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Biomedical subjects

J L Ramos

Publications and source records attributed to J L Ramos.

At least 73 records · Page 4Linked to original sources
PubMed1998

[Predictive value of the "Clinical Risk Index for Babies" for the risk of neonatal death].

OBJECTIVE: Several indicators, mainly birthweight and gestational age, have been used to predict the mortality risk in neonatal intensive care units. In order to assess the potential value of CRIB in predicting neonatal mortality, the score was used over the first 12 hours of life of the newborns admitted to this unit, during the year of 1996. METHOD: The inclusion criteria consisted of all infants without inevitably lethal congenital malformations, birthweight below 1,500 g and/or gestational age less than 31 weeks. Newborn children who died within 12 hours after delivery were excluded. The CRIB score covers birth weight, gestational age, the presence of congenital malformations (not inevitably lethal) and three indexes of physiological status during first 12 hours after birth-maximum and minimum appropriate fraction of inspired oxygen and maximum (most acidotic) base excess. RESULTS: In a prospective cohort, seventy one newborn children were studied. The birthweight (average) was 1,119 +/- 275.6 g, gestational age 30 weeks 4/7 +/- 2 weeks 3/7; male (57%); Apgar 1(0) min. score < or = 3 (36.2%) and Apgar 5 degrees min. score < 5 (5.8%). The mortality rate was 29.6% (gold standard). But mortality rate by birthweight less than 1,000 gr. or gestational age lower than 29 weeks was 60.0% and for the CRIB score above 10 was 100%. DISCUSSION: The specificity and predictive positive values for CRIB score above 10 were greater than any other two parameters. The area under the receiver operating characteristic (ROC) curve for predicting death was significantly greater for CRIB than for birthweight alone. It was concluded that the CRIB score is a better predictive indicator for mortality than are birthweight and gestational age.

Birth Weight↗
PubMed1997

Mechanisms for solvent tolerance in bacteria.

The development of tolerance in Pseudomonas putida DOT-T1 to toluene and related highly toxic compounds involves short- and long-term responses. The short-term response is based on an increase in the rigidity of the cell membrane by rapid transformation of the fatty acid cis-9,10-methylene hexadecanoic acid (C17:cyclopropane) to unsaturated 9-cis-hexadecenoic acid (C16:1,9 cis) and subsequent transformation to the trans isomer. The long-term response involves in addition to the changes in fatty acids, alterations in the level of the phospholipid polar head groups: cardiolipin increases and phosphatidylethanolamine decreases. The two alterations lead to increased cell membrane rigidity and should be regarded as physical mechanisms that prevent solvent penetrance. Biochemical mechanisms that decrease the concentration of toluene in the cell membrane also take place and involve: (i) a solvent exclusion system and (ii) metabolic removal of toluene via oxidation. Mutants unable to carry out cis --> trans isomerization of unsaturated lipids, that exhibit altered cell envelopes because of the lack of the OprL protein, or that are unable to exclude toluene from cell membranes are hypersensitive to toluene.

Adaptation, Physiological↗
PubMed1997

Removal of high concentrations of nitrate from industrial wastewaters by bacteria.

Klebsiella oxytoca isolate 15 was isolated from the grounds of a nitration factory and was found to be tolerant to nitrate at concentrations up to 0.5 to 1 M. Physicochemical parameters for optimal growth conditions for K. oxytoca isolate 15 were established. Growth took place when the nitrate concentration in the medium was less than 150 mM, and full nitrate consumption required about 14 g of C per g of N. This strain was able to remove nitrate without accumulating nitrite. The system was scaled up to a 40-liter pilot plant and was operated on-site satisfactorily.

Journal Article↗
PubMed1997

Transcriptional control of the multiple catabolic pathways encoded on the TOL plasmid pWW53 of Pseudomonas putida MT53.

The TOL plasmid pWW53 encodes a catabolic pathway for the metabolism of toluene. It bears an upper-pathway operon for the oxidation of toluene to benzoate and a copy of the gene that encodes regulatory protein XylR. For metabolism of the aromatic carboxylic acid, it bears two functional homologous meta-pathway operons, together with two functional copies of the xylS regulatory gene (xylS1 and xylS3). In cells growing in the absence of pathway substrates, no mRNA from upper- and meta-pathway operons were found; however, the xylR gene was expressed from two sigma70-dependent tandem promoters, and the xylS1 and the xylS3 genes were also expressed from their sigma70-dependent promoters, called Ps2 and Ps3, respectively. In cells grown in the presence of o-xylene, the XylR protein became active and stimulated transcription from the Pu promoter for the upper pathway. Expression from xylS1 but not from xylS3 was also stimulated by XylR; this was due to activation of transcription from the xylS1 Ps1 promoter, which is sigma54 dependent, and the lack of effect on expression from the Ps2 sigma70-dependent promoter. As a result of overexpression of the xylS1 gene, the XylS1 protein was overproduced and activated transcription from Pm1 and Pm2. In cells growing on benzoate, the upper-pathway operon was not expressed, but both meta operons were expressed. Given that XylS1 but not XylS3 recognized benzoate as an effector, stimulation of transcription was found to be mediated by XylS1. This was confirmed with cloned meta-pathway promoters and regulators. When 3-methylbenzoate was present in the medium, both meta operons were also expressed and stimulation of transcription was mediated by both XylS1 and XylS3, which both recognized 3-methylbenzoate as an effector.

Bacterial Proteins↗
PubMed1997

In vivo construction of a hybrid pathway for metabolism of 4-nitrotoluene in Pseudomonas fluorescens.

Pseudomonas fluorescens 410PR grows on 4-nitrobenzoate but does not metabolize 4-nitrotoluene. The TOL pWW0 delta pm plasmid converts 4-nitrotoluene into 4-nitrobenzoate through its upper pathway, but it does not metabolize 4-nitrobenzoate. P. fluorescens 410PR(pWW0 delta pm) transconjugants were isolated and found to be able to grow on 4-nitrotoluene. This phenotype was stable after growth for at least 300 generations without any selective pressure. P. fluorescens 410PR(pWW0 delta pm) converted 4-nitrotoluene into 4-nitrobenzoate via 4-nitrobenzylalcohol and 4-nitrobenzaldehyde. 4-Nitrobenzoate was metabolized via 4-hydroxylaminobenzoate and finally yielded NH4+ and 3,4-dihydroxybenzoate, which was mineralized.

Biotransformation↗
PubMed1997

Arac/XylS family of transcriptional regulators.

The ArC/XylS family of prokaryotic positive transcriptional regulators includes more than 100 proteins and polypeptides derived from open reading frames translated from DNA sequences. Members of this family are widely distributed and have been found in the gamma subgroup of the proteobacteria, low- and high-G + C-content gram-positive bacteria, and cyanobacteria. These proteins are defined by a profile that can be accessed from PROSITE PS01124. Members of the family are about 300 amino acids long and have three main regulatory functions in common: carbon metabolism, stress response, and pathogenesis. Multiple alignments of the proteins of the family define a conserved stretch of 99 amino acids usually located at the C-terminal region of the regulator and connected to a nonconserved region via a linker. The conserved stretch contains all the elements required to bind DNA target sequences and to activate transcription from cognate promoters. Secondary analysis of the conserved region suggests that it contains two potential alpha-helix-turn-alpha-helix DNA binding motifs. The first, and better-fitting motif is supported by biochemical data, whereas existing biochemical data neither support nor refute the proposal that the second region possesses this structure. The phylogenetic relationship suggests that members of the family have recruited the nonconserved domain(s) into a series of existing domains involved in DNA recognition and transcription stimulation and that this recruited domain governs the role that the regulator carries out. For some regulators, it has been demonstrated that the nonconserved region contains the dimerization domain. For the regulators involved in carbon metabolism, the effector binding determinants are also in this region. Most regulators belonging to the AraC/XylS family recognize multiple binding sites in the regulated promoters. One of the motifs usually overlaps or is adjacent to the -35 region of the cognate promoters. Footprinting assays have suggested that these regulators protect a stretch of up to 20 bp in the target promoters, and multiple alignments of binding sites for a number of regulators have shown that the proteins recognize short motifs within the protected region.

Amino Acid Sequence↗
PubMed1997

Transcriptional control of the Pseudomonas TOL plasmid catabolic operons is achieved through an interplay of host factors and plasmid-encoded regulators.

The xyl genes of Pseudomonas putida TOL plasmid that specify catabolism of toluene and xylenes are organized in four transcriptional units: the upper-operon xylUWCAMBN for conversion of toluene/xylenes into benzoate/alkylbenzoates; the meta-operon xylXYZLTEGFJQKIH, which encodes the enzymes for further conversion of these compounds into Krebs cycle intermediates; and xylS and xylR, which are involved in transcriptional control. The XylS and XylR proteins are members of the XylS/AraC and NtrC families, respectively, of transcriptional regulators. The xylS gene is constitutively expressed at a low level from the Ps2 promoter. The XylS protein is activated by interaction with alkylbenzoates, and this active form stimulates transcription from Pm by sigma70- or sigmaS-containing RNA polymerase (the meta loop). The xylR gene is also expressed constitutively. The XylR protein, which in the absence of effectors binds in a nonactive form to target DNA sequences, is activated by aromatic hydrocarbons and ATP; it subsequently undergoes multimerization and structural changes that result in stimulation of transcription from Pu of the upper operon. This latter process is assisted by the IHF protein and mediated by sigma54-containing RNA polymerase. Once activated, the XylR protein also stimulates transcription from the Ps1 promoter of xylS without interfering with expression from Ps2. This process is assisted by the HU protein and is mediated by sigma54-containing RNA polymerase. As a consequence of hyperexpression of the xylS gene, the XylS protein is hyperproduced and stimulates transcription from Pm even in the absence of effectors (the cascade loop). The two sigma54-dependent promoters are additionally subject to global (catabolite repression) control.

Bacterial Proteins↗
PubMed1997

[Clinical-epidemiologic indications for echocardiographic assessment in the neonatal period. Value of risk groups].

PURPOSE: An echocardiographic study was performed in newborns from risk groups, with the aim of to determining prevalence and to evaluate the indications for this test in the neonatal period. METHODS: One hundred fifty six newborns were studied. They were admitted to the Newborn ward of the Department of Obstetric of the HC-FMUSP, in the period of November 91 to April 93, from mothers with congenital heart disease or diabetes, low birth weight newborns, with extracardiac malformations, cardiac signs and/or congenital infections. RESULTS: The observed prevalence was 21.8%, greater than that of the general population (0.8-1.2%). The group composed by 27 children with extracardiac abnormalities presented the largest prevalence when compared with the other groups (40.7%). These data justify the use of echocardiography in high risk newborns for the detection of congenital heart disease.

Female↗
PubMed1996

Metabolism of nitrate esters by a consortium of two bacteria.

The products of condensation of organic alcohols and nitric acid are nitrate esters with the general structure C-O-NO2. These products are widely employed as vasodilators and explosives, and are true xenobiotic compounds, as they do not occur in nature. We have isolated and characterized a consortium of two microorganisms, Arthrobacter ilicis and Agrobacterium radiobacter, that mineralized recalcitrant ethylene glycol dinitrate. The Arthrobacter strain was the actual degrading microorganism, although the second microbe facilitated mineralization. The biodegradation of ethylene glycol dinitrate by A. ilicis involved the progressive elimination of the nitro groups from the organic molecule to generate ethylene glycol, which was then mineralized. Waters polluted with ethylene glycol dinitrate have been shown amenable to biological treatment in a pilot plant with wastewaters generated during the synthesis of the chemical in a factory.

Arthrobacter↗
PubMed1996

Catabolite repression of the toluene degradation pathway in Pseudomonas putida harboring pWW0 under various conditions of nutrient limitation in chemostat culture.

In earlier studies, the pathway of toluene and m- and p-xylene degradation (TOL pathway) in Pseudomonas putida (pWW0) was found to be subject to catabolite repression when the strain was grown at the maximal rate on glucose or succinate in the presence of an inducer. This report describes catabolite repression of the TOL pathway by succinate in chemostat cultures run at a low dilution rate (D = 0.05 h-1) under different conditions of inorganic-nutrient limitation. The activity of benzylalcohol dehydrogenase (BADH) in cell extracts was used as a measure of the expression of the TOL upper pathway. When cells were grown in the presence of 10 to 15 mM succinate under conditions of phosphate or sulfate limitation, the BADH activity in response to the nonmetabolizable inducer o-xylene was less than 2% of that of cells grown under conditions of succinate limitation. Less repression was found under conditions of ammonium or oxygen limitation (2 to 10% and 20 to 35%, respectively, of the BADH levels under succinate limitation). The BADH expression levels determined under the different growth conditions appeared to correlate well with the mRNA transcript levels from the upper pathway promoter (Pu), which indicates that repression was due to a blockage at the transcriptional level. The meta-cleavage pathway was found to be less susceptible to catabolite repression. The results obtained suggest that the occurrence of catabolite repression is related to a high-energy status of the cells rather than to a high growth rate or directly to the presence of growth-saturating concentrations of a primary carbon and energy source.

Alcohol Oxidoreductases↗
PubMed1996

Characterization of an OprL null mutant of Pseudomonas putida.

A Pseudomonas putida oprL null mutant was generated with reverse genetics by using an in vitro-truncated oprL::xylE construct and in vivo allelic exchange. The nature of the mutation introduced in P. putida was confirmed by Southern blotting. Western blots (immunoblots) of peptidoglycan-associated proteins revealed that the OprL protein was not made in the mutant strain, whereas it was detectable as a 19-kDa band in protein preparations of the wild-type strain. The P. putida oprL, mutant exhibited altered cell morphology as revealed by electron microscopy and was more sensitive to sodium dodecyl sulfate, deoxycholate, and EDTA than the wild-type strain. The oprL gene was conserved in a wide variety of the Pseudomonas strains belonging to rRNA group I, which suggests that this gene is important for the maintenance of the cell envelope and cell morphology in this group of microorganisms.

Bacterial Outer Membrane Proteins↗
PubMed1996

The TACAN4TGCA motif upstream from the -35 region in the sigma70-sigmaS-dependent Pm promoter of the TOL plasmid is the minimum DNA segment required for transcription stimulation by XylS regulators.

Transcription from the TOL plasmid meta-cleavage pathway operon promoter Pm is dependent on the XylS regulator activated by benzoate effectors or after XylS overproduction. We have generated 5' deletions in Pm and have analyzed expression from wild-type and mutant promoters with the wild-type XylS regulator and XylS mutant regulators that stimulated transcription constitutively. We have found that the motifs T(C or A)CAN4TGCA located between -46 and -57 and -67 and -78 with respect to the main transcription initiation point are required for maximal stimulation of transcription from Pm with effector-activated wild-type XylS. Deletion of the farthest TCCA submotif decreased but did not abolish transcription mediated by the pair XylS with 3-methylbenzoate; however, removal of the motif between -67 and -78 resulted in the loss of stimulation by the wild-type regulator. XylSG44S and XylSS229I stimulated high levels of transcription in the absence of effectors from the wild-type promoter and from a mutant promoter exhibiting only the -46 to -57 motif only when an effector was present. The point mutation Pm5U (with C-47 replaced by G [C-47-->G]) and Pm4 (C-68-->G), located in each 3' TGCA submotif of each motif, resulted in a 90% decrease in transcription stimulation with wild-type XylS; however, the mutant XylSS229I stimulated high levels of transcription from the point mutation promoters both in the presence and in the absence of effectors, while mutant XylSG44S suppressed the two point mutations only with 3-methylbenzoate. Overexpression of XylS and XylSG44S allowed the two regulators to stimulate high levels of transcription from the wild-type promoter, the point mutation Pm4 and Pm5U promoters, and deltaPm promoters exhibiting at least the -46 to -57 motif. Therefore the TACAN4TGCA motif between -46 and -57 represents the minimal DNA segment required for stimulation of transcription from Pm.

Bacterial Proteins↗
PubMed1996

The Pseudomonas putida peptidoglycan-associated outer membrane lipoprotein is involved in maintenance of the integrity of the cell cell envelope.

Pseudomonas putida 14G-3, a derivative of the natural soil inhabitant P. putida KT2440, exhibited a chromosomal insertion of a mini-Tn5/'phoA transposon that resulted in reduced ability to colonize soil. In vitro characterization of P. putida 14G-3 revealed that it exhibited an altered cell morphology and envelope, as revealed by electron microscopy. The derived strain was sensitive to sodium dodecyl sulfate, deoxycholate, and EDTA, produced clumps when it reached high cell densities in the late logarithmic growth phase, and did not grow on low-osmolarity medium. The P. putida DNA surrounding the mini-Tn5/'phoA insertion was cloned and used as a probe to rescue the wild-type gene, which was sequenced. Comparison of the deduced peptide sequence with sequences in the Swiss-Prot database allowed the knocked-out gene to be identified as that encoding the peptidoglycan-associated lipoprotein (Pal or OprL) of P. putida. The protein was identified in coupled transcription and translation assays in vitro.

Amino Acid Sequence↗
PubMed1996

Expression of the TOL plasmid xylS gene in Pseudomonas putida occurs from a alpha 70-dependent promoter or from alpha 70- and alpha 54-dependent tandem promoters according to the compound used for growth.

Growth of Pseudomonas putida (pWWO) on alkylbenzoates requires the expression of the meta pathway operon, which is mediated by the XylS protein after binding of a benzoate effector. Alternatively, in cells growing on toluene or its aromatic alcohols, overexpression of xylS mediated by XylR activated by these compounds leads to overproduction of the XylS regulator, which even in the absence of benzoate effectors stimulates transcription from the meta cleavage pathway operon promoter. We show here that in bacteria growing on glycerol or alkylbenzoates, the xylS gene is expressed at a low but constitutive level from a newly found sigma 70-dependent promoter called Ps2. The amount of XylS protein made from the transcript originated from Ps2 was sufficient to allow high levels of expression from the meta cleavage pathway operon promoter when the cells were grown in the presence of 3-methylbenzoate. The transcription initiation point of the transcript generated from Ps2 mapped 9 bp upstream from the proposed ATG of the xylS gene; this transcript contains the ribosome-binding site. The Ps2 promoter was located 110 bp downstream from a previously described sigma54-dependent promoter located upstream from the xylS open reading frame, now called Ps1. In cells growing on toluene or benzyl alcohols, the XylS regulator is overproduced as a consequence of increased expression of the gene through the effect of the two promoters working in tandem: the newly found sigma 70-dependent promoter, whose expression is XylR and toluene independent, and the sigma 54-dependent promoter, whose expression is dependent on XylR activated by its effectors. This expression pathway of the xylS gene explains why sigma 54-deficient P. putida bearing the wild-type TOL plasmid, or the wild-type P. putida strain bearing a TOL plasmid with a knocked-out xylR gene, can grow on alkylbenzoates. Until now this has been one of the unresolved paradoxes in the transcriptional control of the TOL meta cleavage pathway.

Bacterial Proteins↗
PubMed1996

Vitamin dependent erythrocyte enzymes in newborns in relation to gestational age and birth weight.

In order to search for any difference in riboflavin and pyridoxine nutrition between term appropriate for gestational age (TAGA), term small for gestational age (TSGA) and preterm appropriate for gestational age (PTAGA) newborns, cord blood from 23, 19 and 20 infants, respectively, were studied, and red cell glutathione reductase (riboflavin-dependent) and glutamic oxaloacetic transaminase (pyridoxine-dependent) activities were measured, as well as their respective activity coefficients (GRase AC and GOT AC). Red cell enzymes were assayed according to BEUTLER [4] in a Gilford recording spectrophotometer model 2451 at 37 degrees C. Values of 1.42, 1.39 and 1.32 of GRase AC and values of 1.17, 1.02 and 1.08 GOT AC, respectively were obtained. These data indicate that there is no significant difference between the three categories of newborns with regard to riboflavin and pyridoxine nutrition, and suggest that neither maturity nor adequacy of intrauterine growth were related to differences in the availability of these vitamins. All groups demonstrate a uniform mild deficiency of riboflavin and an adequate level of pyridoxine, probably the effect of maternal diet.

Aspartate Aminotransferases↗
PubMed1995

Single amino acids changes in the signal receptor domain of XylR resulted in mutants that stimulate transcription in the absence of effectors.

The XylR protein positively controls expression from the Pseudomonas putida TOL plasmid sigma 54-dependent "upper" pathway operon promoter (Pu) and the xylS gene promoter (Ps), in response to the presence of aromatic effectors. Two mutant XylR regulators able to stimulate transcription from Pu and Ps in the absence of effectors were isolated. These mutants exhibited single point mutations, namely Asp135-->Asn and Pro85-->Ser. Both mutations are located in the amino termini domain of XylR, which is thought to be responsible for interactions with effectors. The effector profile of XylRP85S was similar to that of wild-type XylR protein; however, XylRD135N exhibited an altered pattern of effector recognition: with m-nitrotoluene it stimulated transcription from the Pu promoter above the high basal level, whereas this nitroarene inhibited the wild-type regulator. Previous work (Delgado, A., and Ramos, J.L. (1994) J. Biol. Chem. 269, 8059-8062) showed that residue 172 was involved in effector interactions, as mutant XylRE172K also recognized m-nitrotoluene. However, double mutant XylR135N/E172K did not stimulate transcription in the absence of effector, but retained the ability to stimulate transcription with m-nitrotoluene. Transcription mediated by XylRD135N and XylRP85S from Pu::lacZ was analyzed in detail. Like the wild-type regulator, XylRD135N and XylRP85S required sigma 54 for full transcription activation, but in contrast with the wild-type regulator, XylRD135N, but not XylRP85S, stimulated transcription from Pu in the absence of the integration host factor protein. XylRD135N, also in contrast with XylR and XylRP85S, mediated transcription from a mutant Pu promoter that lacked one of the upstream regulator binding sites (delta UAS1), but not when both upstream regulator binding sites were deleted. The level of autoregulation of XylRD135N was at least 2-fold higher than that found with the wild-type XylR regulator and the mutant XylRP85S.

Bacterial Proteins↗
PubMed1995

Effect of amiodarone therapy on mortality in patients with left ventricular dysfunction and asymptomatic complex ventricular arrhythmias: Argentine Pilot Study of Sudden Death and Amiodarone (EPAMSA).

The efficiency of prophylactic antiarrhythmic treatment with amiodarone in reducing 1-year mortality in patients with reduced left ventricular ejection fraction ( < 35%) and asymptomatic ventricular arrhythmias (Lown classes 2 and 4) was investigated in a prospective, multicenter, randomized, controlled study. Among 127 patients who entered the study, 61 were assigned to no antiarrhythmic therapy (control group [CG] and 66 to amiodarone treatment (amiodarone group [AG]). Amiodarone was administered at a dosage of 800 mg/day for 2 weeks followed by 400 mg/day thereafter. A 12-month follow-up was completed for 106 patients (57 in the AG and 49 in the CG). Amiodarone reduced the overall mortality rate, which was 10.5% in the AG versus 28.6% in the CG (odds ratio [OR] 0.29; 95% confidence interval [CI] 0.10 to 0.84; log-rank test 0.02) and sudden death rate, which was 7.0% in the AG versus 20.4% in the CG (OR 0.29; 95% CI 0.08 to 1.00; log-rank test 0.04). Side effects were rare, and in only three patients did amiodarone treatment have to be discontinued.

Aged↗