Functional analysis of the copy 1 of the fixNOQP operon of Ensifer meliloti under free-living micro-oxic and symbiotic conditions.

AIM: In this work, phenotypic analyses of a Ensifer meliloti fixN1 mutant under free-living and symbiotic conditions have been carried out. METHODS AND RESULTS: Ensifer meliloti fixN1 mutant showed a defect in growth as well as in TMPD-dependent oxidase activity when cells were incubated under micro-oxic conditions. Furthermore, haem c staining analyses of a fixN1 and a fixP1 mutant identified two membrane-bound c-type cytochromes of 27 and 32 kDa, present in microaerobically grown cells and in bacteroids, as the FixO and FixP components of the E. meliloti cbb3 oxidase. Under symbiotic conditions, fixN1 mutant showed a clear nitrogen fixation defect in alfalfa plants that were grown in an N-free nutrient solution during 3 weeks. However, in plants grown for a longer period, fixNOQP1 copy was not indispensable for symbiotic nitrogen fixation. CONCLUSIONS: The copy 1 of the fixNOQP operon is involved in E. meliloti respiration and growth under micro-oxic conditions as well as in the expression of the FixO and FixP components of the cbb3 oxidase present in free-living microaerobic cultures and in bacteroids. This copy is important for nitrogen fixation during the early steps of the symbiosis. SIGNIFICANCE AND IMPACT OF THE STUDY: It is the first time that a functional analysis of the E. meliloti copy 1 of the fixNOQP operon is performed. In this work, the cytochromes c that constitute the cbb3 oxidase operating in free-living micro-oxic cultures and in bacteroids of E. meliloti have been identified.

Enhanced expression of Rhizobium etli cbb3 oxidase improves drought tolerance of common bean symbiotic nitrogen fixation.

To investigate the involvement of Rhizobium etli cbb(3) oxidase in the response of Phaseolus vulgaris to drought, common bean plants were inoculated with the R. etli strain, CFNX713, overexpressing this oxidase in bacteroids (cbb(3)(+)) and subjected to drought conditions. The negative effect of drought on plant and nodule dryweight, nitrogen content, and nodule functionality was more pronounced in plants inoculated with the wild-type (WT) strain than in those inoculated with the cbb(3)(+) strain. Regardless of the plant treatment, bacteroids produced by the cbb(3)(+) strain showed higher respiratory capacity than those produced by the WT strain. Inoculation of plants with the cbb(3)(+) strain alleviated the negative effect of a moderate drought on the respiratory capacity of bacteroids and the energy charge of the nodules. Expression of the FixP and FixO components of the cbb(3) oxidase was higher in bacteroids of the cbb(3)(+) strain than in those of the WT strain under all experimental conditions. The decline in sucrose synthase activity and the decrease in dicarboxylic acids provoked by moderate drought stress were more pronounced in nodules from plants inoculated with the WT strain than in those inoculated with the cbb(3)(+) strain. Taken together, these results suggest that inoculation of plants with a R. etli strain having enhanced expression of cbb(3) oxidase in bacteroids reduces the sensitivity of P. vulgaris-R. etli symbiosis to drought and can modulate carbon metabolism in nodules.

Theoretical study of the neutral hydrolysis of hydrogen isocyanate in aqueous solution via assisted-concerted mechanisms.

A procedure is described for the theoretical study of chemical reactions in solution by means of molecular dynamics simulation, with solute-solvent interaction potentials derived from ab initio quantum calculations. We apply the procedure to the case of neutral hydrogen isocyanate hydrolysis, HNCO + 2 H(2)O --> H(2)NCOOH + H(2)O, in aqueous solution, via the assisted-concerted mechanisms and the two-water model. We used the solvent as a reaction coordinate and the free-energy curves for the calculation of the properties related to the reaction mechanisms, with a particular focus on the reaction and activation energies. The results showed that the mechanism with two water molecules attacking the C=N bond is preferred to the mechanism with three waters forming a ring of eight members. In addition, the aqueous medium significantly reduces the activation barrier (DeltaG(double dagger) = 13.9 kcal/mol) and makes the process more exothermic (DeltaG = -11.1 kcal/mol) relative to the gas-phase reaction, increasing the rate constant of the process to k = 4.25 x 10(5) s(-1).

[Cardiovascular risk and the new diagnostic categories for diabetes mellitus proposed by the American Diabetes Association]. / Riesgo cardiovascular asociado a las nuevas categorías diagnósticas de la diabetes mellitus propuestas por la Asociación Americana de Diabetes.

OBJECTIVE: To study the cardiovascular risk (CVR) associated with the diagnostic criteria for diabetes proposed by the American Diabetes Association. DESIGN: Cross-sectional, descriptive study. SETTING: Urban health district. PATIENTS: 1840 patients > 14 years old, selected by simple randomised sampling from the clinical records (CR) archive, were studied. METHOD: Through review of the CR, the patients were classified as having: normal glycaemia, disturbed basal glycaemia (DBG) and type-2 diabetes mellitus (DM2). CVR was studied through the simplified Framingham method and the CVR factors of obesity, tobacco dependency, hypertension, hypercholesterolaemia and hypertriglyceridaemia. The likelihood of having high or very high CVR and the CVR factors described in the patients with DBG or DM2 were compared with the same in those with normal glycaemia, through logistical regression with the odds ratio adjusted for age and sex. RESULTS: 1351 patients were classified: 995 with normal glycaemia, 146 with DBG and 210 with DM2. Patients with DBG or DM2 had greater likelihood of high or very high CVR, with some OR at 2.26 (95% CI, 1.39-3.69) and 2.74 (95% CI, 1.81-4.15), respectively. They also had differences (p < 0.05) for obesity (OR, 1.76 and 1.58), hypertension (OR, 1.75 and 2.15) and hypertriglyceridaemia (OR, 1.73 and 2.70), respectively. There were no differences (p > 0.05) for tobacco dependency and hypercholesterolaemia. No differences were found (p > 0.05) between DBG and DM2 for CVR and the CVR factors studied. CONCLUSIONS: Patients with DBG and DM2 are at high CVR.

[Attitude of primary care physicians to early detection of prostatic cancer through prostate-specific antigen]. / Actitud del médico de atención primaria en la detección precoz del cáncer de próstata mediante el antígeno prostático específico.

OBJECTIVES: To study the attitude of primary care doctors when a high (> or = 4 ng/ml) prostate-specific antigen (PSA) is found and to examine the variables linked to a prostate biopsy and the diagnosis of prostate cancer (PC). DESIGN: Descriptive, observational study. SETTING: Urban health district. PATIENTS: Ninety-four men not previously diagnosed with PC who in 1998 had a PSA figure > or = 4 ng/ml. The list was obtained from the pertinent laboratory. MEASUREMENTS: The following variables were gathered from review of clinical records: family background of PC, age, PSA figure, reason for request for PSA (if not given, it was considered a screening), referral to the urologist, rectal touch, transrectal echography, prostate biopsy and final diagnosis. RESULTS: Average age was 70 (SD, 9.31). The reason for requesting PSA was: urine symptoms in 25 (26.6%), other signs or symptoms in 25 (26.6%), request of patient in 2 cases (2.1%) and screening in 42 (44.7%). Rectal touch took place in 16 cases. Twenty-nine people were referred for examination to the urologist. 36 patients had an echography and biopsy. Variables linked to the prostate biopsy in the logistic model were: higher value of the PSA (OR 1.1; 95% CI, 1.03-1.18), being older (OR 0.92; CI, 0.87-0.98) and rectal touch performed (OR 3.58; CI, 1.02-12.51). Ten cases of PC were diagnosed. CONCLUSIONS: The most common reason for a PSA request was screening. Prostate biopsy was not requested for 58 men. A primary care guide to action concerning PC diagnosis in cases of PSA > or = 4 ng/ml would be useful.