Microbial communities in a porphyry copper tailings impoundment and their impact on the geochemical dynamics of the mine waste

The distribution and diversity of acidophilic bacteria of a tailings impoundment at the La Andina copper mine, Chile, was examined. The tailings have low sulfide (1.7% pyrite equivalent) and carbonate (1.4% calcite equivalent) contents and are stratified into three distinct zones: a surface (0-70-80 cm) `oxidation zone' characterized by low-pH (2.5-4), a `neutralization zone' (70-80 to 300-400 cm) and an unaltered `primary zone' below 400 cm. A combined cultivation-dependent and biomolecular approach (terminal restriction enzyme fragment length polymorphism and 16S rRNA clone library analysis) was used to characterize the indigenous prokaryotic communities in the mine tailings. Total cell counts showed that the microbial biomass was greatest in the top 125 cm of the tailings. The largest numbers of bacteria (10(9) g(-1) dry weight of tailings) were found at the oxidation front (the junction between the oxidation and neutralization zones), where sulfide minerals and oxygen were both present. The dominant iron-/sulfur-oxidizing bacteria identified at the oxidation front included bacteria of the genus Leptospirillum (detected by molecular methods), and Gram-positive iron-oxidizing acidophiles related to Sulfobacillus (identified both by molecular and cultivation methods). Acidithiobacillus ferrooxidans was also detected, albeit in relatively small numbers. Heterotrophic acidophiles related to Acidobacterium capsulatum were found by molecular methods, while another Acidobacterium-like bacterium and an Acidiphilium sp. were isolated from oxidation zone samples. A conceptual model was developed, based on microbiological and geochemical data derived from the tailings, to account for the biogeochemical evolution of the Piuquenes tailings impoundment.

Methadone enantiomer plasma levels, CYP2B6, CYP2C19, and CYP2C9 genotypes, and response to treatment.

BACKGROUND AND OBJECTIVE: Recent in vitro studies have suggested an important role of cytochrome P450 (CYP) 2B6 and CYP2C19 in methadone metabolism. We aimed to determine the influence of CYP2B6, CYP2C9, and CYP2C19 genetic polymorphism on methadone pharmacokinetics and on the response to treatment. METHODS: We included 209 patients in methadone maintenance treatment on the basis of their response to treatment and their daily methadone dose. Patients were genotyped for CYP2B6, CYP2C9, and CYP2C19. Steady-state trough and peak (R)-, (S)-, and (R,S)-plasma levels and peak-to-trough plasma level ratios were measured. RESULTS: CYP2B6 genotype influences (S)-methadone and, to a lesser extent, (R)-methadone plasma levels, with the median trough (S)-methadone plasma levels being 105, 122, and 209 ng . kg/mL . mg for the noncarriers of allele *6, heterozygous carriers, and homozygous carriers (*6/*6), respectively (P = .0004). CYP2C9 and CYP2C19 genotypes do not influence methadone plasma levels. Lower peak and trough plasma levels of methadone and higher peak-to-trough ratios were measured in patients considered as nonresponders [median (R,S)-methadone trough plasma levels of 183 and 249 ng . kg/mL . mg (P = .0004) and median peak-to-trough ratios of 1.82 and 1.58 for high-dose nonresponders and high-dose responders, respectively (P = .0003)]. CONCLUSION: Although CYP2B6 influences (S)-methadone plasma levels, given that only (R)-methadone contributes to the opioid effect of this drug, a major influence of CYP2B6 genotype on response to treatment is unlikely and has not been shown in this study. Lower plasma levels of methadone in nonresponders, suggesting a higher clearance, and higher peak-to-trough ratios, suggesting a shorter elimination half-life, are in agreement with the usual clinical measures taken for such patients, which are to increase methadone dosages and to split the daily dose into several intakes.

Maintenance of polymorphism : a theoretical approach using multilocus systems

Résumé La thématique de cette thèse peut être résumée par le célèbre paradoxe de biologie évolutive sur le maintien du polymorphisme face à la sélection et par l'équation du changement de fréquence gamétique au cours du temps dû, à la sélection. La fréquence d'un gamète xi à la génération (t + 1) est: !!!Equation tronquée!!! Cette équation est utilisée pour générer des données utlisée tout au long de ce travail pour 2, 3 et 4 locus dialléliques. Le potentiel de l'avantage de l'hétérozygote pour le maintien du polymorphisme est le sujet de la première partie. La définition commune de l'avantage de l'hétérozygote n'etant applicable qu'a un locus ayant 2 allèles, cet avantage est redéfini pour un système multilocus sur les bases de précédentes études. En utilisant 5 définitions différentes de l'avantage de l'hétérozygote, je montre que cet avantage ne peut être un mécanisme général dans le maintien du polymorphisme sous sélection. L'étude de l'influence de locus non-détectés sur les processus évolutifs, seconde partie de cette thèse, est motivée par les travaux moléculaires ayant pour but de découvrir le nombre de locus codant pour un trait. La plupart de ces études sous-estiment le nombre de locus. Je montre que des locus non-détectés augmentent la probabilité d'observer du polymorphisme sous sélection. De plus, les conclusions sur les facteurs de maintien du polymorphisme peuvent être trompeuses si tous les locus ne sont pas détectés. Dans la troisième partie, je m'intéresse à la valeur attendue de variance additive après un goulot d'étranglement pour des traits sélectionés. Une études précédente montre que le niveau de variance additive après goulot d'étranglement augmente avec le nombre de loci. Je montre que le niveau de variance additive après un goulot d'étranglement augmente (comparé à des traits neutres), mais indépendamment du nombre de loci. Par contre, le taux de recombinaison a une forte influence, entre autre en regénérant les gamètes disparus suite au goulot d'étranglement. La dernière partie de ce travail de thèse décrit un programme pour le logiciel de statistique R. Ce programme permet d'itérer l'équation ci-dessus en variant les paramètres de sélection, recombinaison et de taille de populations pour 2, 3 et 4 locus dialléliques. Cette thèse montre qu'utiliser un système multilocus permet d'obtenir des résultats non-conformes à ceux issus de systèmes rnonolocus (la référence en génétique des populations). Ce programme ouvre donc d'intéressantes perspectives en génétique des populations. Abstract The subject of this PhD thesis can be summarized by one famous paradox of evolu-tionary biology: the maintenance of polymorphism in the face of selection, and one classical equation of theoretical population genetics: the changes in gametic frequencies due to selection and recombination. The frequency of gamete xi at generation (t + 1) is given by: !!! Truncated equation!!! This equation is used to generate data on selection at two, three, and four diallelic loci for the different parts of this work. The first part focuses on the potential of heterozygote advantage to maintain genetic polymorphism. Results of previous studies are used to (re)define heterozygote advantage for multilocus systems, since the classical definition is for one diallelic locus. I use 5 different definitions of heterozygote advantage. And for these five definitions, I show that heterozygote advantage is not a general mechanism for the maintenance of polymorphism. The study of the influence of undetected loci on evolutionary processes (second part of this work) is motivated by molecular works which aim at discovering the loci coding for a trait. For most of these works, some coding loci remains undetected. I show that undetected loci increases the probability of maintaining polymorphism under selection. In addition, conclusions about the factor that maintain polymorphism can be misleading if not all loci are considered. This is, therefore, only when all loci are detected that exact conclusions on the level of maintained polymorphism or on the factor(s) that maintain(s) polymorphism could be drawn. In the third part, the focus is on the expected release of additive genetic variance after bottleneck for selected traits. A previous study shows that the expected release of additive variance increases with an increase in the number of loci. I show that the expected release of additive variance after bottleneck increases for selected traits (compared with neutral), but this increase is not a function of the number of loci, but function of the recombination rate. Finally, the last part of this PhD thesis is a description of a package for the statistical software R that implements the Equation given above. It allows to generate data for different scenario regarding selection, recombination, and population size. This package opens perspectives for the theoretical population genetics that mainly focuses on one locus, while this work shows that increasing the number of loci leads not necessarily to straightforward results.

Enzyme activities and pectin breakdown of sapodilla submitted to 1-methylcyclopropene

The objective of this work was to investigate the influence of 1-methylcyclopropene (1-MCP) at 300 nL L-1 on activities of cell wall hidrolytic enzymes and pectin breakdown changes which Sapodilla (Manilkara zapota cv. Itapirema 31) cell wall undergoes during ripening. Sapodilla were treated with ethylene antagonist 1-MCP at 300 nL L-1 for 12 hours and then, stored under a modified atmosphere at 25º C for 23 days. Firmness, total and soluble pectin and cell wall enzymes were monitored during storage. 1-MCP at 300 nL L-1 for 12 hours delayed significantly softening of sapodilla for 11 days at 25º C. 1-MCP postharvest treatment affected the activities of cell wall degrading enzymes pectinmethylesterase and polygalacturonase and completely suppressed increases in beta-galactosidase for 8 days, resulting in less pectin solubilization. Beta-galactosidase seems relevant to softening of sapodilla and is probably responsible for modification of both pectin and xyloglucan-cellulose microfibril network.

The deubiquitinating enzyme AMSH3 is required for intracellular trafficking and vacuole biogenesis in Arabidopsis thaliana.

Ubiquitination, deubiquitination, and the formation of specific ubiquitin chain topologies have been implicated in various cellular processes. Little is known, however, about the role of ubiquitin in the development of cellular organelles. Here, we identify and characterize the deubiquitinating enzyme AMSH3 from Arabidopsis thaliana. AMSH3 hydrolyzes K48- and K63-linked ubiquitin chains in vitro and accumulates both ubiquitin chain types in vivo. amsh3 mutants fail to form a central lytic vacuole, accumulate autophagosomes, and mis-sort vacuolar protein cargo to the intercellular space. Furthermore, AMSH3 is required for efficient endocytosis of the styryl dye FM4-64 and the auxin efflux facilitator PIN2. We thus present evidence for a role of deubiquitination in intracellular trafficking and vacuole biogenesis.

Alternative genotyping method for the single nucleotide polymorphism A2959G (AF159246) of the bovine CAST gene

The objective of this work was to genotype the single nucleotide polymorphism (SNP) A2959G (AF159246) of bovine CAST gene by PCR-RFLP technique, and to report its use for the first time. For this, 147 Bos indicus and Bos taurus x Bos indicus animals were genotyped. The accuracy of the method was confirmed through the direct sequencing of PCR products of nine individuals. The lowest frequency of the meat tenderness favorable allele (A) in Bos indicus was confirmed. The use of PCR-RFLP for the genotyping of the bovine CAST gene SNP was shown to be robust and inexpensive, which will greatly facilitate its analysis by laboratories with basic structure.

The T309G Murine Double Minute 2 Gene Polymorphism Is an Independent Prognostic Factor for Patients with Renal Cell Carcinoma.

The purpose of this study was to evaluate the association of the T309G MDM2 gene polymorphism with renal cell carcinoma (RCC) risk, pathology, and cancer-specific survival (CSS). T309G MDM2 was genotyped in 449 Caucasians, including 240 with RCC and 209 cancer-free controls. The T309G MDM2 genotype was TT in 174 (38.8%), GT in 214 (47.7%), and GG in 61 (13.6%) subjects, without any significant differences between cases and controls on both univariable (p=0.58) and multivariable logistic regression (each p>0.25). Furthermore, T309G MDM2 was not linked with T stage (p=0.75), N stage (p=0.37), M stage (p=0.94), grade (p=0.21), and subtype (p=0.55). There was, however, a statistically significant association of T309G MDM2 with CSS (p=0.022): patients with TT had significantly worse survival than GG/GT (p=0.009), while those with GT and GG had similar outcomes (p=0.92). The 5-year survival rate for patients with TT, GT, and GG was 69.5%, 84.5%, and 89.7%, respectively. On the multivariable analysis, T309G was identified as an independent prognostic factor. The T309G MDM2 polymorphism is an independent prognostic factor for patients with RCC, with the TT genotype being associated with worse prognosis. In this study, there were no significant associations with RCC risk and pathology.

Hypertension. Angio-oedème sous inhibition de l'enzyme de conversion de l'angiotensine et de la dipeptidyl-peptidase-4 [Angioedema during ACE and DPP-4 inhibition]

Angioedema is a rare side effect of angiotensin converting enzyme (ACE) inhibitors. Its cause is probably related to the accumulation of bradykinin and substance P, i.e. two proinflammatory peptides normally inactivated by ACE. Angioedema occurs most of the time at the early phase of treatment, but may also develop during long-term treatment. It might involve the gastro-intestinal tract, leading to abdominal pain, vomiting and/or diarrhea, as well as pancreatitis. Dipeptidyl-ptidase-4 (DPP-4) is another enzyme allowing the degradation of bradykinin and substance P. Co-administering an ACE inhibitor and a DPP-4 inhibitor (as an antidiabetic agent) increases significantly the risk of angioedema.

Mutations in B3GALT6, which encodes a glycosaminoglycan linker region enzyme, cause a spectrum of skeletal and connective tissue disorders.

Proteoglycans (PGs) are a major component of the extracellular matrix in many tissues and function as structural and regulatory molecules. PGs are composed of core proteins and glycosaminoglycan (GAG) side chains. The biosynthesis of GAGs starts with the linker region that consists of four sugar residues and is followed by repeating disaccharide units. By exome sequencing, we found that B3GALT6 encoding an enzyme involved in the biosynthesis of the GAG linker region is responsible for a severe skeletal dysplasia, spondyloepimetaphyseal dysplasia with joint laxity type 1 (SEMD-JL1). B3GALT6 loss-of-function mutations were found in individuals with SEMD-JL1 from seven families. In a subsequent candidate gene study based on the phenotypic similarity, we found that B3GALT6 is also responsible for a connective tissue disease, Ehlers-Danlos syndrome (progeroid form). Recessive loss-of-function mutations in B3GALT6 result in a spectrum of disorders affecting a broad range of skeletal and connective tissues characterized by lax skin, muscle hypotonia, joint dislocation, and spinal deformity. The pleiotropic phenotypes of the disorders indicate that B3GALT6 plays a critical role in a wide range of biological processes in various tissues, including skin, bone, cartilage, tendon, and ligament.

Lack of angiotensin I accumulation after converting enzyme blockade by enalapril or lisinopril in man.

In nine normal volunteers, a series of five venous blood samples was obtained before and up to 24 h after converting enzyme inhibition by a single oral dose of enalapril or lisinopril. Plasma renin activity and blood angiotensin I were measured. A close linear relationship was found between the increase in plasma renin activity and the increase in blood angiotensin I. The linear correlation between plasma renin activity and blood angiotensin I remained after converting enzyme inhibition. Thus, the rise in angiotensin I after inhibition of the conversion of angiotensin I to angiotensin II is due to an enhanced release of renin rather than to accumulation of angiotensin I.

Genome-wide DNA polymorphism analyses using VariScan

BACKGROUND: DNA sequence polymorphisms analysis can provide valuable information on the evolutionary forces shaping nucleotide variation, and provides an insight into the functional significance of genomic regions. The recent ongoing genome projects will radically improve our capabilities to detect specific genomic regions shaped by natural selection. Current available methods and software, however, are unsatisfactory for such genome-wide analysis. RESULTS: We have developed methods for the analysis of DNA sequence polymorphisms at the genome-wide scale. These methods, which have been tested on a coalescent-simulated and actual data files from mouse and human, have been implemented in the VariScan software package version 2.0. Additionally, we have also incorporated a graphical-user interface. The main features of this software are: i) exhaustive population-genetic analyses including those based on the coalescent theory; ii) analysis adapted to the shallow data generated by the high-throughput genome projects; iii) use of genome annotations to conduct a comprehensive analyses separately for different functional regions; iv) identification of relevant genomic regions by the sliding-window and wavelet-multiresolution approaches; v) visualization of the results integrated with current genome annotations in commonly available genome browsers. CONCLUSION: VariScan is a powerful and flexible suite of software for the analysis of DNA polymorphisms. The current version implements new algorithms, methods, and capabilities, providing an important tool for an exhaustive exploratory analysis of genome-wide DNA polymorphism data.

DnaSP v5: A software for comprehensive analysis of DNA polymorphism data

DnaSP is a software package for a comprehensive analysis of DNA polymorphism data. Version 5 implements a number of new features and analytical methods allowing extensive DNA polymorphism analyses on large datasets. Among other features, the newly implemented methods allow for: (i) analyses on multiple data files; (ii) haplotype phasing; (iii) analyses on insertion/deletion polymorphism data; (iv) visualizing sliding window results integrated with available genome annotations in the UCSC browser.