1997, another survey was conducted. POPULATION AND METHODS: Six-hundred and sixty, 1 to 30-month-old infants and children were recruited. Food records were completed by their parents during a 3-day period. Energy, proteins, carbohydrates, total lipids, minerals, vitamins, linoleic acid contents were calculated. Energy contribution of various food groups and the pattern of the distribution of the different meals were indicated. RESULTS: Between 1989 and 1997, the contribution of milk-based infant formulas increased at 4, 5, 7 and 8-9 months. From the age of 5 months, approximately 75% of the children had a protein intake twice as high as the official recommendations. Mean iron intakes were higher than those calculated in 1989 at the age of 7, 8-9 and 10-12 months. The average daily intake of linoleic acid was lower than the recommended daily allowance from the age of 6 months. CONCLUSION: Trends in dietary intakes between 1989 and 1997 indicated that the nutritional needs of French infants and toddlers are better covered as recommended by scientific committees. More effective efforts are needed for improvement of their nutritional status particularly for iron and essential fatty acids. For example, weaning foods should be introduced later and toddler's formulas should be used more systematically and longer.

Hydrocortisone 17-butyrate degradation in the presence of micro-organisms.

OBJECTIVE: This study compared the degradation of hydrocortisone 17-butyrate (H17B) in the presence of six different bacteria, commonly found on psoriatic skin. METHOD: H17B and its degradation products (hydrocortisone and hydrocortisone 21-butyrate (H21B)) were assayed by HPLC. RESULTS: In the absence of micro-organisms, we observed 16.6 +/- 7.1% degradation. In the presence of micro-organisms and otherwise similar conditions, we noted that H17B degradation was not modified by cocci (Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus agalactiae). Three bacilli increased degradation, Escherichia coli 59.1 +/- 19.4%, Klebsiella oxytoca 62.1 +/- 6.7% and Pseudomonas aeruginosa 56.0 +/- 17.9%. CONCLUSION: The degradation of H17B into hydrortisone and H21B may produce a loss of therapeutic activity.

Stability study of cloxacillin in an oily suspension, by high performance liquid chromatography.

A high performance liquid chromatography method, using spectrophotometric detection (254 nm) has been developed for the analysis of cloxacillin in an oily suspension. Analysis was performed by isocratic elution with 0.02M potassium dihydrogen phosphate solution, methanol, 50/50 (V/V). The method was specific, linear, accurate (99.8 +/- 1.4%) and precise (less than 1.0%). A stability study of cloxacillin in the formulation, showed that about 10% degradation occurred for 6 months storage at room temperature, when lyophilized sodium cloxacillin was used. On the contrary, when sodium cloxacillin obtained by precipitation from non aqueous solvent was used, about 6% degradation appeared in the drug for 3 years storage in the same conditions.