RESUMO
Polar biotransformation products have been identified as causative agents for the eventual increase in genotoxicity observed after the bioremediation of PAH-contaminated soils. Their further biodegradation has been described under certain biostimulation conditions; however, the underlying microorganisms and mechanisms remain to be elucidated. 9,10-Anthraquinone (ANTQ), a transformation product from anthracene (ANT), is the most commonly detected oxygenated PAH (oxy-PAH) in contaminated soils. Sand-in-liquid microcosms inoculated with creosote-contaminated soil revealed the existence of a specialized ANTQ degrading community, and Sphingobium sp. AntQ-1 was isolated for its ability to grow on this oxy-PAH. Combining the metabolomic, genomic, and transcriptomic analyses of strain AntQ-1, we comprehensively reconstructed the ANTQ biodegradation pathway. Novel mechanisms for polyaromatic compound degradation were revealed, involving the cleavage of the central ring catalyzed by Baeyer-Villiger monooxygenases (BVMO). Abundance of strain AntQ-1 16S rRNA and its BVMO genes in the sand-in-liquid microcosms correlated with maximum ANTQ biodegradation rates, supporting the environmental relevance of this mechanism. Our results demonstrate the existence of highly specialized microbial communities in contaminated soils responsible for processing oxy-PAHs accumulated by primary degraders. Also, they underscore the key role that BVMO may play as a detoxification mechanism to mitigate the risk posed by oxy-PAH formation during bioremediation of PAH-contaminated soils.
Assuntos
Hidrocarbonetos Policíclicos Aromáticos , Poluentes do Solo , Areia , RNA Ribossômico 16S/genética , Multiômica , Biodegradação Ambiental , Microbiologia do Solo , SoloRESUMO
This work examines the role of mycelia in enhancing the degradation by attached bacteria of organic pollutants that have poor bioavailability. Two oomycetes, Pythium oligandrum and Pythium aphanidermatum, were selected as producers of mycelial networks, while Mycobacterium gilvum VM552 served as a model polycyclic aromatic hydrocarbon (PAH) degrading bacterium. The experiments consisted of bacterial cultures exposed to a nondisturbed nonaqueous phase liquid (NAPL) layer containing a heavy fuel spiked with 14C-labeled phenanthrene that were incubated in the presence or absence of the mycelia of the oomycetes in both shaking and static conditions. At the end of the incubation, the changes in the total alkane and PAH contents in the NAPL residue were quantified. The results revealed that with shaking and the absence of mycelia, the strain VM552 grew by utilizing the bulk of alkanes and PAHs in the fuel; however, biofilm formation was incipient and phenanthrene was mineralized following zero-order kinetics, due to bioavailability limitations. The addition of mycelia favored biofilm formation and dramatically enhanced the mineralization of phenanthrene, up to 30 times greater than the rate without mycelia, possibly by providing a physical support to bacterial colonization and by supplying nutrients at the NAPL/water interface. The results in the static condition were very different because the bacterial strain alone degraded phenanthrene with sigmoidal kinetics but could not degrade alkanes or the bulk of PAHs. We suggest that bacteria/oomycete interactions should be considered not only in the design of new inoculants in bioremediation but also in biodegradation assessments of chemicals present in natural environments.
Assuntos
Biodegradação Ambiental , Micélio , Hidrocarbonetos Policíclicos Aromáticos , Bactérias , Disponibilidade Biológica , Poluentes do SoloRESUMO
A novel biphasic system containing mineral medium and sand coated with a biologically weathered creosote-PAH mixture was developed to specifically enrich the high molecular weight polycyclic aromatic hydrocarbon (HMW PAH)-degrading community from a creosote-polluted soil. This consortium (UBHP) removed 70% of the total HMW PAHs and their alkyl-derivatives in 12 weeks. Based on a combined culture-dependent/independent approach, including clone library analysis, detection of catabolic genes, metabolomic profiles, and characterization of bacterial isolates, 10 phylotypes corresponding to five major genera (Sphingobium, Sphingomonas, Achromobacter, Pseudomonas, and Mycobacterium) were pointed out as key players within the community. In response to exposure to different single PAHs, members of sphingomonads were associated to the utilization of phenanthrene, fluoranthene, benzo[a]anthracene, and chrysene, while the degradation of pyrene was mainly associated to low-abundance mycobacteria. In addition to them, a number of uncultured phylotypes were detected, being of special relevance a group of Gammaproteobacteria closely related to a group previously associated with pyrene degradation that were here related to benzo(a)anthracene degradation. The overall environmental relevance of these phylotypes was confirmed by pyrosequencing analysis of the microbial community shift in the creosote-polluted soil during a lab-scale biostimulation.
Assuntos
Achromobacter/metabolismo , Gammaproteobacteria/metabolismo , Mycobacterium/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Pseudomonas/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Sphingomonadaceae/metabolismo , Achromobacter/classificação , Achromobacter/genética , Achromobacter/isolamento & purificação , Antracenos/metabolismo , Biodegradação Ambiental , Gammaproteobacteria/classificação , Gammaproteobacteria/genética , Gammaproteobacteria/isolamento & purificação , Biblioteca Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Consórcios Microbianos/genética , Mycobacterium/classificação , Mycobacterium/genética , Mycobacterium/isolamento & purificação , Fenantrenos/metabolismo , Filogenia , Pseudomonas/classificação , Pseudomonas/genética , Pseudomonas/isolamento & purificação , Pirenos/metabolismo , Solo/química , Espanha , Sphingomonadaceae/classificação , Sphingomonadaceae/genética , Sphingomonadaceae/isolamento & purificaçãoRESUMO
Marine microbial consortium UBF, enriched from a beach polluted by the Prestige oil spill and highly efficient in degrading this heavy fuel, was subcultured in pyrene minimal medium. The pyrene-degrading subpopulation (UBF-Py) mineralized 31 % of pyrene without accumulation of partially oxidized intermediates indicating the cooperation of different microbial components in substrate mineralization. The microbial community composition was characterized by culture dependent and PCR based methods (PCR-DGGE and clone libraries). Molecular analyses showed a highly stable community composed by Alphaproteobacteria (84 %, Breoghania, Thalassospira, Paracoccus, and Martelella) and Actinobacteria (16 %, Gordonia). The members of Thalasosspira and Gordonia were not recovered as pure cultures, but five additional strains, not detected in the molecular analysis, that classified within the genera Novosphingobium, Sphingopyxis, Aurantimonas (Alphaproteobacteria), Alcanivorax (Gammaproteobacteria) and Micrococcus (Actinobacteria), were isolated. None of the isolates degraded pyrene or other PAHs in pure culture. PCR amplification of Gram-positive and Gram-negative dioxygenase genes did not produce results with any of the cultured strains. However, sequences related to the NidA3 pyrene dioxygenase present in mycobacterial strains were detected in UBF-Py consortium, suggesting the representative of Gordonia as the key pyrene degrader, which is consistent with a preeminent role of actinobacteria in pyrene removal in coastal environments affected by marine oil spills.
Assuntos
Dioxigenases/genética , Genes Bacterianos , Consórcios Microbianos/genética , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Pirenos/metabolismo , Água do Mar/microbiologia , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/metabolismo , Biodegradação Ambiental , Eletroforese em Gel de Gradiente Desnaturante , Processos Heterotróficos , Hidroxilação , Dados de Sequência Molecular , Fenantrenos/metabolismo , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
Polycyclic aromatic hydrocarbons (PAHs) are major environmental pollutants in a number of point source contaminated sites, where they are found embedded in complex mixtures containing different polyaromatic compounds. The application of bioremediation technologies is often constrained by unpredictable end-point concentrations enriched in recalcitrant high molecular weight (HMW)-PAHs. The aim of this study was to elucidate the microbial populations and potential interactions involved in the biodegradation of benz(a)anthracene (BaA) in PAH-contaminated soils. The combination of DNA stable isotope probing (DNA-SIP) and shotgun metagenomics of 13C-labeled DNA identified a member of the recently described genus Immundisolibacter as the key BaA-degrading population. Analysis of the corresponding metagenome assembled genome (MAG) revealed a highly conserved and unique genetic organization in this genus, including novel aromatic ring-hydroxylating dioxygenases (RHD). The influence of other HMW-PAHs on BaA degradation was ascertained in soil microcosms spiked with BaA and fluoranthene (FT), pyrene (PY) or chrysene (CHY) in binary mixtures. The co-occurrence of PAHs resulted in a significant delay in the removal of PAHs that were more resistant to biodegradation, and this delay was associated with relevant microbial interactions. Members of Immundisolibacter, associated with the biodegradation of BaA and CHY, were outcompeted by Sphingobium and Mycobacterium, triggered by the presence of FT and PY, respectively. Our findings highlight that interacting microbial populations modulate the fate of PAHs during the biodegradation of contaminant mixtures in soils.
Assuntos
Hidrocarbonetos Policíclicos Aromáticos , Poluentes do Solo , Sphingomonadaceae , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Peso Molecular , Biodegradação Ambiental , Sphingomonadaceae/genética , Sphingomonadaceae/metabolismo , Benzo(a)Antracenos/metabolismo , Solo , Poluentes do Solo/metabolismo , Microbiologia do SoloRESUMO
Cyanobacteria are capable of transforming CO2 into polyhydroxybutyrate (PHB). In this study, different inorganic carbon concentrations (0-2 gC L-1) were evaluated for a Synechocystis sp. strain isolated from wastewater. Quantitative RT-qPCR was also performed to decipher the links between inorganic carbon and PHB and glycogen metabolism. 2 gC L-1 of bicarbonate stimulated cell growth, nutrients consumption and production of PHB. Using this concentration, a 14%dcw of PHB and an average productivity of 2.45 mgPHB L-1 d-1 were obtained. Gene expression analysis revelated that these conditions caused the overexpression of genes related to glycogen and PHB synthesis. Moreover, a positive correlation between the genes codifying for the glycogen phosphorylase, the acetyl-CoA reductase and the poly(3-hydroxyalkanoate) polymerase was found, meaning that PHB synthesis and glycogen catabolism are strongly related. These results provide an exhaustive evaluation of the effect of carbon on the PHB production and cyanobacterial metabolism.
Assuntos
Synechocystis , Carbono/metabolismo , Dióxido de Carbono/metabolismo , Glicogênio/metabolismo , Poliésteres/metabolismo , Synechocystis/genética , Synechocystis/metabolismo , Águas ResiduáriasRESUMO
We studied how sunflower plants affect rhamnolipid biosurfactant mobilization of slowly desorbing fractions of polycyclic aromatic hydrocarbons (PAHs) in soil from a creosote-contaminated site. Desorption kinetics of 13 individual PAHs revealed that the soil contained initially up to 50% slowly desorbing fractions. A rhamnolipid biosurfactant was applied to the soil at the completion of the sunflower cycle (75 days in greenhouse conditions). After this period, the PAHs that remained in the soil were mainly present in a slowly desorbing form as a result of the efficient biodegradation of fast-desorbing PAHs by native microbial populations. The rhamnolipid enhanced the bioavailable fraction of the remaining PAHs by up to 30%, as evidenced by a standardized desorption extraction with Tenax, but the enhancement occurred with only planted soils. The enhanced bioavailability did not decrease residual PAH concentrations under greenhouse conditions, possibly due to ecophysiological limitations in the biodegradation process that were independent of the bioavailability. However, biodegradation was enhanced during slurry treatment of greenhouse planted soils that received the biosurfactant. The addition of rhamnolipids caused a dramatic shift in the soil bacterial community structure, which was magnified in the presence of sunflower plants. The stimulated groups were identified as fast-growing and catabolically versatile bacteria. This new rhizosphere microbial biomass possibly interacted with the biosurfactant to facilitate intra-aggregate diffusion of PAHs, thus enhancing the kinetics of slow desorption. Our results show that the usually limited biosurfactant efficiency with contaminated field soils can be significantly enhanced by integrating the sunflower ontogenetic cycle into the bioremediation design.
Assuntos
Rizosfera , Biodegradação Ambiental , Hidrocarbonetos Policíclicos Aromáticos , Solo , Microbiologia do Solo , Poluentes do SoloRESUMO
The aim of this study was to investigate the PHB and glycogen accumulation dynamics in two photobioreactors inoculated with different monocultures of wastewater-borne cyanobacteria, using a three-stage feeding strategy (growth phase, feast-famine phase and feast phase). Two cyanobacterial monocultures containing members of Synechocystis sp. or Synechococcus sp. were collected from treated wastewater and inoculated in lab-scale photobioreactors to evaluate the PHB and glycogen accumulation. A third photobioreactor with a complex microbial community grown in real wastewater was also set up. During each experimental phase different concentrations of inorganic carbon were applied to the cultures, these shifts allowed to discern the accumulation mechanism of carbon storage polymers (PHB and glycogen) in cyanobacteria. Conversion of one into the other was directly related to the carbon content. The highest PHB and glycogen contents (5.04%dcw and 69%dcw, respectively) were achieved for Synechocystis sp.
Assuntos
Fotobiorreatores , Synechocystis , Carbono , Glicogênio , Águas ResiduáriasRESUMO
The pyrene-degrading Mycobacterium sp. strain AP1 grew in nutrient-supplemented artificial seawater with a heavy fuel oil as the sole carbon source, causing the complete removal of all linear (C(12) to C(40)) and branched alkanes from the aliphatic fraction, as well as an extensive degradation of the three- and four-ring polycyclic aromatic hydrocarbons (PAHs) phenanthrene (95%), anthracene (80%), fluoranthene (80%), pyrene (75%), and benzo(a)anthracene (30%). Alkylated PAHs, which are more abundant in crude oils than the nonsubstituted compounds, were selectively attacked at extents that varied from more than 90% for dimethylnaphthalenes, methylphenanthrenes, methylfluorenes, and methyldibenzothiophenes to about 30% for monomethylated fluoranthenes/pyrenes and trimethylated phenanthrenes and dibenzothiophenes. Identification of key metabolites indicated the utilization of phenanthrene, pyrene, and fluoranthene by known assimilatory metabolic routes, while other components were cooxidized. Detection of mono- and dimethylated phthalic acids demonstrated ring cleavage and further oxidation of alkyl PAHs. The extensive degradation of the alkanes, the two-, three-, and four-ring PAHs, and their 1-, 2-, and 3-methyl derivatives from a complex mixture of hydrocarbons by Mycobacterium sp. strain AP1 illustrates the great substrate versatility of alkane- and PAH-degrading mycobacteria.
Assuntos
Óleos Combustíveis , Hidrocarbonetos/metabolismo , Mycobacterium/crescimento & desenvolvimento , Mycobacterium/metabolismo , Água do Mar/microbiologia , Meios de Cultura/químicaRESUMO
The application of a rhamnolipid biosurfactant for enhanced solubilization and biodegradation of slowly desorbing polycyclic aromatic hydrocarbons (PAHs) in contaminated soils was determined in this study. The soil samples exhibited different levels of pollution and different bioremediation stages: the first soil originated from a creosote-polluted site, contained 4370â¯mgâ¯kg -1 of PAHs and had not been bioremediated; the second soil was the same as the first but had received bioremediation treatment with nutrient amendment in biopiles for a period of 5â¯months and contained 580â¯mgâ¯kg -1 of PAHs after this treatment; the third soil was treated by bioremediation for several years to reduce the concentration of PAHs to 275â¯mgâ¯kg -1. The kinetics of PAH desorption were determined to assess the magnitude of the slowly desorbing fractions present in the polluted soil and to optimize the biosurfactant effectiveness in terms of biodegradation. The soils that had been treated by bioremediation were enriched in slowly desorbing PAHs. The rhamnolipid at a concentration above its critical micelle concentration enhanced biodegradation in the soils that had been bioremediated previously. The measurement of residual concentrations of native PAHs showed the promoting effect of the biosurfactant on the biodegradation of the slowly desorbing fractions. Interestingly, benzo(a)pyrene was biodegraded in the soil that had been bioremediated for a long time. Rhamnolipid can constitute a valid alternative to chemical surfactants in promoting the biodegradation of slow-desorption PAHs, which is one of the most important problems in bioremediation, but the efficiency depends strongly on the bioremediation stage in which the biosurfactant is applied.
Assuntos
Biodegradação Ambiental , Glicolipídeos/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Poluentes do Solo/metabolismo , Microbiologia do Solo , TensoativosRESUMO
Cycling of pollutants is essential to preserve functional marine and terrestrial ecosystems. Progress in optimizing these natural biological processes relies on the identification of the underlying microbial actors and deciphering their interactions at molecular, cellular, community, and ecosystem level. Novel advances on PAH biodegradation are built on a progressive approach that span from pure cultures to environmental communities, illustrating the complex metabolic networks within a single cell, and their further implications in higher complexity systems. Recent analytical chemistry and molecular tools allow a deeper insight into the active microbial processes actually occurring in situ, identifying active functions, metabolic pathways and key players. Understanding these processes will provide new tools to assess biodegradation occurrence and, as a final outcome, predict the success of bioremediation thus reducing its uncertainties, the main drawback of this environmental biotechnology.
Assuntos
Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Biodegradação Ambiental , Ecossistema , Humanos , Oceanos e Mares , Microbiologia do SoloRESUMO
This study seeks to improve our understanding of the conceptual model of pollutant transport and fate in cases of DNAPL contamination at sites with a complex contamination history. The study was carried out in an unconfined aquifer of alluvial fans in the Tarragona Petrochemical Complex (Spain). Two boreholes were drilled and continuous cores were recovered in order to carry out a detailed core description at centimeter scale and a comprehensive sampling of borehole cores. The biogeochemical heterogeneity at these sites is controlled by the conjunction of lithological, hydrochemical and microbiological heterogeneities. Biodegradation processes of contaminant compounds take place not only at the level of the dissolved fraction in the aquifer but also at the level of the fraction retained in the fine, less conductive materials as shown by the biodegradation haloes of parent and metabolite compounds. Sampling the low-conductivity levels also allowed us to identify compounds, e.g. BTEX, that are the remaining traces of the passage of old contaminant plumes whose sources no longer exist. This enabled us to describe past biogeochemical processes and to partially account for the processes occurring today. Transition zones, characterized by numerous textural changes, constitute ecotones whose biostimulation could be effective in promoting the acceleration of the remediation of the multiple pollution at these sites.
Assuntos
Sedimentos Geológicos , Água Subterrânea , Modelos Teóricos , Microbiologia do Solo , Solo , Poluentes Químicos da Água/análise , Biodegradação Ambiental , Tetracloreto de Carbono/análise , Clorofórmio/análise , Monitoramento Ambiental , Cloreto de Metila/análise , Espanha , Compostos Orgânicos Voláteis/análiseRESUMO
New Gram-positive and Gram-negative bacteria were isolated from Poeni oily sludge, using enrichment procedures. The six Gram-positive strains belong to Bacillus, Lysinibacillus and Rhodococcus genera. The eight Gram-negative strains belong to Shewanella, Aeromonas, Pseudomonas and Klebsiella genera. Isolated bacterial strains were tolerant to saturated (i.e., n-hexane, n-heptane, n-decane, n-pentadecane, n-hexadecane, cyclohexane), monoaromatic (i.e., benzene, toluene, styrene, xylene isomers, ethylbenzene, propylbenzene) and polyaromatic (i.e., naphthalene, 2-methylnaphthalene, fluorene) hydrocarbons, and also resistant to different antimicrobial agents (i.e., ampicillin, kanamycin, rhodamine 6G, crystal violet, malachite green, sodium dodecyl sulfate). The presence of hydrophilic antibiotics like ampicillin or kanamycin in liquid LB-Mg medium has no effects on Gram-positive and Gram-negative bacteria resistance to toxic compounds. The results indicated that Gram-negative bacteria are less sensitive to toxic compounds than Gram-positive bacteria, except one bacteria belonging to Lysinibacillus genus. There were observed cellular and molecular modifications induced by ampicillin or kanamycin to isolated bacterial strains. Gram-negative bacteria possessed between two and four catabolic genes (alkB, alkM, alkB/alkB1, todC1, xylM, PAH dioxygenase, catechol 2,3-dioxygenase), compared with Gram-positive bacteria (except one bacteria belonging to Bacillus genus) which possessed one catabolic gene (alkB/alkB1). Transporter genes (HAE1, acrAB) were detected only in Gram-negative bacteria.
Assuntos
Anti-Infecciosos/farmacologia , Sistema Enzimático do Citocromo P-450/genética , Farmacorresistência Bacteriana Múltipla , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Hidrocarbonetos/farmacologia , Alcanos/farmacologia , Ampicilina/farmacologia , Transporte Biológico/genética , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Bactérias Gram-Negativas/crescimento & desenvolvimento , Bactérias Gram-Negativas/isolamento & purificação , Bactérias Gram-Negativas/metabolismo , Bactérias Gram-Positivas/crescimento & desenvolvimento , Bactérias Gram-Positivas/isolamento & purificação , Bactérias Gram-Positivas/metabolismo , Hidrocarbonetos Aromáticos/farmacologia , Canamicina/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Oxigenases de Função Mista/genética , Reação em Cadeia da Polimerase , Proteínas/efeitos dos fármacos , Proteínas/metabolismo , RNA Ribossômico 16S/genética , Esgotos/microbiologia , beta-Galactosidase/efeitos dos fármacos , beta-Galactosidase/metabolismoRESUMO
A marine microbial consortium obtained from a beach contaminated by the Prestige oil spill proved highly efficient in removing the different hydrocarbon families present in this heavy fuel oil. Seawater cultures showed a complete removal of all the linear and branched alkanes, an extensive attack on three to five-ring polycyclic aromatic hydrocarbons [PAHs; including anthracene, fluoranthene, pyrene, benzo(a)anthracene, chrysene, and benzo(a)pyrene] (30-100%), and a considerable depletion of their alkyl derivatives. Community dynamics analysis revealed that Alcanivorax species, known alkane degraders, predominated in the initial stages. This was followed by an increase in Alphaproteobacteria (i.e. Maricaulis, Roseovarius), which coincided with the depletion of low molecular PAHs. Finally, these were succeeded by Gammaproteobacteria (mainly Marinobacter and Methylophaga), which were involved in the degradation of the high molecular-weight PAHs. The role of these populations in the removal of the specific components was confirmed by the analysis of subcultures established using the aliphatic or the aromatic fraction of the fuel oil, or single PAHs, as carbon sources. The genus Marinobacter seemed to play a major role in the degradation of a variety of hydrocarbons, as several members of this group were isolated from the different enrichment cultures and grew on plates with hexadecane or single PAHs as sole carbon sources.
Assuntos
Bactérias/metabolismo , Óleos Combustíveis/microbiologia , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Água do Mar/microbiologia , Microbiologia da Água , Bactérias/genética , Bactérias/isolamento & purificação , Biodegradação Ambiental , DNA Bacteriano/genética , Poluição Ambiental , Biblioteca Gênica , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNARESUMO
A Gram-negative bacterium designated UBF-P1(T) was isolated from an enrichment culture established in nutrient supplemented artificial sea water with pyrene as a carbon source, and inoculated with a marine fuel oil-degrading consortium obtained from a sand sample collected from the beach of Corrubedo (A Coruña, Galicia, Spain) after the Prestige accidental oil spill. Phylogenetic analysis based on the almost complete 16S rRNA gene sequence affiliated strain UBF-P1(T) with the family Cohaesibacteraceae, Cohaesibacter gelatinilyticus (DSM 18289(T)) being the closest relative species with 92% sequence similarity. Cells were irregular rods, motile, strictly aerobic, catalase and oxidase positive. Ubiquinone 10 was the major respiratory lipoquinone. The major polar lipids comprised diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), phosphatidylmonomethylethanolamine (PME), and phosphatidylcholine (PC). The major fatty acids detected were C(18:1)ω7c, C(19:0) cycloω8c, and C(16:0). The G+C content of strain UBF-P1(T) was 63.9 mol%. The taxonomic comparison with the closest relative based on genotypic, phenotypic and chemotaxonomic characteristics supported that strain UBF-P1(T) could be classified as a novel genus and species, for which the name Breoghania corrubedonensis gen. nov., sp. nov. is proposed. The type strain of this new taxon is UBF-P1(T) (CECT 7622, LMG 25482, DSM 23382).
Assuntos
Alphaproteobacteria/classificação , Alphaproteobacteria/isolamento & purificação , Microbiologia Ambiental , Alphaproteobacteria/genética , Alphaproteobacteria/crescimento & desenvolvimento , Composição de Bases , Carbono/metabolismo , Análise por Conglomerados , Meios de Cultura/química , DNA Bacteriano/química , DNA Bacteriano/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Poluentes Ambientais , Ácidos Graxos/análise , Óleos Combustíveis , Dados de Sequência Molecular , Fosfolipídeos/análise , Filogenia , Pirenos/metabolismo , Quinonas/análise , RNA Ribossômico 16S/genética , Água do Mar/microbiologia , Análise de Sequência de DNA , EspanhaRESUMO
During bioremediation of polycyclic aromatic hydrocarbon (PAH)-polluted soils accumulation of polar metabolites resulting from the biological activity may occur. Since these polar metabolites are potentially more toxic than the parental products, a better understanding of the processes involved in the production and fate of these oxidation products in soil is needed. In the present work we describe the design and set-up of a static soil microcosm system and an analytical methodology for detection of PAHs and their oxidation products in soils. When applied to a soil contaminated with phenanthrene, as a model PAH, and 1-hydroxy-2-naphthoic acid, diphenic acid, and phthalic acid as putative metabolites, the extraction and fractionation procedures resulted in recoveries of 93%, 89%, 100%, and 89%, respectively. The application of the standardized system to study the biodegradation of phenanthrene in an agricultural soil with and without inoculation of the high molecular weight PAH-degrading strain Mycobacterium sp. AP1, demonstrates its suitability for determining the environmental fate of PAHs in polluted soils and for evaluating the effect of bioremediative treatments. In inoculated microcosms 35% of the added phenanthrene was depleted, 19% being recovered as CO(2) and 3% as diphenic acid. The latter, together with other two unidentified metabolites, accumulated in soil.
Assuntos
Mycobacterium/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/análise , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Solo/análise , CinéticaRESUMO
When incubated with a creosote-polycyclic aromatic hydrocarbons (PAHs) mixture, the pyrene-degrading strain Mycobacterium sp. AP1 acted on three- and four-ring components, causing the simultaneous depletion of 25% of the total PAHs in 30 days. The kinetics of disappearance of individual PAHs was consistent with differences in aqueous solubility. During the incubation, a number of acid metabolites indicative of distinctive reactions carried out by high-molecular-weight PAH-degrading mycobacteria accumulated in the medium. Most of these metabolites were dicarboxylic aromatic acids formed as a result of the utilization of growth substrates (phenanthrene, pyrene, or fluoranthene) by multibranched pathways including meta- and ortho-ring-cleavage reactions: phthalic acid, naphthalene-1,8-dicarboxylic acid, phenanthrene-4,5-dicarboxylic acid, diphenic acid, Z-9-carboxymethylenefluorene-1-carboxylic acid, and 6,6'-dihydroxy-2,2'-biphenyl dicarboxylic acid. Others were dead-end products resulting from cometabolic oxidations on nongrowth substrates (fluorene meta-cleavage product). These results contribute to the general knowledge of the biochemical processes that determine the fate of the individual components of PAH mixtures in polluted soils. The identification of the partially oxidized compounds will facilitate to develop analytical methods to determine their potential formation and accumulation in contaminated sites.
Assuntos
Creosoto/metabolismo , Mycobacterium/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Pirenos/metabolismo , Biodegradação Ambiental , Meios de Cultura/química , Ácidos Dicarboxílicos/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Oxirredução , Fatores de TempoRESUMO
The pyrene-degrading Mycobacterium strain AP1 was found to utilize fluoranthene as a sole source of carbon and energy. Identification of metabolites formed from fluoranthene (by growing cells and washed-cell suspensions), the kinetics of metabolite accumulation, and metabolite-feeding studies all indicated that strain AP1 oxidizes fluoranthene using three alternative routes. The first route is initiated by dioxygenation at C-7 and C-8 and, following meta cleavage and pyruvate release, produces a hydroxyacenaphthoic acid that is decarboxylated to acenaphthenone (V). Monooxygenation of this ketone to the quinone and subsequent hydrolysis generates naphthalene-1,8-dicarboxylic acid (IV), which is further degraded via benzene-1,2,3-tricarboxylic acid (III). A second route involves dioxygenation at C-1 and C-2, followed by dehydrogenation and meta cleavage of the resulting diol. A two-carbon fragment excision of the meta cleavage product yields 9-fluorenone-1-carboxylic acid (II), which appears to undergo angular dioxygenation and further degradation to produce benzene-1,2,3-tricarboxylic acid (III), merging this route with the 7,8-dioxygenation route. Decarboxylation of benzene-1,2,3-tricarboxylic acid to phthalate (VIII), as well as further oxidation of the latter, would connect both routes with the central metabolism. The identification of Z-9-carboxymethylenefluorene-1-carboxylic acid (I) suggests a third route for fluoranthene degradation involving dioxygenation at C-2, C-3, and ortho cleavage. There is no evidence of any further degradation of this compound.
Assuntos
Fluorenos/metabolismo , Mycobacterium/metabolismo , Microbiologia do Solo , Biodegradação Ambiental , Cromatografia Líquida de Alta Pressão , Meios de Cultura , Cromatografia Gasosa-Espectrometria de Massas , Espectroscopia de Ressonância Magnética , Mycobacterium/classificação , Mycobacterium/crescimento & desenvolvimento , Mycobacterium/isolamento & purificação , Petróleo/metabolismo , Ácidos Ftálicos/metabolismo , Poluentes do Solo/metabolismoRESUMO
Mycobacterium sp. strains CP1, CP2, CFt2 and CFt6 were isolated from creosote-contaminated soil due to their ability to grow in pyrene (CP1 and CP2) or fluoranthene (CFt2 and CFt6). All these strains utilized fluoranthene as a sole source of carbon and energy. Strain CP1 exhibited the best growth, with a cellular assimilation of fluoranthene carbon of approximately 45%. Identification of the metabolites accumulated during growth in fluoranthene, the kinetics of metabolites, and metabolite feeding studies, indicated that all these isolates oxidized fluoranthene by the following two routes: the first involves dioxygenation at C-1 and C-2, meta cleavage, and a 2-carbon fragment excision to produce 9-fluorenone-1-carboxylic acid. An angular dioxygenation of the latter yields cis-1,9a-dihydroxy-1-hydrofluorene-9-one-8-carboxylic acid, which is further degraded via 8-hydroxy-3,4-benzocoumarin-1-carboxylic acid, benzene-1,2,3-tricarboxylic acid, and phthalate; the second route involves dioxygenation at C-2 and C-3 and ortho cleavage to give Z-9-carboxymethylenefluorene-1-carboxylic acid. In addition, the pyrene-degrading strains CP1 and CP2 possess a third route initiated by dioxygenation at positions C-7 and C-8, which--following meta cleavage, an aldolase reaction, and a C(1)-fragment excision--yields acenaphthenone. Monooxygenation of this ketone to the corresponding quinone, and its subsequent hydrolysis, produces naphthalene-1,8-dicarboxylic acid. The results obtained in this study not only complete and confirm the three fluoranthene degradation routes previously proposed for the pyrene-degrading strain Mycobacterium sp. AP1, but also suggest that such routes represent general microbial processes for environmental fluoranthene removal.