The surface free energy of Leishmania mexicana amazonensis.

Surface charge of Leishmania mexicana amazonensis was investigated by direct zeta-potential determination and ultrastructural cytochemistry, and its surface tension was studied by measurements of the advancing contact angle formed by the parasite monolayers with drops of liquids of different polarities. Both virulent and avirulent promastigotes exhibited negatively charged surfaces with a zeta-potential of about -15 mV. Treatment of these cells with trypsin, alkaline phosphatase, or phospholipase C rendered their surfaces less negatively charged, whereas neuraminidase did not alter the parasite negativeness. Cytochemically, we could observe a reduction in the cationized ferritin binding after the parasite treatment with each of the former enzymes, but not with neuraminidase. The surface free energy of parasites was calculated by taken to account the London dispersion, the Keeson dipole-dipole, and the Debye dipole-induced forces, as well as the surface polarity of the parasites and their zeta-potentials, by considering their adhesion to polystyrene surfaces. The delta G values of -6.4 and -18.1 mJ.m-2 were obtained for avirulent and virulent promstigotes, respectively.     

Geographic differences in the allele frequencies of the human Y-linked tetranucleotide polymorphism DYS19

We have studied the allele frequency distribution of the microsatellite locus DYS 19 in several populations with different geographical origins worldwide. Three new alleles were found. In addition, remarkable geographic and ethnic differences were observed in the allele frequency profiles and DNA marker (gene) diversity among populations and major ethnic groups. Amerindians showed an overwhelming predominance of the A allele, while in Caucasians the B allele was modal, and in Greater Asians and Africans allele C became predominant. Even within these geographic regions there were significant gradients, as exemplified by the decreasing frequency profile of the B allele from Great Britain over Germany to Slovakia. Thus, DYS 19 emerges as a useful tool for studying the structure and dynamics of human populations.     

The magic-bead concept: an integrated approach to nitrogen removal with co-immobilized micro-organisms

This paper describes both qualitative and quantitative aspects of simultaneous autotrophic nitrification and heterotrophic denitrification by, respectively, the nitrifierNitrisomonas europaea and either of the denitrifiersPseudomonas denitrificans orParacoccus denitrificans co-immobilized in double-layer gel beads. The system is based on the establishment of well-defined oxic and anoxic zones within the cell supports and on physical separation of the nitrifying and denitrifying populations. Nitrification and denitrification rates were obtained from measured bulk concentrations and head-space analysis. The latter analyses showed that ammonia was primarily converted into molecular nitrogen. Nitrous oxide was not detected. High nitrogen removal rates (up to 5.1 mmol N m^–3 gel s^–1) were achieved in continuous reactors under aerobic conditions. The overall rate of nitrogen removal was controlled by the nitrifying step. The approach followed is, in principle, also suitable to the coupling of other oxidative and reductive bioprocesses having complementary metabolic routes. Two-stage bioconversion processes can be thus conducted as if single-staged, which results in more compact reactor systems.     

Transfer RNA structural change is a key element in the reassignment of the CUG codon in Candida albicans.

The human pathogenic yeast Candida albicans and a number of other Candida species translate the standard leucine CUG codon as serine. This is the latest addition to an increasing number of alterations to the standard genetic code which invalidate the theory that the code is frozen and universal. The unexpected finding that some organisms evolved alternative genetic codes raises two important questions: how have these alternative codes evolved and what evolutionary advantages could they create to allow for their selection? To address these questions in the context of serine CUG translation in C.albicans, we have searched for unique structural features in seryl-tRNA(CAG), which translates the leucine CUG codon as serine, and attempted to reconstruct the early stages of this genetic code switch in the closely related yeast species Saccharomyces cerevisiae. We show that a purine at position 33 (G33) in the C.albicans Ser-tRNA(CAG) anticodon loop, which replaces a conserved pyrimidine found in all other tRNAs, is a key structural element in the reassignment of the CUG codon from leucine to serine in that it decreases the decoding efficiency of the tRNA, thereby allowing cells to survive low level serine CUG translation. Expression of this tRNA in S.cerevisiae induces the stress response which allows cells to acquire thermotolerance. We argue that acquisition of thermotolerance may represent a positive selection for this genetic code change by allowing yeasts to adapt to sudden changes in environmental conditions and therefore colonize new ecological niches.     

Enrichment of mixed cultures capable of aerobic degradation of 1,2-dibromoethane.

1,2-dibromoethane (DBE) is a common environmental contaminant; it is potentially carcinogenic and has been detected in soil and groundwater supplies. Most of the biodegradation studies to date have been performed under anaerobic conditions or in the context of soil remediation, where the pollutant concentration was in the parts per billion range. In this work a mixed bacterial culture capable of complete aerobic mineralization of concentrations of DBE up to 1 g liter(-1) under well-controlled laboratory conditions was enriched. In order to verify biodegradation, formation of biodegradation products as well as the disappearance of DBE from the biological medium were measured. Complete mineralization was verified by measuring stoichiometric release of the biodegradation products. This mixed culture was found to be capable of degrading other halogenated compounds, including bromoethanol, the degradation of which has not been reported previously.     

Uncoupling effect of nitrite during denitrification by Pseudomonas fluorescens: An in vivo (31)P-NMR study

In vivo (31)P-NMR was used to investigate the basis for the inhibition of denitrification by nitrite accumulated endogenously by Pseudomonas fluorescens ATCC 17822 (biotype II) at pH 7.0. Cells were immobilized in kappa-carrageenan to obtain high cell concentrations in the NMR tube. Acetate and nitrate in two concentration ratios were supplied as electron donor and acceptor, respectively, to achieve different levels of nitrite accumulation. During denitrification, cells were able to maintain a pH gradient of approximately 0.4 to 0.5 units, but when nitrite accumulation reached values approximating 27 mM the transmembrane DeltapH collapsed sharply. Nitrite stimulated the reduction rate of nitrate; furthermore, at nitrite concentrations below 1 mM, activation of oxygen respiratory rates was observed in cells grown under aerobic conditions. The results provide evidence for nitrite acting as a protonophore (an uncoupler that increases the proton permeability of membranes by a shuttling mechanism). (c) 1996 John Wiley & Sons, Inc.     

Prediction of total fecal output in sheep fed silage using the Captec chrome controlled-release capsule

Six Finnish wethers (average weight, 52.3 kg) were used to determine the effectiveness of a controlled-release Cr₂O₃ bolus and the internal marker acid insoluble ash (AIA) for predicting fecal output. Wethers were fed grass silage for ad libitum intake. Each wether was dosed orally with one continuous-release Cr₂O₃ bolus (CRC) on the first day of the experiment. Chronology of the trial was as follows: Day 0 to 14, marker adaptation period; Day 15 to 21, total fecal collection; Day 22 to 23, fecal grab sampling every 4 h; Day 25 to 35, once-daily fecal grab sampling. Accuracy of marker-estimated fecal output derived from each method was compared within a marker with total fecal collection values using a paired t-test. Concentrations of AIA and Cr varied (P < 0.10) between sampling times. Accuracy of fecal output estimates was better with the controlled-release Cr₂O₃ bolus (r = 0.97; P = 0.03) than with AIA (r = −0.08; P = 0.89). Grab samples taken once produced reliable (r = 0.96; P = 0.02) estimates of fecal output of silage-fed wethers using the controlled-release Cr₂O₃ bolus. The controlled-release Cr₂O₃ bolus is a better marker than acid-insoluble ash for predicting fecal output of silage-fed wethers.     

Enzymatic hydrolysis of wood: an engineering assessment

Eucalyptus wood samples were delignified with HCl-catalysed acetic acid solutions under selected experimental conditions and treated with NaClO solutions. The solid residues obtained were employed as substrates for enzymatic hydrolysis. The NaClO concentration used in the pretreatment step, the liquor/solid ratio and the enzyme/substrate ratio were considered as operational variables. The experimental data allowed the development of generalized kinetic models which provided the necessary information for design calculations. The operational conditions were compared from an engineering viewpoint on the basis of economic estimates. Optimum conditions were established from these estimates.     

Detachment of transformed cells

A parallel-plate flow chamber was used to quantify the detachment of normal cloned rat embryo fibroblasts (CREF) fibroblasts,ras-transformed CREF fibroblasts (CREF T24), and CREF T24 fibroblasts transfected with a Krev/RAP1A suppressor gene (HK B1) from a confluent monolayer of normal CREF fibroblasts to determine if the expression patterns of CD44 variants (mol wt 110 and 140 kDa) corresponded with detachment properties and metastatic potential. In the detachment assay, known shear stresses ranging from 20–24 dyn/cm² were applied to the adherent cells and the number of cells detached from the monolayer after 180 s was determined. Results showed that cellular expression of CD44 variants correlated with the metastatic potential of the cells and with the cells’ ability to detach from a monolayer of normal cells. Western blot analysis showed a low level of expression of the CD44 variants in the normal cell line, CREF, and the lowly metastatic cell line, HK B1. Detachment studies showed a low percentage of detachment of both of these cell lines from a normal cell monolayer. Tumor-derived (HK B1-T) and lung nodule-derived (HK B1-M) cell lines were established and both formed tumors and metastasis with reduced latency periods as compared to HK B1, but still showed a markedly delayed latency period compared to the highly metastatic cell line, CREF T24. Both of these cell lines showed a higher expression of the CD44 variants as compared to CREF and HK B1, and detached easier than CREF and HK B1. CREF T24 showed a much higher level of expression of the variants and had a higher percentage detachment than all other cell lines. To further test the role of the CD44 variants in the ability of the cells to detach from the normal monolayer, CREF cells were transfected with a DNA construct that constitutively expresses the CD44 variants and the detachment properties of three randomly selected clones were studied. Clones 2 and 3 showed a low level of expression of the CD44 variants after transfection and detached from the normal monolayer similar to CREF. Clone 1 showed a high level of expression of the CD44 variants and the detachment of these cells was significantly higher than CREF. From these results, it is concluded that in the five cell lines studied, expression of the CD44 variants play a significant role in the ability of the cells to detach from a monolayer of normal cells. It is hypothesized that this detachment may be an important component of a cell’s ability to metastasize.