Aerobic Enrichment Cultures Research Articles

Metagenomic analyses of aerobic bacterial enrichment cultures that degraded Tris(2-chloroethyl) phosphate (TCEP) and its transformation products

Tris(2-chloroethyl) phosphate (TCEP) is of growing public concern worldwide because of its ubiquitous contamination, toxicity, and persistence. In this study, we investigated bacterial communities in aerobic enrichment cultures with TCEP and its two major transformation products bis(2-chloroethyl) phosphate (BCEP) and 2-chloroethanol (2-CE) as the sole carbon source. Burkholderiales and Rhizobiales were likely two main bacterial guilds involved in the hydrolysis of TCEP, while Rhizobiales and Sphingomonadales may play an important role in the hydrolysis of BCEP, given the increase of Rhizobiales and Sphingomonadales-related phosphoesterase genes when the carbon source was switched from TCEP to BCEP. All Burkholderiales, Rhizobiales, Sphingomonadales were probably efficient in 2-CE metabolism, because their dehydrogenase genes and dehalogenase genes increased after 2-CE enrichment. The different substrate preference of different bacterial guilds highlighted the importance to understand the diversity and collaboration among functional bacteria. Meanwhile, two TCEP-degrading strains affiliated with Xanthobacter and Ancylobacter were isolated. Xanthobacter sp. strain T2-1 was able to degrade both TCEP and BCEP following the pseudo-first-order kinetics with reaction rates of 1.66 h−1 for TCEP and 1.02 h−1 for BCEP, respectively. Ancylobacter sp. strain T3-4 could degrade TCEP following the pseudo-first-order kinetics with a reaction rate of 2.54 h−1, but is unable to degrade BCEP. Additionally, strains that were phylogenetically closely related were found to have different degradation capabilities toward TCEP and/or BCEP, indicating the importance to investigate functional genes such as phosphoesterase genes.

Aerobic and anaerobic enrichment cultures highlight the pivotal role of facultative anaerobes in soil hydrocarbon degradation

Aliphatic and aromatic hydrocarbons are ubiquitous in the environment due to natural and anthropogenic processes. Under aerobic conditions hydrocarbons can be rapidly biodegraded but oxygenated environments often quickly become anaerobic when microbial respiration is coupled to contaminant oxidation. Most studies in literature usually focus on the initial microbial diversity of the hydrocarbon impacted environment and examine either aerobic or anaerobic conditions for enrichment. Hence, the aim of the present study was to enrich bacterial consortiums from two diesel impacted soil samples under both these conditions to assess the enrichment diversities and hydrocarbon degradation potentials. This would shed light upon how an environmental population shift would correlate to oxygen intrusion and depletion and still continue hydrocarbon degradation. Analysis of the 16S rRNA gene sequences showcases the different microbial populations that could emerge as the environmental factors change, resulting in different populations that are still capable of hydrocarbon degradation. Microbial diversity analysis also highlights the role of facultative anaerobic bacteria like Pseudomonas spp. and Citrobacter spp. in maintaining hydrocarbon degradation. This study shows that microorganisms capable of surviving under both oxic and anoxic (aerobic and anaerobic) conditions are the most crucial to the long term degradation of hydrocarbons in the environment.

Isolation of a novel Co2+-resistant bacterium and the application of its siderophore in Co2+ recovery from an aqueous solution.

Siderophores are considered to have a good potential as decontamination agents owing to their metal-chelating abilities. In order to confirm whether siderophores can be used in the recovery of metal ions, a siderophore (or metallophore) exhibiting Co2+-chelating activity was screened to demonstrate its ability to recover Co2+ from an aqueous solution. A siderophore-producing bacterium, Pandoraea sp. HCo-4B, was identified from a screen of Co2+-resistant bacteria grown in an aerobic enrichment culture with a Co2+-supplemented medium. After incubation of the crude extracted siderophore in a Co2+-containing solution, the Co2+-siderophore complex was adsorbed on to a C18 column. The bound Co2+ was eluted from the column by the addition of 10 mM H2SO4. The recovered amount of Co2+ was proportional to the amount of the added siderophore. We observed that the siderophore identified in this study binds to Co2+ at a 1:1 ratio.

Effects of dissolved organic matter and nitrification on biodegradation of pharmaceuticals in aerobic enrichment cultures

Natural dissolved organic matter (DOM) and nitrification can play an important role in biodegradation of pharmaceutically active compounds (PhACs) in aerobic zones of constructed wetlands (CWs). This study used an enrichment culture originating from CW sediment to study the effect of DOM and nitrification on aerobic biodegradation of seven PhACs. The enriched culture degraded caffeine (CAF), metoprolol (MET), naproxen (NAP), and ibuprofen (IBP) with a consistent biodegradability order of CAF>MET>NAP>IBP. Biodegradation of propranolol, carbamazepine, and diclofenac was insignificant (<15%). CAF biodegradation was inhibited by the easily biodegradable DOM. Conversely, DOM enhanced biodegradation of MET, NAP, and IBP, potentially by contributing more biomass capable of degrading PhACs. Nitrification enhanced biodegradation of NAP and IBP and mineralization of the PhAC mixture as well as less biodegradable DOM, which may result from co-metabolism of ammonia oxidizing bacteria or enhanced heterotrophic microbial activity under nitrification. MET biodegradation was inhibited in the presence of nitrification. DOM and nitrification effects on PhAC biodegradation in CWs gained from this study can be used in strategies to improve CW operation, namely: designing hydraulic retention times based on the biodegradability order of specific PhACs; applying DOM amendment; and introducing consistent ammonium streams to increase removal of PhACs of interest.

Microbial Processes Mediating the Evolution of Methylarsine Gases from Dimethylarsenate in Paddy Soils.

Arsenic (As) biovolatilization is an important component of the global As biogeochemical cycle. Soils can emit various methylarsine gases, but the underlying microbial processes remain unclear. Here, we show that the addition of molybdate (Mo), an inhibitor of sulfate-reducing bacteria, greatly enhanced dimethylarsine evolution from dimethylarsenate [DMAs(V)] added to two paddy soils. Molybdate addition significantly affected the microbial community structure. The aerobic enrichment cultures from both soils volatilized substantial amounts of dimethylarsine from DMAs(V) in the presence of Mo, whereas the anaerobic enrichment cultures did not. A Bacillus strain (CZ-2) capable of reducing DMAs(V) to dimethylarsine was isolated from the aerobic enrichment culture, and its volatilization ability was enhanced by Mo. RNA-seq analysis identified 10 reductase genes upregulated by Mo. Addition of the reducing agent NADH increased dimethylarsine volatilization by strain CZ-2, suggesting that DMAs(V) reductase is an NADH-dependent enzyme. The strain could not methylate arsenite or convert monomethylarsenate and DMAs(V) to trimethylarsine. Our results show that dimethylarsine evolution from DMAs(V) is independent of the As methylation pathway and that Mo enhances dimethylarsine evolution from paddy soils by shifting the microbial community structure and enhancing the reduction of DMAs(V) to dimethylarsine, possibly through upregulating the expression of DMAs(V) reductase gene(s).

Effect of Anaerobic Granular Sludge in Degradation of Two Azo dyes

The main theme to conduct present research work was to evaluate the effect of anaerobic granular sludge in degradation of two azo dyes. The study was evaluated in batch experiments where anaerobic and aerobic conditions were integrated by exposing anaerobic granular sludge to oxygen. Under these conditions, the azo dyes were reduced, resulting in a temporal accumulation of aromatic amines. Subsequently, aniline was degraded further in the presence of oxygen by the facultative aerobic bacteria present in the anaerobic granular sludge. Acid Orange and Reactive Black were also degraded, if inoculum from aerobic enrichment cultures were added to the batch experiments. Due to rapid autoxidation of Acid Orange, no enrichment culture could be established for this compound. The results of this study indicate that aerobic enrichment cultures developed on aromatic amines combined with oxygen tolerant anaerobic granular sludge can potentially be used to completely biodegrade azo dyes under integrated anaerobic/aerobic conditions.

Molecular analysis of atrazine-degrading bacteria and catabolic genes in the water column and sediment of a created wetland in an agricultural/urban watershed

Abstract Atrazine-mineralizing bacteria were enriched from water and sediment samples from a constructed flow-through wetland that received water from a river with an agricultural watershed. Bio–Sep beads laden with atrazine were submerged in the wetland sediment and water column in an attempt to sorb atrazine-metabolizing microbes on beads. Biometer experiments with in-situ incubated beads did not detect 14CO2 formation from 14C-atrazine. HPLC analysis was used to rank atrazine biodegradation by enrichment cultures from wetland samples. Analysis of the 16S rRNA gene sequences of 60 clones from aerobic enrichment cultures revealed the predominance of the genera Acidovorax (33%), Ralstonia (22%), Azorhizobium (12%), and Xanthobacter (12%) with sequence match ≥0.90 in wetland samples. Similar analysis of 12 clones from anaerobic enrichment cultures revealed the predominance of the genera Hydrogenophaga (42%) and Rhizobium (25%). Pure culture isolates (n = 18) were primarily Arthrobacter spp. (89%) at sequence match ≥0.97. Pure cultures were screened qualitatively for the presence of 8 catabolic genes associated with known pathways of atrazine biodegradation. None of the cultures had all Atz pathway genes and none contained atzD and trzD genes. AtzC was found in all isolates. The data indicate that the diversity of atrazine degraders was comparable to findings in agricultural soil studies, with the notable cluster of Arthrobacter spp. which may be favored by the culture conditions used for isolation.

Microbial Degradation of Acid Orange and Reactive Black in Presence of Anaerobic Granular Sludge

This study mainly focuses on the biodegradation of two azo dyes, Acid Orange and Reactive Black, was evaluated in batch experiments where anaerobic and aerobic conditions were integrated by exposing anaerobic granular sludge to oxygen. Under these conditions, the azo dyes were reduced, resulting in a temporal accumulation of aromatic amines. Subsequently, aniline was degraded further in the presence of oxygen by the facultative aerobic bacteria present in the anaerobic granular sludge. Acid Orange and Reactive Black were also degraded, if inoculum from aerobic enrichment cultures were added to the batch experiments. Due to rapid autoxidation of Acid Orange, no enrichment culture could be established for this compound. The results of this study indicate that aerobic enrichment cultures developed on aromatic amines combined with oxygen tolerant anaerobic granular sludge can potentially be used to completely biodegrade azo dyes under integrated anaerobic/aerobic conditions.

Ethyl tert-butyl ether (ETBE)-degrading microbial communities in enrichments from polluted environments

The ethyl tert-butyl ether (ETBE) degradation capacity and phylogenetic composition of five aerobic enrichment cultures with ETBE as the sole carbon and energy source were studied. In all cases, ETBE was entirely degraded to biomass and CO2. Clone libraries of the 16S rRNA gene were prepared from each enrichment. The analyses of the DNA sequences obtained showed different taxonomic compositions with a majority of Proteobacteria in three cases. The two other enrichments have different microbiota with an abundance of Acidobacteria in one case, whereas the microbiota in the second was more diverse (majority of Actinobacteria, Chlorobi and Gemmatimonadetes). Actinobacteria were detected in all five enrichments. Several bacterial strains were isolated from the enrichments and five were capable of degrading ETBE and/or tert-butyl alcohol (TBA), a degradation intermediate. The five included three Rhodococcus sp. (IFP 2040, IFP 2041, IFP 2043), one Betaproteobacteria (IFP 2047) belonging to the Rubrivivax/Leptothrix/Ideonella branch, and one Pseudonocardia sp. (IFP 2050). Quantification of these five strains and two other strains, Rhodococcus sp. IFP 2042 and Bradyrhizobium sp. IFP2049, which had been previously isolated from one of the enrichments was carried out on the different enrichments based on quantitative PCR with specific 16S rRNA gene primers and the results were consistent with the hypothesized role of Actinobacteria and Betaproteobacteria in the degradation of ETBE and the possible role of Bradyrhizobium strains in the degradation of TBA.

Novel multispecies microbial consortia involved in lignocellulose and 5-hydroxymethylfurfural bioconversion

To develop a targeted metagenomics approach for the analysis of novel multispecies microbial consortia involved in the bioconversion of lignocellulose and furanic compounds, we applied replicated sequential batch aerobic enrichment cultures with either pretreated or untreated wheat straw as the sources of carbon and energy. After each transfer, exponential growth of bacteria was detected using microscopic cell counts, indicating that the substrate was being utilized. In batch, the final bacterial abundances increased from an estimated 5 to 8.7-9.5 log 16S rRNA gene copy numbers/ml. The abundances of fungal propagules showed greater variation, i.e., between 5.4 and 8.0 log ITS1 copies/ml. Denaturing gradient gel electrophoresis analyses showed that the bacterial consortia in both treatments reached approximate structural stability after six transfers. Moreover, the structures of the fungal communities were strongly influenced by substrate treatment. A total of 124 bacterial strains were isolated from the two types of enrichment cultures. The most abundant strains were affiliated with the genera Raoultella/Klebsiella, Kluyvera, Citrobacter, Enterobacter, Pseudomonas, Acinetobacter, Flavobacterium and Arthrobacter. Totals of 43 and 11 strains obtained from the untreated and pretreated substrates, respectively, showed (hemi)cellulolytic activity (CMC-ase and xylanase), whereas 96 strains were capable of growth in 7.5 mM 5-hydroxymethylfurfural. About 50 % of the latter showed extracellular oxidoreductase activity as detected by a novel iodide oxidation method. Also, (hemi)cellulolytic fungal strains related to Coniochaeta, Plectosphaerella and Penicillium were isolated. One Trichosporon strain was isolated from pretreated wheat straw. The two novel bacterial-fungal consortia are starting points for lignocellulose degradation applications.

Halophilic archaea in the human intestinal mucosa

The human gastrointestinal tract microbiota, despite its key roles in health and disease, remains a diverse, variable and poorly understood entity. Current surveys reveal a multitude of undefined bacterial taxa and a low diversity of methanogenic archaea. In an analysis of the microbiota in colonic mucosal biopsies from patients with inflammatory bowel disease we found 16S rDNA sequences representing a phylogenetically rich diversity of halophilic archaea from the Halobacteriaceae (haloarchaea), including novel phylotypes. As the human colon is not considered a salty environment and haloarchaea are described as extreme halophiles, we evaluated and further discarded the possibility that these sequences originated from pre-colonoscopy saline lavage solutions. Furthermore, aerobic enrichment cultures prepared from a patient biopsy at low salinity (2.5% NaCl) yielded haloarchaeal sequence types. Microscopic observation after fluorescence in situ hybridization provided evidence of the presence of viable archaeal cells in these cultures. These results prove the survival of haloarchaea in the digestive system and suggest that they may be members of the mucosal microbiota, even if present in low numbers in comparison with methanogenic archaea. Investigation of a potential physiological basis of this association may lead to new insights into gastrointestinal health and disease.

Thiohalobacter thiocyanaticus gen. nov., sp. nov., a moderately halophilic, sulfur-oxidizing gammaproteobacterium from hypersaline lakes, that utilizes thiocyanate

A moderately halophilic, obligately chemolithoautotrophic, sulfur-oxidizing bacterium, designated strain HRh1(T), was obtained from mixed sediment samples from hypersaline chloride-sulfate lakes in the Kulunda Steppe, in south-western Siberia (Russia), using aerobic enrichment culture at 1 M NaCl with thiocyanate as substrate. Cells of the isolate were short, non-motile rods with a Gram-negative type of cell wall. The bacterium was an obligate aerobe capable of chemolithoautotrophic growth with thiocyanate and thiosulfate. With thiosulfate, it grew at NaCl concentrations of 0.2-3.0 M (optimum 0.5 M) and at pH 6.3-8.0 (optimum pH 7.3-7.5). During growth on thiocyanate, cyanate was identified as an intermediate. The dominant cellular fatty acids were C(16 : 0) and C(18 : 1)omega7. Phylogenetic analysis based on 16S rRNA gene sequencing placed the isolate in the class Gammaproteobacteria as an independent lineage, with an unclassified marine sulfur-oxidizing bacterium as the closest culturable relative (93 % sequence similarity). A single cbbL gene (coding for the key enzyme of the Calvin-Benson cycle of autotrophic CO(2) assimilation) with relatively low similarity to any homologous genes found in chemolithoautotrophs was detected in strain HRh1(T). On the basis of our phenotypic and phylogenetic analysis, the halophilic isolate is proposed to represent a new genus and novel species, Thiohalobacter thiocyanaticus gen. nov., sp. nov. The type strain of Thiohalobacter thiocyanaticus is HRh1(T) (=DSM 21152(T) =UNIQEM U249(T)).

Different bacterial strategies to degrade taurocholate

Aerobic enrichment cultures with taurocholate or alkanesulfonates as sole sources of carbon and energy for growth were successful and yielded nine bacterial isolates, all of which utilized taurocholate. Growth was complex and involved not only many, usually transient, excretion products but also sorption of taurocholate and cholate to cells. Three metabolic strategies to dissimilate taurocholate were elucidated, all of which involved bile salt hydrolase cleaving taurocholate to cholate and taurine. Comamonas testosteroni KF-1 utilized both the taurine and the cholate moieties for growth. Pseudomonas spp., e.g. strain TAC-K3 and Rhodococcus equi TAC-A1 grew with the cholate moiety and released taurine quantitatively. Delftia acidovorans SPH-1 utilized the taurine moiety and released cholate.

N-Acetyltaurine dissimilated via taurine by Delftia acidovorans NAT

The naturally occurring sulfonate N-acetyltaurine was synthesized chemically and its identity was confirmed. Aerobic enrichment cultures for bacteria able to utilize N-acetyltaurine as sole source of fixed nitrogen or as sole source of carbon were successful. One representative isolate, strain NAT, which was identified as a strain of Delftia acidovorans, grew with N-acetyltaurine as carbon source and excreted stoichiometric amounts of sulfate and ammonium. Inducible enzyme activities were measured in crude extracts of this organism to elucidate the degradative pathway. Cleavage of N-acetyltaurine by a highly active amidase yielded acetate and taurine. The latter was oxidatively deaminated by taurine dehydrogenase to ammonium and sulfoacetaldehyde. This key intermediate of sulfonate catabolism was desulfonated by the known reaction of sulfoacetaldehyde acetyltransferase to sulfite and acetyl phosphate, which was further degraded to enter central metabolism. A degradative pathway including transport functions is proposed.

Isolation and characterization of aromatics-degrading microorganisms from the gut of the lower termite Coptotermes formosanus.

We isolated aromatics-degrading bacteria from the gut of a lower termite, Coptotermes formosanus, using a mineral salt medium containing various aromatic compounds as the sole carbon source. Two species, Burkholderia sp. strain VE22 and Citrobacter sp. strain VA53, were isolated by aerobic enrichment culture with veratraldehyde and vanillin, respectively. Strain VA53 could also grow and metabolize vanillin anaerobically.

Cometabolism of cis-1,2-dichloroethene by aerobic cultures grown on vinyl chloride as the primary substrate.

An aerobic enrichment culture was grown on vinyl chloride (VC) as the sole source of carbon and energy. In the absence of VC, the enrichment culture cometabolized cis-1,2-dichloroethene (cDCE) and, to a lesser extent, trans1,2-dichloroethene (tDCE), beginning with oxidation to the corresponding DCE-epoxides. When provided with VC (1.3 mM) and cDCE (0.2-0.3 mM), the enrichment culture cometabolized repeated additions of cDCE for over 85 days. Cometabolism of repeated additions of tDCE was also demonstrated but at a lower ratio of nongrowth substrate to VC. VC-grown Pseudomonas aeruginosa MF1 (previously isolated from the enrichment culture) also readily cometabolizes cDCE, with an observed transformation capacity (Tc,obs) of 0.82 micromol of cDCE/mg of total suspended solids (TSS). When provided with VC and cDCE, MF1 did not begin cometabolizing cDCE until nearly all of the VC was consumed. The presence of cDCE reduces the maximum specific rate of VC utilization. A kinetic model was developed that describes these phenomena via Monod parameters for substrate and nongrowth substrate, plus inactivation and inhibition coefficients. MF1 did not show any cometabolic activity on tDCE or trichloroethene and very limited activity on 1,1-DCE (Tc,obs = 2 x 10(-5) micromol/mg TSS). Above 40 microM, tDCE and TCE noticeably increased the maximum specific rate of VC utilization, even though neither compound was consumed during or after VC consumption. High concentrations of 1,1-DCE (950 microM) completely inhibited VC biodegradation. As there is currently no evidence for aerobic biodegradation of cDCE as a sole source of carbon and energy, the results of this study provide a potential explanation for in situ disappearance of cDCE when the only other significant substrate available is VC. It is fortuitous that the VC-grown cultures tested exhibit their highest cometabolic activity toward cDCE, because it is the predominant DCE isomer formed during anaerobic reductive dechlorination of trichloroethene and tetrachloroethene.

Bioremediation of Chlorobenzene-Contaminated Groundwater on Granular Activated Carbon Barriers

During the past decade, various promising technologies have been developed for the decontamination of groundwater insitu which do not require long-term pumping or high energy consumption. One approach is to use funnel and gate technology. In the case described here, the combination of adsorption of contaminants on granular activated carbon (GAC) and its biodegradation is applied to considerably extend the operating time of the filling material in the barrier system. Monochlorobenzene (MCB), a recalcitrant groundwater contaminant under anaerobic conditions, undergoes high-capacity adsorption on GAC up to about 450 mg per gram. Aerobic enrichment cultures, obtained from a contaminated aquifer, were able to mineralize initially adsorbed MCB. In respirometer experiments the rate of carbon dioxide formation was dependent on the equilibrium concentration of MCB. The oxygen consumption of activated carbon by means of autoxidative reactions may delay aerobic biodegradation in GAC filters. The oxygen uptake of pristine activated carbon amounted to 5.6 mg per gram GAC in laboratory column experiments. When GAC was pre-loaded with MCB, autoxidation rates were considerably reduced. Hence, it is advisable not to stimulate the biodegradation of MCB by oxygen supply in GAC biobarriers until after an initial period of solely sorptive MCB removal from the groundwater flow.

Transformation of 2,2'-dichlorodiisopropyl ether in mixed and pure culture.

An aerobic enrichment culture derived from a groundwater contaminated with organic and chloroorganic compounds was adapted to the transformation of 2,2'-dichlorodiisopropyl ether (DDE) in a continuous fixed-bed bioreactor. Continuous DDE removal efficiencies over 90% were achieved with a model water containing 3.3 mM methanol as co-substrate at DDE loading rates of up to 150 micromol l(-1) day(-1) with a hydraulic retention time of 24 h. In batch cultures, a stoichiometric release of 2 micromol chloride per micromol DDE transformed was observed. From the mixed culture, a strain was isolated that is able to grow on DDE as the sole energy and carbon source, tolerating DDE concentrations of up to 1 mM. Based on 16S rRNA sequencing, the strain is affiliated with the genus Rhodococcus.

Ethanedisulfonate is degraded via sulfoacetaldehyde in Ralstonia sp. strain EDS1.

Aerobic enrichment cultures (11) yielded three cultures able to utilise ethane-1,2-disulfonate as sole source of carbon and energy in salts medium. Two pure cultures were obtained and we worked with strain EDS1, which was assigned to the genus Ralstonia on the basis of its 16S rDNA sequence and simple taxonomic tests. Strain EDS1 utilised at least seven alkane(di)sulfonates, ethane-1,2-disulfonate, taurine, isethionate, sulfoacetate, sulfoacetaldehyde and propane-1,3-disulfonate, as well as methanesulfonate and formate. Growth with ethanedisulfonate was concomitant with substrate disappearance and the formation of 2 mol sulfate per mol substrate. The growth yield, 7 g protein (mol C)(-1), indicated quantitative utilisation of the substrate. Ethanedisulfonate-dependent oxygen uptake of whole cells during growth rose to a maximum before the end of growth and then sank rapidly; this was interpreted as evidence for an inducible desulfonative oxygenase that was not active in cell extracts. Inducible sulfoacetaldehyde sulfo-lyase was detected at high activity. Inducible degradation of taurine or isethionate or sulfoacetate via sulfoacetaldehyde sulfo-lyase is interpreted from the data.

Characterization of an isolate that uses vinyl chloride as a growth substrate under aerobic conditions.

An aerobic enrichment culture was developed by using vinyl chloride (VC) as the sole organic carbon and electron donor source. VC concentrations as high as 7.3 mM were biodegraded without apparent inhibition. VC use did not occur when nitrate was provided as the electron acceptor. A gram-negative, rod-shaped, motile isolate was obtained from the enrichment culture and identified based on biochemical characteristics and the sequence of its 16S rRNA gene as Pseudomonas aeruginosa, designated strain MF1. The observed yield of MF1 when it was grown on VC was 0.20 mg of total suspended solids (TSS)/mg of VC. Ethene, acetate, glyoxylate, and glycolate also served as growth substrates, while ethane, chloroacetate, glycolaldehyde, and phenol did not. Stoichiometric release of chloride and minimal accumulation of soluble metabolites following VC consumption indicated that the predominant fate for VC is mineralization and incorporation into cell material. MF1 resumed consumption of VC after at least 24 days when none was provided, unlike various mycobacteria that lost their VC-degrading ability after brief periods in the absence of VC. When deprived of oxygen for 2.5 days, MF1 did not regain the ability to grow on VC, and a portion of the VC was transformed into VC-epoxide. Acetylene inhibited VC consumption by MF1, suggesting the involvement of a monooxygenase in the initial step of VC metabolism. The maximum specific VC utilization rate for MF1 was 0.41 micromol of VC/mg of TSS/day, the maximum specific growth rate was 0.0048/day, and the Monod half-saturation coefficient was 0.26 microM. A higher yield and faster kinetics occurred when MF1 grew on ethene. When grown on ethene, MF1 was able to switch to VC as a substrate without a lag. It therefore appears feasible to grow MF1 on a nontoxic substrate and then apply it to environments that do not exhibit a capacity for aerobic biodegradation of VC.