Synergistic degradation of linuron by a bacterial consortium and isolation of a single linuron-degrading variovorax strain

The bacterial community composition of a linuron-degrading enrichment culture and the role of the individual strains in linuron degradation have been determined by a combination of methods, such as denaturing gradient gel electrophoresis of the total 16S rRNA gene pool, isolation and identification of strains, and biodegradation assays. Three strains, Variovorax sp. strain WDL1, Delftia acidovorans WDL34, and Pseudomonas sp. strain WDL5, were isolated directly from the linuron-degrading culture. In addition, subculture of this enrichment culture on potential intermediates in the degradation pathway of linuron (i.e., N,O-dimethylhydroxylamine and 3-chloroaniline) resulted in the isolation of, respectively, Hyphomicrobium sulfonivorans WDL6 and Comamonas testosteroni WDL7. Of these five strains, only Variovorax sp. strain WDL1 was able to use linuron as the sole source of C, N, and energy. WDL1 first converted linuron to 3,4-dichloroaniline (3,4-DCA), which transiently accumulated in the medium but was subsequently degraded. To the best of our knowledge, this is the first report of a strain that degrades linuron further than the aromatic intermediates. Interestingly, the rate of linuron degradation by strain WDL1 was lower than that for the consortium, but was clearly increased when WDL1 was coinoculated with each of the other four strains. D. acidovorans WDL34 and C. testosteroni WDL7 were found to be responsible for degradation of the intermediate 3,4-DCA, and H. sulfonivorans WDL6 was the only strain able to degrade N,O-dimethylhydroxylamine. The role of Pseudomonas sp. strain WDL5 needs to be further elucidated. The degradation of linuron can thus be performed by a single isolate, Variovorax sp. strain WDL1, but is stimulated by a synergistic interaction with the other strains isolated from the same linuron-degrading culture.     

HylA,an Alternative Hydrolase for Initiation of Catabolism of the Phenylurea Herbicide Linuron in Variovorax sp. Strains

Variovorax sp. strain WDL1, which mineralizes the phenylurea herbicide linuron, expresses a novel linuron-hydrolyzing enzyme, HylA, that converts linuron to 3,4-dichloroaniline (DCA). The enzyme is distinct from the linuron hydrolase LibA enzyme recently identified in other linuron-mineralizing Variovorax strains and from phenylurea-hydrolyzing enzymes (PuhA, PuhB) found in Gram-positive bacteria. The dimeric enzyme belongs to a separate family of hydrolases and differs in K_m, temperature optimum, and phenylurea herbicide substrate range. Within the metal-dependent amidohydrolase superfamily, HylA and PuhA/PuhB belong to two distinct protein families, while LibA is a member of the unrelated amidase signature family. The hylA gene was identified in a draft genome sequence of strain WDL1. The involvement of hylA in linuron degradation by strain WDL1 is inferred from its absence in spontaneous WDL1 mutants defective in linuron hydrolysis and its presence in linuron-degrading Variovorax strains that lack libA. In strain WDL1, the hylA gene is combined with catabolic gene modules encoding the downstream pathways for DCA degradation, which are very similar to those present in Variovorax sp. SRS16, which contains libA. Our results show that the expansion of a DCA catabolic pathway toward linuron degradation in Variovorax can involve different but isofunctional linuron hydrolysis genes encoding proteins that belong to evolutionary unrelated hydrolase families. This may be explained by divergent evolution and the independent acquisition of the corresponding genetic modules.     

Characterization of novel linuron-mineralizing bacterial consortia enriched from long-term linuron-treated agricultural soils

Linuron-mineralizing cultures were enriched from two linuron-treated agricultural soils in the presence and absence of a solid support. The cultures contained linuron-degrading bacteria, which coexisted with bacteria degrading either 3,4-dichloroaniline (3,4-DCA) or N,O-dimethylhydroxylamine (N,O-DMHA), two common metabolites in the linuron degradation pathway. For one soil, the presence of a solid support enriched for linuron-degrading strains phylogenetically related to but different from those enriched without support. Most linuron-degrading consortium members were identified as Variovorax, but a Hydrogenophaga and an Achromobacter strain capable of linuron degradation were also obtained. Several of the linuron-degrading isolates also degraded 3,4-DCA. Isolates that degraded 3,4-DCA but not linuron belonged to the genera Variovorax, Cupriavidus and Afipia. Hyphomicrobium spp. were involved in the metabolism of N,O-DMHA. Whereas several isolates degraded linuron independently, more efficient degradation was achieved by combining linuron and 3,4-DCA-degraders or by adding casamino acids. These data suggest that (1) linuron degradation is performed by a group of metabolically interacting bacteria rather than by individual strains, (2) there are other genera in addition to Variovorax that degrade linuron beyond 3,4-DCA, (3) linuron-degrading consortia of different origins have a similar composition, and (4) interactions between consortium members can be complex and can involve exchange of both metabolites and other nutrients.     

Biodegradation of the herbicide propanil, and its 3,4-dichloroaniline by-product in a continuously operated biofilm reactor

The persistence of propanil in soil and aquatic environments along with the possible accumulation of toxic degradation products, such as chloroanilines, is of environmental concern. In this work, a continuous small-scale bioprocess to degrade the herbicide propanil, its main catabolic by-product, 3,4-dichloroaniline (3,4-DCA), and the herbicide adjuvants is carried out. A microbial consortium, constituted by nine bacterial genera, was selected. The isolated strains, identified by amplification and sequencing of their 16S rDNA, were: Acidovorax sp., Luteibacter (rhizovicinus), Xanthomonas sp., Flavobacterium sp., Variovorax sp., Acinetobacter (calcoaceticus), Pseudomonas sp., Rhodococcus sp., and Kocuria sp. The ability of the microbial consortium to degrade the herbicide was evaluated in a biofilm reactor at propanil loading rates ranging from 1.9 to 36.8 mg L^?1 h^?1. Complete removal of propanil, 3,4-DCA, chemical oxygen demand and total organic carbon was obtained at propanil loading rates up to 24.9 mg L^?1 h^?1. At higher loading rates, the removal efficiencies decayed. Four of the identified strains could grow individually in propanil, and 3,4-DCA: Pseudomonas sp., Acinetobacter calcoaceticus, Rhodococcus sp., and Xanthomonas sp. The Kokuria strain grew on 3,4-DCA, but not on propanil. The first three bacteria have been related to biodegradation of phenyl urea herbicides or chlorinated anilines. Although some strains of the genera Xanthomonas and Kocuria have a role in the biodegradation of several xenobiotic compounds, as far as we know, there are no reports about degradation of propanil by Xanthomonas or 3,4-DCA by Kocuria species.     

Construction of a Novel Polychlorinated Biphenyl-Degrading Bacterium: Utilization of 3,4′-Dichlorobiphenyl by Pseudomonas acidovorans M3GY

Pseudomonas acidovorans M3GY is a recombinant bacterium with the novel capacity to utilize a biphenyl congener chlorinated on both rings, 3,4′-dichlorobiphenyl (3,4′-DCBP), as a sole carbon and energy source. Strain M3GY was constructed with a continuous amalgamated culture apparatus (L. Kröckel and D. D. Focht, Appl. Environ. Microbiol. 53:2470-2475, 1987) with P. acidovorans CC1(19), a chloroacetate and biphenyl degrader, and Pseudomonas sp. strain CB15(1), a biphenyl and 3-chlorobenzoate degrader. Genetic and phenotypic data showed the recipient parental strain to be P. acidovorans CC1 and the donor parental strain to be Pseudomonas sp. strain CB15. In growth experiments with 3,4′-DCBP as a sole source of carbon, cultures of strain M3GY increased in absorbance from 0.07 to 0.39 in 29 days while reaching a protein concentration of 58 μg ml^-1 and 67% substrate dehalogenation. 4-Chlorobenzoate was identified from culture supernatants of strain M3GY by gas chromatography-infrared spectrometry-mass spectrometry; this would be consistent with the oxidation of the m-chlorinated ring through the standard biphenyl pathway. 4-Chlorobenzoate was converted to 4-chlorocatechol, which was metabolized through the meta-fission pathway. The construction of P. acidovorans M3GY, with the novel capability to utilize 3,4′-DCBP, thus involves the complete use of meta-fission pathways for sequential rupture of the biphenyl and chlorobenzoate rings.     

Effect of immobilization of a bacterial consortium on diuron dissipation and community dynamics

This work intended to study the relationship between diuron herbicide dissipation and the population dynamics of co-cultivated Delftia acidovorans WDL34 (WDL34) and Arthrobacter sp. N4 (N4) for different cell formulations: free cells or immobilization in Ca-alginate beads of one or both strains. GFP-tagged WDL34 and N4 Gram staining allowed analyzing the cell growth and distribution of each strain in both beads and culture medium in the course of the time. Compared to the free cell co-culture of WDL34 and N4, immobilization of WDL34 in Ca-alginate beads co-cultivated with free N4 increased the dissipation rate of diuron by 53% (0.141 mg ml⁻¹ h⁻¹). In that case, immobilization strongly modified the final equilibrium among both strains (highest total N4 to WDL34 ratio). Our results demonstrated that the inoculant formulation played a major role in the cell growth of each cultivated strain possibly increasing diuron dissipation. This optimized cell formulation may allow improving water and soil treatment.     

Mineralization of diuron [3-(3,4-dichlorophenyl)-1, 1-dimethylurea] by co-immobilized <Emphasis Type="Italic">Arthrobacter</Emphasis> sp. and <Emphasis Type="Italic">Delftia acidovorans</Emphasis>

Mineralization of diuron has not been previously demonstrated despite the availability of some bacteria to degrade diuron into 3,4-dichloroaniline (3,4-DCA) and others that can mineralize 3,4-DCA. A bacterial co-culture of Arthrobacter sp. N4 and Delftia acidovorans W34, which respectively degraded diuron (20 mg l⁻¹) to 3,4-DCA and mineralized 3,4-DCA, were able to mineralize diuron. Total diuron mineralization (20 mg l⁻¹) was achieved with free cells in co-culture. When the bacteria were immobilized (either one bacteria or both), the degradation rate was higher. Best results were obtained with free Arthrobacter sp. N4 cells co-cultivated with immobilized cells of D. acidovorans W34 (mineralization of diuron in 96 h, i.e., 0.21 mg l⁻¹ h⁻¹ vs. 0.06 mg l⁻¹ h⁻¹ with free cells in co-culture).     

The quantity and quality of dissolved organic matter as supplementary carbon source impacts the pesticide-degrading activity of a triple-species bacterial biofilm

Effects of environmental dissolved organic matter (eDOM) that consists of various low concentration carbonic compounds on pollutant biodegradation by bacteria are poorly understood, especially when it concerns synergistic xenobiotic-degrading consortia where degradation depends on interspecies metabolic interactions. This study examines the impact of the quality and quantity of eDOM, supplied as secondary C-source, on the structure, composition and pesticide-degrading activity of a triple-species bacterial consortium in which the members synergistically degrade the phenylurea herbicide linuron, when grown as biofilms. Biofilms developing on 10 mg L^?1 linuron showed a steady-state linuron degradation efficiency of approximately 85 %. The three bacterial strains co-localized in the biofilms indicating syntrophic interactions. Subsequent feeding with eDOM or citrate in addition to linuron resulted into changes in linuron-degrading activity. A decrease in linuron-degrading activity was especially recorded in case of co-feeding with citrate and eDOM of high quality and was always associated with accumulation of the primary metabolite 3,4-dichloroaniline. Improvement of linuron degradation was especially observed with more recalcitrant eDOM. Addition of eDOM/citrate formulations altered biofilm architecture and species composition but without loss of any of the strains and of co-localization. Compositional shifts correlated with linuron degradation efficiencies. When the feed was restored to only linuron, the linuron-degrading activity rapidly changed to the level before the mixed-substrate feed. Meanwhile only minor changes in biofilm composition and structure were recorded, indicating that observed eDOM/citrate effects had been primarily due to repression/stimulation of linuron catabolic activity rather than to biofilm characteristics.     

Environmental Dissolved Organic Matter Governs Biofilm Formation and Subsequent Linuron Degradation Activity of a Linuron-Degrading Bacterial Consortium

It was examined whether biofilm growth on dissolved organic matter (DOM) of a three-species consortium whose members synergistically degrade the phenylurea herbicide linuron affected the consortium''s integrity and subsequent linuron-degrading functionality. Citrate as a model DOM and three environmental DOM (eDOM) formulations of different quality were used. Biofilms developed with all DOM formulations, and the three species were retained in the biofilm. However, biofilm biomass, species composition, architecture, and colocalization of member strains depended on DOM and its biodegradability. To assess the linuron-degrading functionality, biofilms were subsequently irrigated with linuron at 10 mg liter⁻¹ or 100 μg liter⁻¹. Instant linuron degradation, the time needed to attain maximal linuron degradation, and hence the total amount of linuron removed depended on both the DOM used for growth and the linuron concentration. At 10 mg liter⁻¹, the final linuron degradation efficiency was as high as previously observed without DOM except for biofilms fed with humic acids which did not degrade linuron. At 100 μg liter⁻¹ linuron, DOM-grown biofilms degraded linuron less efficiently than biofilms receiving 10 mg liter⁻¹ linuron. The amount of linuron removed was more correlated with biofilm species composition than with biomass or structure. Based on visual observations, colocalization of consortium members in biofilms after the DOM feed appears essential for instant linuron-degrading activity and might explain the differences in overall linuron degradation. The data show that DOM quality determines biofilm structure and composition of the pesticide-degrading consortium in periods with DOM as the main carbon source and can affect subsequent pesticide-degrading activity, especially at micropollutant concentrations.     

Architecture and spatial organization in a triple-species bacterial biofilm synergistically degrading the phenylurea herbicide linuron

Members of a triple-species 3-(3,4-dichlorophenyl)-1-methoxy-1-methyl urea (linuron)-mineralizing consortium, i.e. the linuron- and 3,4-dichloroaniline-degrading Variovorax sp. WDL1, the 3,4-dichloroaniline-degrading Comamonas testosteroni WDL7 and the N,O-dimethylhydroxylamine-degrading Hyphomicrobium sulfonivorans WDL6, were cultivated as mono- or multi-species biofilms in flow cells irrigated with selective or nonselective media, and examined with confocal laser scanning microscopy. In contrast to mono-species biofilms of Variovorax sp. WDL1, the triple-species consortium biofilm degraded linuron completely through apparent synergistic interactions. The triple-species linuron-fed consortium biofilm displayed a heterogeneous structure with an irregular surface topography that most resembled the topography of linuron-fed mono-species WDL1 biofilms, indicating that WDL1 had a dominating influence on the triple-species biofilm architecture. This architecture was dependent on the carbon source supplied, as the biofilm architecture of WDL1 growing on alternative carbon sources was different from that observed under linuron-fed conditions. Linuron-fed triple-species consortium biofilms consisted of mounds composed of closely associated WDL1, WDL7 and WDL6 cells, while this association was lost when the consortium was grown on a nonselective carbon source. In addition, under linuron-fed conditions, microcolonies displaying associated growth developed rapidly after inoculation. These observations indicate that the spatial organization in the linuron-fed consortium biofilm reflected the metabolic interactions within the consortium.     

Inactivation of 2,3-dihydroxybiphenyl 1,2-dioxygenase fromPseudomonas sp. strain CB406 by 3,4-dihydroxybiphenyl (4-phenylcatechol)

Summary 3,4-dihydroxybiphenyl is not a substrate for the 2,3-dihydroxybiphenyl 1,2-dioxygenase (BphC) from biphenyldegradingPseudomonas sp. strain CB406. It acts as both a reversible inhibitor and a potent inactivator of the enzyme. The inactivation process requires the presence of O₂ and can be reversed by the removal of the 3,4-dihydroxybiphenyl followed by incubation of the enzyme in the presence of dithioerythritol and Fe²⁺ under anaerobic conditions. Two other extradiol dioxygenases behave similarly, the catechol 2,3-dioxygenase (BphE) from strain CB406 and the BphC fromPseudomonas sp. strain LB400. The BphC fromP. testosteroni B-356 also did not cleave 3,4-dihydroxybiphenyl but it was not inactivated.Abbreviations C23o Catechol 2,3-dioxygenase - 34DHBP 3,4-dihydroxybiphenyl     

Aerobic Biodegradation of 2,2′-Dithiodibenzoic Acid Produced from Dibenzothiophene Metabolites

Dibenzothiophene is a sulfur heterocycle found in crude oils and coal. The biodegradation of dibenzothiophene through the Kodama pathway by Pseudomonas sp. strain BT1d leads to the formation of three disulfides: 2-oxo-2-(2-thiophenyl)ethanoic acid disulfide, 2-oxo-2-(2-thiophenyl)ethanoic acid-2-benzoic acid disulfide, and 2,2′-dithiodibenzoic acid. When provided as the carbon and sulfur source in liquid medium, 2,2′-dithiodibenzoic acid was degraded by soil enrichment cultures. Two bacterial isolates, designated strains RM1 and RM6, degraded 2,2′-dithiodibenzoic acid when combined in the medium. Isolate RM6 was found to have an absolute requirement for vitamin B₁₂, and it degraded 2,2′-dithiodibenzoic acid in pure culture when the medium was supplemented with this vitamin. Isolate RM6 also degraded 2,2′-dithiodibenzoic acid in medium containing sterilized supernatants from cultures of isolate RM1 grown on glucose or benzoate. Isolate RM6 was identified as a member of the genus Variovorax using the Biolog system and 16S rRNA gene analysis. Although the mechanism of disulfide metabolism could not be determined, benzoic acid was detected as a transient metabolite of 2,2′-dithiodibenzoic acid biodegradation by Variovorax sp. strain RM6. In pure culture, this isolate mineralized 2,2′-dithiodibenzoic acid, releasing 59% of the carbon as carbon dioxide and 88% of the sulfur as sulfate.     

The <Emphasis Type="Italic">cnr</Emphasis>-like operon in strain <Emphasis Type="Italic">Comamonas</Emphasis> sp. encoding resistance to cobalt and nickel

Plasmid pBS501 was detected in the strain Comamonas sp. BS501. This plasmid specifies high level of induced resistance (5 mM) to cobalt/nickel both in the host strain and in related strains C. testosteroni B-1241 and C. acidovorans B-1251. Hybridization analysis revealed a homology of pBS501 restriction fragments with the only well-characterized operon cnrXYHCBAT that resides in plasmid pMOL28 from Cupriavidus metallidurans CH34. Essential differences in the structural organization of the cobalt/nickel resistance determinant were found between plasmid pBS501 and the cnr operon.     

A simple method for detection of enzyme activities involved in the initial step of phthalate degradation in microorganisms

We have developed a simple method for the detection of phthalate 4,5-dioxygenase and 4,5-dihydro-4,5-dihydroxyphthalate dehydrogenase activities in the initial step of phthalate degradation in bacteria. It was found that cells of a Pseudomonas putida strain adapted for phthalate could convert quinolinic acid to a hydroxylated product having λ_max at 315 nm. The occurrence of this compound was visualized by reaction with diazotized p-nitroaniline with which a red compound having λ_max at 512 nm was produced. In practice, if cells in colonies developed on an agar plate containing mineral salt medium supplemented with 0.4% of disodium phthalate and 0.1% of quinolinic acid are active with respect to the 4,5-dihydroxyphthalate pathway, then the colonies would be colored red immediately upon spraying with the diazotized p-nitroaniline reagent. The method was used to identify the phthalate degradative pathway for 27 phthalate-utilizing strains of the genera Pseudomonas (18 strains), Agrobacterium (3 strains), Alcaligenes (5 strains), and Micrococcus (1 strain). It was found that 24 of the 26 Gram-negative bacteria have the 4,5-dihydroxyphthalate pathway and that the remaining two strains of Pseudomonas sp. may metabolize via an unidentified pathway other than the dihydroxyphthalate pathways, and the Gram-positive strain of Micrococcus sp. metabolizes phthalate via the 3,4-dihydroxyphthalate pathway.     

Isolation and Characterization of Diuron-degrading Bacteria from Lotic Surface Water

The bacterial community structure of a diuron-degrading enrichment culture from lotic surface water samples was analyzed and the diuron-degrading strains were selected using a series of techniques combining temporal temperature gradient gel electrophoresis (TTGE) of 16 S rDNA gene V1–V3 variable regions, isolation of strains on agar plates, colony hybridization methods, and biodegradation assays. The TTGE fingerprints revealed that diuron had a strong impact on bacterial community structure and highlighted both diuron-sensitive and diuron-adapted bacterial strains. Two bacterial strains, designated IB78 and IB93 and identified as belonging to Pseudomonas sp. and Stenotrophomonas sp., were isolated and shown to degrade diuron in pure resting cells in a first-order kinetic reaction during the first 24 h of incubation with no 3,4-DCA detected. The percentages of degradation varied from 25% to 60% for IB78 and 20% to 65% for IB93 and for a diuron concentration range from 20 mg/L to 2 mg/L, respectively. It is interesting to note that diuron was less degraded by single isolates than by mixed resting cells, thereby underlining a cumulative effect between these two strains. To the best of our knowledge, this is the first report of diuron-degrading strains isolated from lotic surface water.     

Mineralization of Paraoxon and Its Use as a Sole C and P Source by a Rationally Designed Catabolic Pathway in Pseudomonas putida

Organophosphate compounds, which are widely used as pesticides and chemical warfare agents, are cholinesterase inhibitors. These synthetic compounds are resistant to natural degradation and threaten the environment. We constructed a strain of Pseudomonas putida that can efficiently degrade a model organophosphate, paraoxon, and use it as a carbon, energy, and phosphorus source. This strain was engineered with the pnp operon from Pseudomonas sp. strain ENV2030, which encodes enzymes that transform p-nitrophenol into β-ketoadipate, and with a synthetic operon encoding an organophosphate hydrolase (encoded by opd) from Flavobacterium sp. strain ATCC 27551, a phosphodiesterase (encoded by pde) from Delftia acidovorans, and an alkaline phosphatase (encoded by phoA) from Pseudomonas aeruginosa HN854 under control of a constitutive promoter. The engineered strain can efficiently mineralize up to 1 mM (275 mg/liter) paraoxon within 48 h, using paraoxon as the sole carbon and phosphorus source and an inoculum optical density at 600 nm of 0.03. Because the organism can utilize paraoxon as a sole carbon, energy, and phosphorus source and because one of the intermediates in the pathway (p-nitrophenol) is toxic at high concentrations, there is no need for selection pressure to maintain the heterologous pathway.     

Genetics of naphthalene and phenanthrene degradation by Comamonas testosteroni

Naphthalene and phenanthrene have long been used as model compounds to investigate the ability of bacteria to degrade polycyclic aromatic hydrocarbons. The catabolic pathways have been determined, several of the enzymes have been purified to homogeneity, and genes have been cloned and sequenced. However, the majority of this work has been performed with fast growing Pseudomonas strains related to the archetypal naphthalene-degrading P. putida strains G7 and NCIB 9816-4. Recently Comamonas testosteroni strains able to degrade naphthalene and phenanthrene have been isolated and shown to possess genes for polycyclic aromatic hydrocarbon degradation that are different from the canonical genes found in Pseudomonas species. For instance, C. testosteroni GZ39 has genes for naphthalene and phenanthrene degradation which are not only different from those found in Pseudomonas species but are also arranged in a different configuration. C. testosteroni GZ42, on the other hand, has genes for naphthalene and phenanthrene degradation which are arranged almost the same as those found in Pseudomonas species but show significant divergence in their sequences. Received 10 August 1997/ Accepted in revised form 15 August 1997     

Rapid mineralization of the phenylurea herbicide diuron by Variovorax sp. strain SRS16 in pure culture and within a two-member consortium

The phenylurea herbicide diuron [N-(3,4-dichlorophenyl)-N,N-dimethylurea] is widely used in a broad range of herbicide formulations, and consequently, it is frequently detected as a major water contaminant in areas where there is extensive use. We constructed a linuron [N-(3,4-dichlorophenyl)-N-methoxy-N-methylurea]- and diuron-mineralizing two-member consortium by combining the cooperative degradation capacities of the diuron-degrading organism Arthrobacter globiformis strain D47 and the linuron-mineralizing organism Variovorax sp. strain SRS16. Neither of the strains mineralized diuron alone in a mineral medium, but combined, the two strains mineralized 31 to 62% of the added [ring-U-(14)C]diuron to (14)CO(2), depending on the initial diuron concentration and the cultivation conditions. The constructed consortium was used to initiate the degradation and mineralization of diuron in soil without natural attenuation potential. This approach led to the unexpected finding that Variovorax sp. strain SRS16 was able to mineralize diuron in a pure culture when it was supplemented with appropriate growth substrates, making this strain the first known bacterium capable of mineralizing diuron and representatives of both the N,N-dimethyl- and N-methoxy-N-methyl-substituted phenylurea herbicides. The ability of the coculture to mineralize microgram-per-liter levels of diuron was compared to the ability of strain SRS16 alone, which revealed the greater extent of mineralization by the two-member consortium (31 to 33% of the added [ring-U-(14)C]diuron was mineralized to (14)CO(2) when 15.5 to 38.9 mug liter(-1) diuron was used). These results suggest that the consortium consisting of strains SRS16 and D47 could be a promising candidate for remediation of soil and water contaminated with diuron and linuron and their shared metabolite 3,4-dichloroaniline.     

Efficient biotransformation of herbicide diuron by bacterial strain <Emphasis Type="Italic">Micrococcus</Emphasis> sp. PS-1

A Gram-positive, Micrococcus sp. strain PS-1 capable of utilizing phenylurea herbicide diuron as a sole carbon source at a high concentration (up to 250 ppm) was isolated from diuron storage site by selective enrichment study. The taxonomic characterization with 16S rRNA gene sequencing (1,477 bp) identified PS-1 as a member of Micrococcus sp. It was studied for the degradation of diuron and a range of its analogues (monuron, linuron, monolinuron, chlortoluron and fenuron). The shake flasks experiments demonstrated fast degradation of diuron (up to 96% at 250 ppm within 30 h incubation) with the addition of small quantity (0.01%) of non-ionic detergent. The relative degradation profile by the isolate was in the order of fenuron > monuron > diuron > linuron > monolinuron > chlortoluron. Further, the biochemical characterization of catabolic pathway by spectroscopic and chromatographic techniques demonstrated that the degradation proceeded via formation of dealkylated metabolites to form 3,4-dichloroaniline (3,4-DCA). It was the major metabolite formed, associated with profound increase in degradation kinetics in presence of appropriate additive.