RESUMEN
AIMS: Isooctane (2,2,4-trimethylpentane), a major component of gasoline formulations, is recalcitrant to biodegradation probably because of the quaternary carbon group it contains. Information on the biodegradability of this hydrocarbon is essential to evaluate its fate in the environment. For these reasons, the degradation kinetics and the catabolic pathway of isooctane were investigated in Mycobacterium austroafricanum IFP 2173, the only strain characterized to use it as sole carbon and energy source. METHODS AND RESULTS: The selected strain exhibited a rather moderate maximum growth rate (micromax = 0.053 h(-1)) but degraded isooctane up to 99% with a mineralization yield of 45%, indicating attack of the quaternary carbon group. The GC/MS identification of metabolites, 2,4,4-trimethylpentanoic and dimethylpropanoic (pivalic) acids, which transiently accumulated in the cultures indicated that degradation started from the isopropyl extremity of the molecule and subsequently proceeded by catabolism of the tert-butyl moiety. The degradation of putative metabolic intermediates was investigated. The initial isooctane oxidation system was tentatively characterized. CONCLUSIONS: The isooctane-degrading strain harboured two candidate systems for initial alkane oxidation. Although a cytochrome P450 was induced by isooctane degradation, the functional oxidation system was probably a nonheme alkane monooxygenase as indicated by PCR amplification and RT-PCR expression of an alkB gene. SIGNIFICANCE AND IMPACT OF THE STUDY: Isooctane is a recalcitrant branched alkane. A plausible pathway of its degradation by Myco. austroafricanum was put forward.
Asunto(s)
Mycobacterium/metabolismo , Octanos/metabolismo , Alcanos/metabolismo , Biodegradación Ambiental , Biomasa , Dióxido de Carbono/metabolismo , Medios de Cultivo , Citocromo P-450 CYP4A/análisis , Citocromo P-450 CYP4A/genética , Sistema Enzimático del Citocromo P-450/análisis , Exposición a Riesgos Ambientales , Cromatografía de Gases y Espectrometría de Masas/métodos , Gasolina , Genes Bacterianos/genética , Malonatos/metabolismo , Mycobacterium/crecimiento & desarrollo , Ácidos Pentanoicos/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodosRESUMEN
An activated carbon fibre supporting silver (ACF(Ag)) was tested for its antibacterial capacity against Escherichia coli (E. coli). Water that has passed through ACF(Ag) demonstrated strong bactericidal ability. This activity decreased over the time suggesting that generated bactericidal species were short lifespan. Since formation of reactive oxygen species (ROS) might be catalysed by silver impregnated and/or ACF itself, implication of ROS and silver was evaluated by the use of ROS scavengers and a silver ions neutralizing agent. The role of ROS in the E. coli mortality was confirmed by the use of a molecular approach which revealed a strong expression of oxidative stress genes.
Asunto(s)
Escherichia coli/metabolismo , Nanotubos de Carbono , Especies Reactivas de Oxígeno/metabolismo , Plata , Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica/genética , Nanotubos de Carbono/química , Estrés Oxidativo/genética , Plata/químicaRESUMEN
The biodegradability under aerobic conditions of volatile hydrocarbons (4-6 carbons) contained in gasoline and consisting of n-alkanes, iso-alkanes, cycloalkanes and alkenes, was investigated. Activated sludge was used as the reference microflora. The biodegradation test involved the degradation of the volatile fraction of gasoline in closed flasks under optimal conditions. The kinetics of biodegradation was monitored by CO2 production. Final degradation was determined by gas chromatographic analysis of all measurable hydrocarbons (12 compounds) in the mixture after sampling the headspace of the flasks. The degradation of individual hydrocarbons was also studied with the same methodology. When incubated individually, all hydrocarbons used as carbon sources, except 2,2-dimethylbutane and 2,3-dimethylbutane, were completely consumed in 30 days or less with different velocities and initial lag periods. When incubated together as constituents of the light gasoline fraction, all hydrocarbons were metabolised, often with higher velocities than for individual compounds. Cometabolism was involved in the degradation of dimethyl isoalkanes.
Asunto(s)
Gasolina , Hidrocarburos/metabolismo , Biodegradación Ambiental , Cromatografía de Gases , Medios de Cultivo , VolatilizaciónRESUMEN
A bacterial strain (strain IFP 2173) was selected from a gasoline-polluted aquifer on the basis of its capacity to use 2,2, 4-trimethylpentane (isooctane) as a sole carbon and energy source. This isolate, the first isolate with this capacity to be characterized, was identified by 16S ribosomal DNA analysis, and 100% sequence identity with a reference strain of Mycobacterium austroafricanum was found. Mycobacterium sp. strain IFP 2173 used an unusually wide spectrum of hydrocarbons as growth substrates, including n-alkanes and multimethyl-substituted isoalkanes with chains ranging from 5 to 16 carbon atoms long, as well as substituted monoaromatic hydrocarbons. It also attacked ethers, such as methyl t-butyl ether. During growth on gasoline, it degraded 86% of the substrate. Our results indicated that strain IFP 2173 was capable of degrading 3-methyl groups, possibly by a carboxylation and deacetylation mechanism. Evidence that it attacked the quaternary carbon atom structure by an as-yet-undefined mechanism during growth on 2,2,4-trimethylpentane and 2,2-dimethylpentane was also obtained.
Asunto(s)
Gasolina , Hidrocarburos/metabolismo , Mycobacterium/clasificación , Mycobacterium/metabolismo , Microbiología del Agua , Biodegradación Ambiental , Medios de Cultivo , Ciclohexanos/metabolismo , ADN Bacteriano/análisis , ADN Bacteriano/genética , ADN Ribosómico/análisis , ADN Ribosómico/genética , Cromatografía de Gases y Espectrometría de Masas , Hidrocarburos/química , Datos de Secuencia Molecular , Mycobacterium/crecimiento & desarrollo , Mycobacterium/aislamiento & purificación , Octanos/metabolismo , Filogenia , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Contaminantes del Agua/metabolismoRESUMEN
We have studied the meiotic segregation of a chromosome length polymorphism (CLP) in the yeast Saccharomyces cerevisiae. The neopolymorphism frequently observed within the smallest chromosomes (I, VI, III and IX) is not completely understood. We focused on the analysis of the structure of chromosome I in 88 segregants from a cross between YNN295 and FL100trp. Strain FL100trp is known to carry a reciprocal translocation between the left arm of chromosome III and the right arm of chromosome I. PCR and Southern hybridization analyses were performed and a method for the rapid detection of chromosome I rearrangements was developed. Seven chromosome I types were identified among the 88 segregants. We detected 22 recombination events between homologous chromosomes I and seven ectopic recombination events between FL100trp chromosome III and YNN295 chromosome I. These recombination events occurred in 20 of the 22 tetrads studied (91%). Nine tetrads (41%) showed two recombination events. This showed that homologous recombination involving polymorphic homologues or heterologous chromosomes is the main source of neopolymorphism. Only one of the seven chromosome I variants resulted from a transposition event rather than a recombination event. We demonstrated that a Tyl element had transposed within the translocated region of chromosome I, generating mutations in the 3' LTR, at the border between U5 and PBS.
Asunto(s)
Cromosomas Fúngicos , Polimorfismo Genético , Recombinación Genética , Saccharomyces cerevisiae/genética , Secuencias Repetidas Terminales/genética , Secuencia de Bases , Mapeo Cromosómico , Cruzamientos Genéticos , Cartilla de ADN , ADN de Hongos/genética , ADN de Hongos/aislamiento & purificación , Electroforesis en Gel de Campo Pulsado , Meiosis/genética , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Mapeo Restrictivo , Saccharomyces cerevisiae/citología , Alineación de Secuencia , Homología de Secuencia de Ácido NucleicoRESUMEN
A methodology was devised and was found useful for the selection of populations degrading recalcitrant hydrocarbons. The work was part of a programme aiming at developing knowledge of the intrinsic capacities of autochtonous microflorae of the environment for gasoline biodegradation. The methodology involved monitoring the progress of degradation in enrichment liquid cultures on the selected hydrocarbon by gas chromatographic analysis of CO2 production and O2 consumption. Populations degrading in particular o-xylene, 1,2,4-trimethylbenzene, cyclohexane were obtained. Concerning 2,2,4-trimethylpentane (isooctane), one microflora (and a pure strain derived from it) growing on this hydrocarbon were obtained from gasoline-polluted water.
Asunto(s)
Bacterias/aislamiento & purificación , Gasolina/microbiología , Hidrocarburos/metabolismo , Microbiología del Suelo , Microbiología del Agua , Bacterias/metabolismo , Derivados del Benceno/metabolismo , Biodegradación Ambiental , Reactores Biológicos/microbiología , Dióxido de Carbono/análisis , Cromatografía de Gases , Ciclohexanos/metabolismo , Octanos/metabolismo , Consumo de Oxígeno , Xilenos/metabolismoRESUMEN
A methodology allowing the detailed assessment of the capacities of microflorae to degrade gasoline in aerobic conditions has been developed. It consisted in the determination of the degradation of a gasoline model mixture in liquid cultures in optimal conditions. The gasoline model mixture contained 23 representative hydrocarbons of gasoline (GM23). The kinetics and extent of biodegradation were evaluated by continuous overall monitoring of CO2 production and final chromatographic analysis (usually after about 30 days) of the consumption of each hydrocarbon. The methodology was used with soil and water samples from polluted and non polluted sites. The experimentation aimed at assessing the distribution of the degradative capacities in the environment and the prospects for natural attenuation of gasoline. Nine microflorae were tested. The intrinsic biodegradability (existence of mechanisms of biodegradation) appeared total for GM23 as shown by the results obtained with several microflorae. The degradative capacities of microflorae from non polluted samples were high (total degradation rates at least 85%). Incomplete degradation was observed essentially for trimethylalkanes (2,2,4-trimethylpentane and 2,3,4-trimethylpentane) and for cyclohexane. In several cases, samples from polluted sites exhibited more extensive degradative capacities, with total degradation of all hydrocarbons being observed for three out of the six samples.
Asunto(s)
Gasolina , Administración de Residuos/métodos , Aerobiosis , Biodegradación Ambiental , Ciclohexanos/metabolismo , Microbiología del Suelo , Contaminantes del Suelo/metabolismo , Contaminantes Químicos del Agua/metabolismoRESUMEN
The degradation of gasoline by a microflora from an urban waste water activated sludge was investigated in detail. Degradation kinetics were studied in liquid cultures at 30 degrees C by determination of overall O2 consumption and CO2 production and by chromatographic analysis of all 83 identifiable compounds. In a first fast phase (2 d) of biodegradation, 74% of gasoline, involving mostly aromatic hydrocarbons, was consumed. A further 20%, involving other hydrocarbons, was consumed in a second slow phase (23 d). Undegraded compounds (6% of gasoline) were essentially some branched alkanes with a quaternary carbon or/and alkyl chains on consecutive carbons but cycloalkanes, alkenes and C10- and C11-alkylated benzenes were degraded. The degradation kinetics of individual hydrocarbons, determined in separate incubations, followed patterns similar to those observed in cultures on gasoline. Carbon balance experiments of gasoline degradation were performed. The carbon of degraded gasoline was mainly (61.7%) mineralized into CO2, the remaining carbon being essentially converted into biomass.
Asunto(s)
Bacterias/metabolismo , Gasolina , Hidrocarburos/metabolismo , Aguas del Alcantarillado/microbiología , Biodegradación Ambiental , Dióxido de Carbono/metabolismo , Medios de Cultivo , Hidrocarburos/clasificación , Cinética , Consumo de OxígenoRESUMEN
A methodology to determine the intrinsic capacities of a microflora to degrade gasoline was developed, in particular for assessing the potential of autochtonous populations of polluted and non polluted soils for natural attenuation and engineered bioremediation. A model mixture (GM23) constituted of the 23 most representative hydrocarbons of a commercial gasoline was used. The capacities of the microflorae (kinetics and extent of biodegradation) were assessed by chromatographic analysis of hydrocarbon consumption and of CO2 production. The degradation of the components of GM23 was assayed in separate incubations of each component and in the complete mixture. For the microflora of an unpolluted spruce forest soil, all hydrocarbons of GM23 except cyclohexane, 2,2,4- and 2,3,4-trimethylpentane isomers were degraded to below detection limit in 28 days. This microflora was reinforced with two mixed microbial communities selected from gasoline-polluted sites and shown to degrade cyclohexane and 2,2,4-trimethylpentane. With the reinforced microflora, complete degradation of GM23 was observed. The degradation patterns of individual components of GM23 were similar when the compounds were present individually or in the GM23 mixture, as long as the concentrations of 2-ethyltoluene and trimethylbenzene isomers were kept sufficiently low (< or = 35 mg.l-1) to remain below their inhibitory level.