Isolation of a Pseudomonas stutzeri strain that degrades o-xylene

A Pseudomonas stutzeri strain capable of growing on o-xylene was isolated from enrichment cultures. The organism grew on 2,3- and 3,4-dimethylphenol but not on 2-methylbenzyl alcohol, o-tolualdehyde, or o-toluate. P. stutzeri was not able to utilize m-xylene, p-xylene, or 1,2,4-trimethylbenzene, but growth was observed in the presence of the corresponding alcohols and acids. From the Pseudomonas cultures supplied with o-xylene, 2,3-dimethylphenol was isolated and identified. When resting P. stutzeri cells were incubated with 2,3-dimethylphenol, the reaction mixture turned greenish yellow and showed spectral properties identical to those of the 3,4-dimethylcatechol meta ring fission product. Catechol 2,3-oxygenase was induced by growth on o-xylene or on 2,3- or 3,4-dimethylphenol. The suggested hypothesis is that the first metabolic steps of growth on o-xylene involve the direct oxygenation of the aromatic nucleus, followed by meta pathway reactions.     

Plasmid-encoded mercury resistance in a Pseudomonas stutzeri strain that degrades o-xylene

Abstract A Pseudomonas stutzeri strain, previously isolated for its ability to utilize o -xylene, bears a plasmid, pPB, of about 80 kbp. pPB was found to encode resistance to mercuric chloride and organomercury compounds. Loss of the plasmid resulted in a simultaneous loss of the metal resistance, but not of the ability to degrade o -xylene. Transfer of the Hg^r phenotype to an Hg^s strain was achieved by mobilizing pPB with RP4. Mercury reductase activity was induced by mercuric chloride and by phenylmercuric acetate and Thimerosal. pPB may be considered a broad spectrum resistance plasmid.     

Isolation and characterization of a Pseudomonas strain that degrades 4-acetamidophenol and 4-aminophenol

Though many microorganisms that are capable of using phenol as sole sourceof carbon have been isolated and characterized, only a few organisms degradingsubstituted phenols have been described to date. In this study, one strain ofmicroorganism that is capable of using phenol (3000 ppm), 4-aminophenol(4000 ppm) and 4-acetamidophenol (4000 ppm) as sole source of carbon andenergy was isolated and characterized. This strain was obtained by enrichmentculture from a site contaminated with compounds like 4-acetamidophenol,4-aminophenol and phenol in Pakistan at Bhai Pheru. The contaminated siteis able to support large bacterial community as indicated by the viable cellcounts (2 × 10⁴–5 × 10⁸) per gram of soil. Detailed taxonomic studies identified the organisms as Pseudomonas species designated as strain STI. The isolate also showed growth on other organic compounds like aniline, benzene, benzyl alcohol, benzyl bromide, toluene, -cresol, trichloroethylene and o-xylene. Optimum growth temperature and pH were found to be 30 °C and 7, respectively, while growth at 4, 25 and 35 °C and at pH 8 and 9 was also observed. Non growing suspended cells of strain ST1 degraded 68, 96 and 76.8% of 4-aminophenol (1000 ppm), phenol (500 ppm) and 4-acetamidophenol (1000 ppm), respectively, in 72 hrs. The isolation and characterization of Pseudomonas speciesstrain ST1, may contribute to efforts on phenolic bioremediation, particularly in anenvironment with very high levels of 4-acetamidophenol and 4-aminophenol.     

Isolation and characterization of a thermotolerant bacterium Ralstonia sp. strain PHS1 that degrades benzene, toluene, ethylbenzene, and o-xylene

A thermotolerant bacterium, designated as PHS1, was isolated from a hot spring in Pohang, Korea, on the basis of its ability to grow on benzene, toluene, ethylbenzene, and xylenes (BTEX) as a sole carbon source. Strain PHS1 is a gram-negative, rod-shaped aerobe and grows optimally at 42 degrees C and pH 7.2. According to 16 S rDNA analysis, strain PHS1 showed highest similarity to Ralstonia eutropha (previously named Alcaligenes eutrophus). Unlike its closest known Ralstonia species, however, strain PHS1 was able to utilize toluene, ethylbenzene, o-xylene, and both m- and o-cresol. The degradation of o-xylene by strain PHS1 is particularly important, since o-xylene is a compound of considerable environmental interest, owing to its recalcitrance; and very few microorganisms have been reported to utilize o-xylene as a sole carbon source. It was found that strain PHS1 transformed o-xylene to 2,3-dimethylphenol through direct oxygenation of the aromatic ring. The unique properties of strain PHS1, such as thermotolerance and the ability to degrade o-xylene, may have important implications for the treatment of BTEX-contaminated industrial effluents.     

Isolation and characterization of a sulfate-reducing bacterium that anaerobically degrades alkanes.

An alkane-degrading, sulfate-reducing bacterial strain, AK-01, was isolated from an estuarine sediment with a history of chronic petroleum contamination. The bacterium is a short, nonmotile, non-spore-forming, gram-negative rod. It is mesophilic and grows optimally at pH 6.9 to 7.0 and at an NaCl concentration of 1%. Formate, fatty acids (C4 to C16) and hydrogen were readily utilized as electron donors. Sulfate, sulfite, and thiosulfate were used as electron acceptors, but sulfur, nitrite, and nitrate were not. Phenotypic characterization and phylogenetic analysis based on 16S rRNA gene sequence indicate that AK-01 is most closely related to the genera Desulfosarcina, Desulfonema, and Desulfococcus in the delta subdivision of the class Proteobacteria. It is phenotypically and phylogenetically different from strains Hxd3 and TD3, two previously reported isolates of alkane-degrading, sulfate-reducing bacteria. The alkanes tested to support growth of AK-01 had chain lengths of C13 to C18. 1-Alkenes (C15 and C16) and 1-alkanols (C15 and C16) also supported growth. The doubling time for growth on hexadecane was 3 days, about four times longer than that for growth on hexadecanoate. Mineralization of hexadecane was indicated by the recovery of 14CO2 from cultures grown on [1-14C]hexadecane. Degradation of hexadecane was dependent on sulfate reduction. The stoichiometric ratio (as moles of sulfate reduced per mole of hexadecane degraded) was 10.6, which is very close to the theoretical ratio of 12.25, assuming a complete oxidation to CO2. Anaerobic alkane degradation by sulfate reducers may be a more widespread phenomenon than was previously thought.     

Biology of Pseudomonas stutzeri.

Pseudomonas stutzeri is a nonfluorescent denitrifying bacterium widely distributed in the environment, and it has also been isolated as an opportunistic pathogen from humans. Over the past 15 years, much progress has been made in elucidating the taxonomy of this diverse taxonomical group, demonstrating the clonality of its populations. The species has received much attention because of its particular metabolic properties: it has been proposed as a model organism for denitrification studies; many strains have natural transformation properties, making it relevant for study of the transfer of genes in the environment; several strains are able to fix dinitrogen; and others participate in the degradation of pollutants or interact with toxic metals. This review considers the history of the discovery, nomenclatural changes, and early studies, together with the relevant biological and ecological properties, of P. stutzeri.     

The rice inoculant strain Alcaligenes faecalis A15 is a nitrogen-fixing Pseudomonas stutzeri.

The taxonomic position of the nitrogen-fixing rice isolate A15, previously classified as Alcaligenes faecalis, was reinvestigated. On the basis of its small subunit ribosomal RNA (16S rRNA) sequence this strain identifies as Pseudomonas stutzeri. Phenotyping and fatty acid profiling confirm this result. DNA:DNA hybridisations, using the optical renaturation rate method, between strain A15 and Pseudomonas stutzeri LMG 11199T revealed a mean DNA-binding of 77%. The identification was further corroborated by comparative sequence analysis of the oprF gene, which encodes the major outer membrane protein of rRNA homology group I pseudomonads. Furthermore we determined the nifH sequence of this strain and of two putative diazotrophic Pseudomonas spp. and made a comparative analysis with sequences of other diazotrophs. These Pseudomonas NifH sequences cluster with NifH sequences isolated from the rice rhizosphere by PCR and of proteobacteria from the beta and gamma subclasses.     

石油生物脱硫菌Pseudomonas stutzeri UP-1的筛选

以二苯并噻吩(DBT)为模型化合物,筛选到一株能有效降解DBT的菌株,根据其菌落的形态特征、生理生化特征和分子生物学鉴定方法,确定其为Pseudomonms stutzer UP1。该菌株对DBT具有较强的降解能力,降解终产物为水溶性物质。通过对降解产物的分析,初步推断DBT的降解符合Kodama机理。     

Organization and regulation of meta cleavage pathway genes for toluene and o-xylene derivative degradation in Pseudomonas stutzeri OX1.

Pseudomonas stutzeri OX1 meta pathway genes for toluene and o-xylene catabolism were analyzed, and loci encoding phenol hydroxylase, catechol 2,3-dioxygenase, 2-hydroxymuconate semialdehyde dehydrogenase, and 2-hydroxymuconate semialdehyde hydrolase were mapped. Phenol hydroxylase converted a broad range of substrates, as it was also able to transform the nongrowth substrates 2,4-dimethylphenol and 2,5-dimethylphenol into 3,5-dimethylcatechol and 3,6-dimethylcatechol, respectively, which, however, were not cleaved by catechol 2,3-dioxygenase. The identified gene cluster displayed a gene order similar to that of the Pseudomonas sp. strain CF600 dmp operon for phenol catabolism and was found to be coregulated by the tou operon activator TouR. A hypothesis about the evolution of the toluene and o-xylene catabolic pathway in P. stutzeri OX1 is discussed.     

Isolation and characterization of a fluoranthene-utilizing strain of Pseudomonas paucimobilis.

A soil bacterium capable of utilizing fluoranthene as the sole source of carbon and energy for growth was purified from a seven-member bacterial community previously isolated from a creosote waste site for its ability to degrade polycyclic aromatic hydrocarbons. By standard bacteriological methods, this bacterium was characterized taxonomically as a strain of Pseudomonas paucimobilis and was designated strain EPA505. Utilization of fluoranthene by strain EPA 505 was demonstrated by increase in bacterial biomass, decrease in aqueous fluoranthene concentration, and transient formation of transformation products in liquid cultures where fluoranthene was supplied as the sole carbon source. Resting cells grown in complex medium showed activity toward anthraquinone, benzo[b]fluorene, biphenyl, chrysene, and pyrene as demonstrated by the disappearance of parent compounds or changes in their UV absorption spectra. Fluoranthene-grown resting cells were active against these compound as well as 2,3-dimethylnaphthalene, anthracene, fluoranthene, fluorene, naphthalene, and phenanthrene. These studies demonstrate that organic compounds not previously reported to serve as growth substrates can be utilized by axenic cultures of microorganisms. Such organisms may possess novel degradative systems that are active toward other compounds whose biological degradation has been limited because of inherent structural considerations or because of low aqueous solubility.     

Isolation and characterisation of a strain of Pseudomonas putida that can express a periplasmic nitrate reductase

A strain of Pseudomonas putida that can express a nitrate reductase that is located in the periplasmic compartment was isolated from freshwater. The enzyme was active in vivo during arginine fermentation and at the onset of oxygen limitation in batch cultures. The activity of the enzyme increased the yield of bacteria following fermentative growth under anoxic conditions with arginine, but nitrate reduction did not support growth on nonfermentable carbon substrates under anoxic conditions. Cells expressing the periplasmic nitrate reductase were capable of reducing nitrate in the presence of oxygen. Nitrate reduction under oxic conditions was clearly coupled to a respiratory electron transport chain because: (1) the process was sensitive to the respiratory inhibitors rotenone and 2-n-heptyl-4-hydroxyquinoline N-oxide, and (2) membrane-bound and periplasmic cytochromes were involved. This is the first report of the presence of a periplasmic nitrate reductase in a member of the proteobacteria.     

Complete genome sequence of the type strain Pseudomonas stutzeri CGMCC 1.1803

Here we report the complete genome sequence of Pseudomonas stutzeri strain CGMCC 1.1803 (equivalent to ATCC 17588), the type strain of P. stutzeri, which encodes 4,138 open reading frames on a 4,547,930-bp circular chromosome. The CGMCC 1.1803 genome contains genes involved in denitrification, benzoate/catechol degradation, chemotaxis, and other functions.     

Phenol hydroxylase and toluene/o-xylene monooxygenase from Pseudomonas stutzeri OX1: interplay between two enzymes

Degradation of aromatic hydrocarbons by aerobic bacteria is generally divided into an upper pathway, which produces dihydroxylated aromatic intermediates by the action of monooxygenases, and a lower pathway, which processes these intermediates down to molecules that enter the citric acid cycle. Bacterial multicomponent monooxygenases (BMMs) are a family of enzymes divided into six distinct groups. Most bacterial genomes code for only one BMM, but a few cases (3 out of 31) of genomes coding for more than a single monooxygenase have been found. One such case is the genome of Pseudomonas stutzeri OX1, in which two different monooxygenases have been found, phenol hydroxylase (PH) and toluene/o-xylene monooxygenase (ToMO). We have already demonstrated that ToMO is an oligomeric protein whose subunits transfer electrons from NADH to oxygen, which is eventually incorporated into the aromatic substrate. However, no molecular data are available on the structure and on the mechanism of action of PH. To understand the metabolic significance of the association of two similar enzymatic activities in the same microorganism, we expressed and characterized this novel phenol hydroxylase. Our data indicate that the PH P component of PH transfers electrons from NADH to a subcomplex endowed with hydroxylase activity. Moreover, a regulatory function can be suggested for subunit PH M. Data on the specificity and the kinetic constants of ToMO and PH strongly support the hypothesis that coupling between the two enzymatic systems optimizes the use of nonhydroxylated aromatic molecules by the draining effect of PH on the product(s) of oxidation catalyzed by ToMO, thus avoiding phenol accumulation.     

Isolation and characterization of a bacterium that degrades various polyester-based biodegradable plastics

Microorganisms isolated from soil samples were screened for their ability to degrade various biodegradable polyester-based plastics. The most active strain, designated as strain TB-13, was selected as the best strain for degrading these plastics. From its phenotypic and genetic characteristics, strain TB-13 was closely related to Paenibacillus amylolyticus. It could degrade poly(lactic acid), poly(butylene succinate), poly(butylene succinate-co-adipate), poly(caprolactone) and poly(ethylene succinate) but not poly(hydroxybutylate-co-valerate). However, it could not utilize these plastics as sole carbon sources. Both protease and esterase activities, which may be involved in the degradation of plastic, were constitutively detected in the culture broth.     

Construction of a Pseudomonas hybrid strain that mineralizes 2,4,6-trinitrotoluene.

A bacterium, Pseudomonas sp. strain C1S1, able to grow on 2,4,6-trinitrotoluene (TNT), 2,4- and 2,6-dinitrotoluene, and 2-nitrotoluene as N sources, was isolated. The bacterium grew at 30 degrees C with fructose as a C source and accumulated nitrite. Through batch culture enrichment, we isolated a derivative strain, called Pseudomonas sp. clone A, which grew faster on TNT and did not accumulate nitrite in the culture medium. Use of TNT by these two strains as an N source involved the successive removal of nitro groups to yield 2,4- and 2,6-dinitrotoluene, 2-nitrotoluene, and toluene. Transfer of the Pseudomonas putida TOL plasmid pWW0-Km to Pseudomonas sp. clone A allowed the transconjugant bacteria to grow on TNT as the sole C and N source. All bacteria in this study, in addition to removing nitro groups from TNT, reduced nitro groups on the aromatic ring via hydroxylamine to amino derivatives. Azoxy dimers probably resulting from the condensation of partially reduced TNT derivatives were also found.     

Biology of Pseudomonas stutzeri

Pseudomonas stutzeri is a nonfluorescent denitrifying bacterium widely distributed in the environment, and it has also been isolated as an opportunistic pathogen from humans. Over the past 15 years, much progress has been made in elucidating the taxonomy of this diverse taxonomical group, demonstrating the clonality of its populations. The species has received much attention because of its particular metabolic properties: it has been proposed as a model organism for denitrification studies; many strains have natural transformation properties, making it relevant for study of the transfer of genes in the environment; several strains are able to fix dinitrogen; and others participate in the degradation of pollutants or interact with toxic metals. This review considers the history of the discovery, nomenclatural changes, and early studies, together with the relevant biological and ecological properties, of P. stutzeri.