Nitrous oxide inhibition of methanogenesis represents an underappreciated greenhouse gas emission feedback.

Methane (CH4) and nitrous oxide (N2O) are major greenhouse gases that are predominantly generated by microbial activities in anoxic environments. N2O inhibition of methanogenesis has been reported, but comprehensive efforts to obtain kinetic information are lacking. Using the model methanogen Methanosarcina barkeri strain Fusaro and digester sludge-derived methanogenic enrichment cultures, we conducted growth yield and kinetic measurements and showed that micromolar concentrations of N2O suppress the growth of methanogens and CH4 production from major methanogenic substrate classes. Acetoclastic methanogenesis, estimated to account for two-thirds of the annual 1 billion metric tons of biogenic CH4, was most sensitive to N2O, with inhibitory constants (KI) in the range of 18-25 µM, followed by hydrogenotrophic (KI, 60-90 µM) and methylotrophic (KI, 110-130 µM) methanogenesis. Dissolved N2O concentrations exceeding these KI values are not uncommon in managed (i.e. fertilized soils and wastewater treatment plants) and unmanaged ecosystems. Future greenhouse gas emissions remain uncertain, particularly from critical zone environments (e.g. thawing permafrost) with large amounts of stored nitrogenous and carbonaceous materials that are experiencing unprecedented warming. Incorporating relevant feedback effects, such as the significant N2O inhibition on methanogenesis, can refine climate models and improve predictive capabilities.

Integrated Advanced Molecular Tools Predict In Situ cVOC Degradation Rates: Field Demonstration.

Chlorinated volatile organic compound (cVOC) degradation rate constants are crucial information for site management. Conventional approaches generate rate estimates from the monitoring and modeling of cVOC concentrations. This requires time series data collected along the flow path of the plume. The estimates of rate constants are often plagued by confounding issues, making predictions cumbersome and unreliable. Laboratory data suggest that targeted quantitative analysis of Dehalococcoides mccartyi (Dhc) biomarker genes (qPCR) and proteins (qProt) can be directly correlated with reductive dechlorination activity. To assess the potential of qPCR and qProt measurements to predict rates, we collected data from cVOC-contaminated aquifers. At the benchmark study site, the rate constant for degradation of cis-dichloroethene (cDCE) extracted from monitoring data was 11.0 ± 3.4 yr-1, and the rate constant predicted from the abundance of TceA peptides was 6.9 yr-1. The rate constant for degradation of vinyl chloride (VC) from monitoring data was 8.4 ± 5.7 yr-1, and the rate constant predicted from the abundance of TceA peptides was 5.2 yr-1. At the other study sites, the rate constants for cDCE degradation predicted from qPCR and qProt measurements agreed within a factor of 4. Under the right circumstances, qPCR and qProt measurements can be useful to rapidly predict rates of cDCE and VC biodegradation, providing a major advance in effective site management.

Dehalogenation of Chlorinated Ethenes to Ethene by a Novel Isolate, "Candidatus Dehalogenimonas etheniformans".

Dehalococcoides mccartyi strains harboring vinyl chloride (VC) reductive dehalogenase (RDase) genes are keystone bacteria for VC detoxification in groundwater aquifers, and bioremediation monitoring regimens focus on D. mccartyi biomarkers. We isolated a novel anaerobic bacterium, "Candidatus Dehalogenimonas etheniformans" strain GP, capable of respiratory dechlorination of VC to ethene. This bacterium couples formate and hydrogen (H2) oxidation to the reduction of trichloro-ethene (TCE), all dichloroethene (DCE) isomers, and VC with acetate as the carbon source. Cultures that received formate and H2 consumed the two electron donors concomitantly at similar rates. A 16S rRNA gene-targeted quantitative PCR (qPCR) assay measured growth yields of (1.2 ± 0.2) × 108 and (1.9 ± 0.2) × 108 cells per µmol of VC dechlorinated in cultures with H2 or formate as electron donor, respectively. About 1.5-fold higher cell numbers were measured with qPCR targeting cerA, a single-copy gene encoding a putative VC RDase. A VC dechlorination rate of 215 ± 40 µmol L-1 day-1 was measured at 30°C, with about 25% of this activity occurring at 15°C. Increasing NaCl concentrations progressively impacted VC dechlorination rates, and dechlorination ceased at 15 g NaCl L-1. During growth with TCE, all DCE isomers were intermediates. Tetrachloroethene was not dechlorinated and inhibited dechlorination of other chlorinated ethenes. Carbon monoxide formed and accumulated as a metabolic by-product in dechlorinating cultures and impacted reductive dechlorination activity. The isolation of a new Dehalogenimonas species able to effectively dechlorinate toxic chlorinated ethenes to benign ethene expands our understanding of the reductive dechlorination process, with implications for bioremediation and environmental monitoring. IMPORTANCE Chlorinated ethenes are risk drivers at many contaminated sites, and current bioremediation efforts focus on organohalide-respiring Dehalococcoides mccartyi strains to achieve detoxification. We isolated and characterized the first non-Dehalococcoides bacterium, "Candidatus Dehalogenimonas etheniformans" strain GP, capable of metabolic reductive dechlorination of TCE, all DCE isomers, and VC to environmentally benign ethene. In addition to hydrogen, the new isolate utilizes formate as electron donor for reductive dechlorination, providing opportunities for more effective electron donor delivery to the contaminated subsurface. The discovery that a broader microbial diversity can achieve detoxification of toxic chlorinated ethenes in anoxic aquifers illustrates the potential of naturally occurring microbes for biotechnological applications.

Identification and widespread environmental distribution of a gene cassette implicated in anaerobic dichloromethane degradation.

Anthropogenic activities and natural processes release dichloromethane (DCM, methylene chloride), a toxic chemical with substantial ozone-depleting capacity. Specialized anaerobic bacteria metabolize DCM; however, the genetic basis for this process has remained elusive. Comparative genomics of the three known anaerobic DCM-degrading bacterial species revealed a homologous gene cluster, designated the methylene chloride catabolism (mec) gene cassette, comprising 8-10 genes encoding proteins with 79.6%-99.7% amino acid identities. Functional annotation identified genes encoding a corrinoid-dependent methyltransferase system, and shotgun proteomics applied to two DCM-catabolizing cultures revealed high expression of proteins encoded on the mec gene cluster during anaerobic growth with DCM. In a DCM-contaminated groundwater plume, the abundance of mec genes strongly correlated with DCM concentrations (R2  = 0.71-0.85) indicating their potential value as process-specific bioremediation biomarkers. mec gene clusters were identified in metagenomes representing peat bogs, the deep subsurface, and marine ecosystems including oxygen minimum zones (OMZs), suggesting a capacity for DCM degradation in diverse habitats. The broad distribution of anaerobic DCM catabolic potential infers a role for DCM as an energy source in various environmental systems, and implies that the global DCM flux (i.e., the rate of formation minus the rate of consumption) might be greater than emission measurements suggest.

Draft Genome Sequence of Shewanella sp. Strain BF02_Schw, Isolated from Blood Falls, a Feature Where Subglacial Brine Discharges to the Surface of Taylor Glacier, Antarctica.

We report the sequencing, assembly, and draft genome of Shewanella sp. strain BF02_Schw. The assembly contains 5,304,243 bp, with a GC content of 41.43%.

Isolation and characterization of a halophilic Modicisalibacter sp. strain Wilcox from produced water.

We report the isolation a halophilic bacterium that degrades both aromatic and aliphatic hydrocarbons as the sole sources of carbon at high salinity from produced water. Phylogenetic analysis of 16S rRNA-gene sequences shows the isolate is a close relative of Modicisalibacter tunisiensis isolated from an oil-field water in Tunisia. We designate our isolate as Modicisalibacter sp. strain Wilcox. Genome analysis of strain Wilcox revealed the presence of a repertoire of genes involved in the metabolism of aliphatic and aromatic hydrocarbons. Laboratory culture studies corroborated the predicted hydrocarbon degradation potential. The strain degraded benzene, toluene, ethylbenzene, and xylenes at salinities ranging from 0.016 to 4.0 M NaCl, with optimal degradation at 1 M NaCl. Also, the strain degraded phenol, benzoate, biphenyl and phenylacetate as the sole sources of carbon at 2.5 M NaCl. Among aliphatic compounds, the strain degraded n-decane and n-hexadecane as the sole sources of carbon at 2.5 M NaCl. Genome analysis also predicted the presence of many heavy metal resistance genes including genes for metal efflux pumps, transport proteins, and enzymatic detoxification. Overall, due to its ability to degrade many hydrocarbons and withstand high salt and heavy metals, strain Wilcox may prove useful for remediation of produced waters.

Metagenome-Guided Proteomic Quantification of Reductive Dehalogenases in the Dehalococcoides mccartyi-Containing Consortium SDC-9.

At groundwater sites contaminated with chlorinated ethenes, fermentable substrates are often added to promote reductive dehalogenation by indigenous or augmented microorganisms. Contemporary bioremediation performance monitoring relies on nucleic acid biomarkers of key organohalide-respiring bacteria, such as Dehalococcoides mccartyi (Dhc). Metagenome sequencing of the commercial, Dhc-containing consortium, SDC-9, identified 12 reductive dehalogenase (RDase) genes, including pceA (two copies), vcrA, and tceA, and allowed for specific detection and quantification of RDase peptides using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Shotgun (i.e., untargeted) proteomics applied to the SDC-9 consortium grown with tetrachloroethene (PCE) and lactate identified 143 RDase peptides, and 36 distinct peptides that covered greater than 99% of the protein-coding sequences of the PceA, TceA, and VcrA RDases. Quantification of RDase peptides using multiple reaction monitoring (MRM) assays with 13C-/15N-labeled peptides determined 1.8 × 103 TceA and 1.2 × 102 VcrA RDase molecules per Dhc cell. The MRM mass spectrometry approach allowed for sensitive detection and accurate quantification of relevant Dhc RDases and has potential utility in bioremediation monitoring regimes.

Biotransformation of the Fluorinated Nonsteroidal Anti-Inflammatory Pharmaceutical Flurbiprofen in Activated Sludge Results in Accumulation of a Recalcitrant Fluorinated Aromatic Metabolite.

Flurbiprofen is a fluorinated, nonsteroidal, anti-inflammatory pharmaceutical with potential application in a wide range of maladies. Currently, there is no information regarding its environmental fate. To address this, flurbiprofen is spiked at 500 and 50 ppm into activated sewage sludge taken from the municipal treatment plant of Ankara, Turkey. Flurbiprofen is partially degraded after 80 days, with removal proportion varying from 33% to 48%. Isolation of organisms able to use flurbiprofen as a sole carbon and energy source is unsuccessful. A transient, acid-labile yellow coloration appears in supernatants after addition of flurbiprofen. During disappearance, a novel potential metabolite is detected by high-performance liquid chromatography (HPLC) analyses, a chemical that does not appear in killed controls or in nonflurbiprofen-amended controls. Mass spectra of the novel chemical obtained at low and high collision energies are consistent with 4-(1-carboxyethyl)-2-fluorobenzoic acid, suggesting the application of a canonical metabolic paradigm for halogenated biphenyl metabolism by bacteria in which the nonhalogenated ring is metabolized by dioxygenation and metacleavage, leaving the halogenated aromatic ring behind. This metabolite shows no signs of disappearance after the 80-day monitoring period, implying that the environmental release of flurbiprofen might be of concern.

Metagenomes from Coastal Marine Sediments Give Insights into the Ecological Role and Cellular Features of Loki- and Thorarchaeota.

The genomes of Asgard Archaea, a novel archaeal proposed superphylum, share an enriched repertoire of eukaryotic signature genes and thus promise to provide insights into early eukaryote evolution. However, the distribution, metabolisms, cellular structures, and ecology of the members within this superphylum are not well understood. Here we provide a meta-analysis of the environmental distribution of the Asgard archaea, based on available 16S rRNA gene sequences. Metagenome sequencing of samples from a salt-crusted lagoon on the Baja California Peninsula of Mexico allowed the assembly of a new Thorarchaeota and three Lokiarchaeota genomes. Comparative analyses of all known Lokiarchaeota and Thorarchaeota genomes revealed overlapping genome content, including central carbon metabolism. Members of both groups contained putative reductive dehalogenase genes, suggesting that these organisms might be able to metabolize halogenated organic compounds. Unlike the first report on Lokiarchaeota, we identified genes encoding glycerol-1-phosphate dehydrogenase in all Loki- and Thorarchaeota genomes, suggesting that these organisms are able to synthesize bona fide archaeal lipids with their characteristic glycerol stereochemistry.IMPORTANCE Microorganisms of the superphylum Asgard Archaea are considered to be the closest living prokaryotic relatives of eukaryotes (including plants and animals) and thus promise to give insights into the early evolution of more complex life forms. However, very little is known about their biology as none of the organisms has yet been cultivated in the laboratory. Here we report on the ecological distribution of Asgard Archaea and on four newly sequenced genomes of the Lokiarchaeota and Thorarchaeota lineages that give insight into possible metabolic features that might eventually help to identify these enigmatic groups of archaea in the environment and to culture them.

Proteogenomics Reveals Novel Reductive Dehalogenases and Methyltransferases Expressed during Anaerobic Dichloromethane Metabolism.

Dichloromethane (DCM) is susceptible to microbial degradation under anoxic conditions and is metabolized via the Wood-Ljungdahl pathway; however, mechanistic understanding of carbon-chlorine bond cleavage is lacking. The microbial consortium RM contains the DCM degrader "Candidatus Dichloromethanomonas elyunquensis" strain RM, which strictly requires DCM as a growth substrate. Proteomic workflows applied to DCM-grown consortium RM biomass revealed a total of 1,705 nonredundant proteins, 521 of which could be assigned to strain RM. In the presence of DCM, strain RM expressed a complete set of Wood-Ljungdahl pathway enzymes, as well as proteins implicated in chemotaxis, motility, sporulation, and vitamin/cofactor synthesis. Four corrinoid-dependent methyltransferases were among the most abundant proteins. Notably, two of three putative reductive dehalogenases (RDases) encoded within strain RM's genome were also detected in high abundance. Expressed RDase 1 and RDase 2 shared 30% amino acid identity, and RDase 1 was most similar to an RDase of Dehalococcoides mccartyi strain WBC-2 (AOV99960, 52% amino acid identity), while RDase 2 was most similar to an RDase of Dehalobacter sp. strain UNSWDHB (EQB22800, 72% amino acid identity). Although the involvement of RDases in anaerobic DCM metabolism has yet to be experimentally verified, the proteome characterization results implicated the possible participation of one or more reductive dechlorination steps and methyl group transfer reactions, leading to a revised proposal for an anaerobic DCM degradation pathway.IMPORTANCE Naturally produced and anthropogenically released DCM can reside in anoxic environments, yet little is known about the diversity of organisms, enzymes, and mechanisms involved in carbon-chlorine bond cleavage in the absence of oxygen. A proteogenomic approach identified two RDases and four corrinoid-dependent methyltransferases expressed by the DCM degrader "Candidatus Dichloromethanomonas elyunquensis" strain RM, suggesting that reductive dechlorination and methyl group transfer play roles in anaerobic DCM degradation. These findings suggest that the characterized DCM-degrading bacterium Dehalobacterium formicoaceticum and "Candidatus Dichloromethanomonas elyunquensis" strain RM utilize distinct strategies for carbon-chlorine bond cleavage, indicating that multiple pathways evolved for anaerobic DCM metabolism. The specific proteins (e.g., RDases and methyltransferases) identified in strain RM may have value as biomarkers for monitoring anaerobic DCM degradation in natural and contaminated environments.

Normalized Quantitative PCR Measurements as Predictors for Ethene Formation at Sites Impacted with Chlorinated Ethenes.

Quantitative PCR (qPCR) targeting Dehalococcoides mccartyi ( Dhc) biomarker genes supports effective management at sites impacted with chlorinated ethenes. To establish correlations between Dhc biomarker gene abundances and ethene formation (i.e., detoxification), 859 groundwater samples representing 62 sites undergoing monitored natural attenuation or enhanced remediation were analyzed. Dhc 16S rRNA genes and the vinyl chloride (VC) reductive dehalogenase genes bvcA and vcrA were detected in 88% and 61% of samples, respectively, from wells with ethene. Dhc 16S rRNA, bvcA, vcrA, and tceA (implicated in cometabolic reductive VC dechlorination) gene abundances all positively correlated with ethene formation. Significantly greater ethene concentrations were observed when Dhc 16S rRNA gene and VC RDase gene abundances exceeded 107 and 106 copies L-1, respectively, and when Dhc 16S rRNA- and bvcA + vcrA-to-total bacterial 16S rRNA gene ratios exceeded 0.1%. Dhc 16S rRNA gene-to- vcrA/ bvcA ratios near unity also indicated elevated ethene; however, no increased ethene was observed in 19 wells where vcrA and/or bvcA gene copy numbers exceeded Dhc cell numbers 10- to 10 000-fold. Approximately one-third of samples with detectable ethene lacked bvcA, vcrA, and tceA, suggesting that comprehensive understanding of VC detoxification biomarkers has not been achieved. Although the current biomarker suite is incomplete, the data analysis corroborates the value of the available Dhc DNA biomarkers for prognostic and diagnostic groundwater monitoring at sites impacted with chlorinated ethenes.

Complete Genome Sequence of Dehalobacterium formicoaceticum Strain DMC, a Strictly Anaerobic Dichloromethane-Degrading Bacterium.

Dehalobacterium formicoaceticum utilizes dichloromethane as the sole energy source in defined anoxic bicarbonate-buffered mineral salt medium. The products are formate, acetate, inorganic chloride, and biomass. The bacterium's genome was sequenced using PacBio, assembled, and annotated. The complete genome consists of one 3.77-Mb circular chromosome harboring 3,935 predicted protein-encoding genes.

The biotransformation of ibuprofen to trihydroxyibuprofen in activated sludge and by Variovorax Ibu-1.

A bacterium was isolated from activated sewage sludge that has the ability to use ibuprofen as its sole carbon and energy source. Phylogenetic analysis of the 16S rRNA gene sequence placed the strain in the Variovorax genus within the ß-proteobacteria. When grown on ibuprofen it accumulated a transient yellow intermediate that disappeared upon acidification, a trait consistent with meta ring-fission metabolites. GC/MS analysis of derivatized culture supernatant yielded two spectra consistent with trihydroxyibuprofen bearing all three hydroxyl groups on the aromatic ring. These metabolites were only detected when 3-fluorocatechol, a meta ring-fission inhibitor, was added to Ibu-1 cultures and the supernatant was then derivatized with aqueous acetic anhydride and diazomethane. These findings suggest the possibility of ibuprofen metabolism proceeding via a trihydroxyibuprofen meta ring-fission pathway. Identical spectra, consistent with these putative ring-hydroxylated trihydroxyibuprofen metabolites, were also obtained from ibuprofen-spiked sewage sludge, but only when it was poisoned with 3-fluorocatechol. The presence of the same trihydroxylated metabolites in both spiked sewage sludge and culture supernatants suggests that this trihydroxyibuprofen extradiol ring-cleavage pathway for the degradation of ibuprofen may have environmental relevance.

Diversity and community analysis of ammonia oxidizing bacteria in a streambed surrounding an artificial dam.

The degree to which small natural dams affect the native bacterial nitrogen cycling community was explored by molecular methods. The identities and relative abundances of ammonia oxidizing bacteria in the sediment surrounding an artificial dam both at the surface and in the hyporheic zone were characterized. Analyses were performed using tRFLP of the conserved amoA gene using a semi-nested degenerate PCR approach. Additionally, an amoA gene library was constructed to characterize the most dominant sediment genotypes. The results of the tRFLP analyses showed clear differences between the upstream and downstream communities at different depths in the sediment column. Non-metric multidimensional scaling ordination of the tRFLP data set produced a stable one-dimensional solution with significant correlations to oxygen, pH, nitrate, and dissolved organic nitrogen levels. The sample corresponding to the hyporheic zone downstream of the dam showed 28-50% higher amoA richness and higher diversity than the other samples. All gene fragments sequenced from the samples grouped with sequences of the Nitrosospira type. Ordination of 16S rDNA tRFLP data revealed a two dimensional data structure, one axis of which had similar chemical correlation characteristics as the amoA model axis. Taken together, the results from this study suggest that the presence of the dam creates physical and chemical heterogeneity that may foster genetic diversity and community changes amongst ammonia oxidizing bacteria.

Genetic and chemical characterization of ibuprofen degradation by Sphingomonas Ibu-2.

Sphingomonas Ibu-2 has the unusual ability to cleave the acid side chain from the pharmaceutical ibuprofen and related arylacetic acid derivatives to yield corresponding catechols under aerobic conditions via a previously uncharacterized mechanism. Screening a chromosomal library of Ibu-2 DNA in Escherichia coli EPI300 allowed us to identify one fosmid clone (pFOS3G7) that conferred the ability to metabolize ibuprofen to isobutylcatechol. Characterization of pFOS3G7 loss-of-function transposon mutants permitted identification of five ORFs, ipfABDEF, whose predicted amino acid sequences bore similarity to the large and small units of an aromatic dioxygenase (ipfAB), a sterol carrier protein X (SCPx) thiolase (ipfD), a domain of unknown function 35 (DUF35) protein (ipfE) and an aromatic CoA ligase (ipfF). Two additional ORFs, ipfH and ipfI, which encode putative ferredoxin reductase and ferredoxin components of an aromatic dioxygenase system, respectively, were also identified on pFOS3G7. Complementation of a markerless loss-of-function ipfD deletion mutant restored catechol production as did complementation of the ipfF Tn mutant. Expression of subcloned ipfABDEF alone in E. coli did not impart full metabolic activity unless coexpressed with ipfHI. CoA ligation followed by ring oxidation is common to phenylacetic acid pathways. However, the need for a putative SCPx thiolase (IpfD) and DUF35 protein (IpfE) in aerobic arylacetic acid degradation is unprecedented. This work provides preliminary insights into the mechanism behind this novel arylacetic acid-deacylating, catechol-generating activity.

Biodegradation of pharmaceutical and personal care products.

Medical treatments and personal hygiene lead to the steady release of pharmaceutical and personal care products (PPCPs) into the environment. Some of these PPCPs have been shown to have detrimental environmental effects and could potentially impact human health. Understanding the biological transformation of PPCPs is essential for accurately determining their ultimate environmental fate, conducting accurate risk assessments, and improving PPCP removal. We summarize the current literature concerning the biological transformation of PPCPs in wastewater treatment plants, the environment, and by pure cultures of bacterial isolates. Although some PPCPs, such as ibuprofen, are readily degraded under most studied conditions, others, such as carbamazepine, tend to be recalcitrant. This variation in the biodegradability of PPCPs can be attributed to structural differences, because PPCPs are classified by application, not chemical structure. The degradation pathways of octylphenol by Sphingomonas sp. strain PWE1, ibuprofen by Sphingomonas sp. strain Ibu-2, and DEET by Pseudomonas putida DTB are discussed in more detail.

Laboratory and field evaluation of crystallized DOW 704 oil on the performance of the Well Impactor Ninety-Six fFine particulate matter fractionator.

Subsequent to the 1997 promulgation of the Federal Reference Method (FRM) for monitoring fine particulate matter (PM2.5) in ambient air, U.S. Environmental Protection Agency (EPA) received reports that the DOW 704 diffusion oil used in the method's Well Impactor Ninety-Six (WINS) fractionator would occasionally crystallize during field use, particularly under wintertime conditions. Although the frequency of occurrence on a nationwide basis was low, uncertainties existed as to whether crystallization of the DOW 704 oil may adversely affect a sampling event's data quality. In response to these concerns, EPA and the State of Connecticut Department of Environmental Protection jointly conducted a series of specialized tests to determine whether crystallized oil adversely affected the performance of the WINS fractionator. In the laboratory, an experimental setup used dry ice to artificially induce crystallization of the diffusion oil under controlled conditions. Using primary polystyrene latex calibration aerosols, standard size-selective performance tests of the WINS fractionator showed that neither the position nor the shape of the WINS particle size fractionation curve was substantially influenced by the crystallization of the DOW 704 oil. No large particle bounce from the crystallized impaction surface was observed. During wintertime field tests, crystallization of the DOW 704 oil did not adversely affect measured PM2.5 concentrations. Regression of measurements with crystallized DOW 704 versus liquid dioctyl sebacate (DOS) oil produced slope, intercept, and R2 values of 0.98, 0.1, and 0.997 microg/m3, respectively. Additional field tests validated the use of DOS as an effective impaction substrate. As a result of these laboratory and field tests, DOS oil has been approved by EPA as a substitute for DOW 704 oil. Since the field deployment of DOS oil in 2001, users of this alternative oil have not reported any operational problems associated with its use in the PM2.5 FRM. Limited field evaluation of the BGI very sharp cut cyclone indicates that it provides a viable alternative to the WINS fractionator.

Formation of catechols via removal of acid side chains from ibuprofen and related aromatic acids.

Although ibuprofen [2-(4-isobutylphenyl)-propionic acid] is one of the most widely consumed drugs in the world, little is known regarding its degradation by environmental bacteria. Sphingomonas sp. strain Ibu-2 was isolated from a wastewater treatment plant based on its ability to use ibuprofen as a sole carbon and energy source. A slight preference toward the R enantiomer was observed, though both ibuprofen enantiomers were metabolized. A yellow color, indicative of meta-cleavage, accumulated transiently in the culture supernatant when Ibu-2 was grown on ibuprofen. When and only when 3-flurocatechol was used to poison the meta-cleavage system, isobutylcatechol was identified in the culture supernatant via gas chromatography-mass spectrometry analysis. Ibuprofen-induced washed-cell suspensions also metabolized phenylacetic acid and 2-phenylpropionic acid to catechol, while 3- and 4-tolylacetic acids and 2-(4-tolyl)-propionic acid were metabolized to the corresponding methyl catechols before ring cleavage. These data suggest that, in contrast to the widely distributed coenzyme A ligase, homogentisate, or homoprotocatechuate pathway for metabolism of phenylacetic acid and similar compounds, Ibu-2 removes the acidic side chain of ibuprofen and related compounds prior to ring cleavage.