Biomedical subjects

Joseph M Suflita

Publications and source records attributed to Joseph M Suflita.

13 recordsLinked to original sources

PubMed2006

Anaerobic biodegradation of natural gas condensate can be stimulated by the addition of gasoline.

Biodegradation of a broad range of linear and branched alkanes, parent and alkyl alicyclic hydrocarbons, and benzene and alkyl-substituted benzenes was observed when sediment and groundwater samples collected from a gas condensate-contaminated aquifer were incubated under methanogenic and especially under sulfate-reducing conditions, even though no exogenous nitrogen or phosphorus was added. This finding expands the range of hydrocarbon molecules known to undergo anaerobic decay and confirms that natural attenuation is an important process at this site. The addition of 1 mul of gasoline to the samples (approximately 10 ppm) had minimal impact on the biodegradation of saturated compounds, but substantially increased the diversity and extent of aromatic compounds undergoing transformation. We attribute this to the promotion or induction of biodegradation pathways in the indigenous microflora following the addition of the gasoline components. The promoting compounds are not precisely known, but may have been present in the initial condensate and reduced in concentration by various mechanisms (dissolution, biodegradation, etc.) such that their concentration in the aquifer fell below necessary levels. A variety of aromatic hydrocarbons would appear to be likely candidates.

Anaerobiosis↗

PubMed2006

Changes in organic matter biodegradability influencing sulfate reduction in an aquifer contaminated by landfill leachate.

In situ experiments were conducted to measure sulfate reduction rates and identify rate-limiting factors in a shallow, alluvial aquifer contaminated with municipal landfill leachate. Single-well, push-pull tests conducted in a well adjacent to the landfill with > 8 mM dissolved organic carbon (DOC) exhibited a sulfate reduction rate of 3.2 mumol SO4(-2) (L sediment)(-1) day(-1), a value in close agreement with laboratory-derived estimates. Identical tests conducted in wells located 90 m downgradient where DOC levels remained high (> 3 mM) showed no detectable sulfate consumption, and laboratory assays confirmed this observation. However, the rates of sulfate reduction in sediment samples obtained from this site were three times larger when they were amended with filter-sterilized groundwater from the upgradient location. The effect of various amendments on sulfate reduction rates was further examined in laboratory incubations using sediment collected from the downgradient site amended with 35S sulfate. Unamended sediments showed only weak conversion of the tracer to 35S sulfide (5 to 7 cpm/cm2), whereas the addition of Desulfovibrio cells increased 35S sulfide production to 44 cpm/cm2. However, the application of heat-killed Desulfovibrio had a similar stimulatory effect, as did a lactate amendment. Collectively, these findings indicate that the lack of measurable sulfate reduction at the downgradient site was not due to the absence of the necessary metabolic potential, the presence of lower sulfate concentration, or the quantity of electron donor, but by its biodegradability. The findings also indicate that field bioaugmentation attempts should be interpreted with caution.

Biodegradation, Environmental↗

PubMed2006

Desulfoglaeba alkanexedens gen. nov., sp. nov., an n-alkane-degrading, sulfate-reducing bacterium.

Two novel sulfate-reducing bacteria, strains ALDC(T) and Lake, which were able to oxidize n-alkanes, were isolated from a naval oily wastewater-storage facility (VA, USA) and from oilfield production water (OK, USA), respectively. The type strain (ALDC(T)) had a narrow substrate specificity and could grow only with n-alkanes (from C(6) to C(12)), pyruvate, butyrate, hexanoic acid and 4-methyloctanoic acid. Cells of strain ALDC(T) stained Gram-negative and were slightly curved, short rods with oval ends (2.5-3.0x1.0-1.4 microm), often occurring in pairs. Cells tended to form aggregates or large clusters and were non-motile and did not form endospores. Optimum growth occurred between 31 and 37 degrees C and at pH 6.5-7.2. NaCl was not required for growth, but salt concentrations up to 55 g l(-1) could be tolerated. The DNA G+C content was 53.6 mol%. Phylogenetic analysis of the 16S rRNA genes revealed that strains ALDC(T) and Lake were closely related, but not identical (99.9 % similarity). The two strains were not closely related to other known alkane-degrading, sulfate-reducing bacteria or to other genera of the Deltaproteobacteria. Therefore, it is proposed that strain ALDC(T) (=JCM 13588(T)=ATCC BAA-1302(T)) represents the type strain of a novel species and genus, with the name Desulfoglaeba alkanexedens gen. nov., sp. nov.

Alkanes↗

PubMed2006

Comparison of mechanisms of alkane metabolism under sulfate-reducing conditions among two bacterial isolates and a bacterial consortium.

Recent studies have demonstrated that fumarate addition and carboxylation are two possible mechanisms of anaerobic alkane degradation. In the present study, we surveyed metabolites formed during growth on hexadecane by the sulfate-reducing isolates AK-01 and Hxd3 and by a mixed sulfate-reducing consortium. The cultures were incubated with either protonated or fully deuterated hexadecane; the sulfate-reducing consortium was also incubated with [1,2-13C2]hexadecane. All cultures were extracted, silylated, and analyzed by gas chromatography-mass spectrometry. We detected a suite of metabolites that support a fumarate addition mechanism for hexadecane degradation by AK-01, including methylpentadecylsuccinic acid, 4-methyloctadecanoic acid, 4-methyloctadec-2,3-enoic acid, 2-methylhexadecanoic acid, and tetradecanoic acid. By using d34-hexadecane, mass spectral evidence strongly supporting a carbon skeleton rearrangement of the first intermediate, methylpentadecylsuccinic acid, was demonstrated for AK-01. Evidence indicating hexadecane carboxylation was not found in AK-01 extracts but was observed in Hxd3 extracts. In the mixed sulfate-reducing culture, however, metabolites consistent with both fumarate addition and carboxylation mechanisms of hexadecane degradation were detected, which demonstrates that multiple alkane degradation pathways can occur simultaneously within distinct anaerobic communities. Collectively, these findings underscore that fumarate addition and carboxylation are important alkane degradation mechanisms that may be widespread among phylogenetically and/or physiologically distinct microorganisms.

Alkanes↗

PubMed2005

Enrichment and isolation of anaerobic hydrocarbon-degrading bacteria.

Recent progress in microbiology resulted in the enrichment and isolation of anaerobic bacteria capable of the biodegradation of various hydrocarbons under a variety of electron-accepting conditions. Problems challenging the enrichment and isolation of anaerobic hydrocarbonclastic organisms required new approaches and modifications of conventional microbiological techniques. This chapter summarizes the collective experience accumulated in this area starting from anaerobic sampling precautions and includes all stages of cultivation from the construction of initial incubations to final isolation steps and the evaluation of culture purity.

Alkanes↗

PubMed2005

Stable isotopic studies of n-alkane metabolism by a sulfate-reducing bacterial enrichment culture.

Gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy were used to study the metabolism of deuterated n-alkanes (C6 to C12) and 1-13C-labeled n-hexane by a highly enriched sulfate-reducing bacterial culture. All substrates were activated via fumarate addition to form the corresponding alkylsuccinic acid derivatives as transient metabolites. Formation of d14-hexylsuccinic acid in cell extracts from exogenously added, fully deuterated n-hexane confirmed that this reaction was the initial step in anaerobic alkane metabolism. Analysis of resting cell suspensions amended with 1-13C-labeled n-hexane confirmed that addition of the fumarate occurred at the C-2 carbon of the parent substrate. Subsequent metabolism of hexylsuccinic acid resulted in the formation of 4-methyloctanoic acid, and 3-hydroxy-4-methyloctanoic acid was tentatively identified. We also found that 13C nuclei from 1-13C-labeled n-hexane became incorporated into the succinyl portion of the initial metabolite in a manner that indicated that 13C-labeled fumarate was formed and recycled during alkane metabolism. Collectively, the findings obtained with a sulfate-reducing culture using isotopically labeled alkanes augment and support the previously proposed pathway (H. Wilkes, R. Rabus, T. Fischer, A. Armstroff, A. Behrends, and F. Widdel, Arch. Microbiol. 177:235-243, 2002) for metabolism of deuterated n-hexane by a denitrifying bacterium.

Alkanes↗

PubMed2004

Progress curve analysis for enzyme and microbial kinetic reactions using explicit solutions based on the Lambert W function.

We present a simple method for estimating kinetic parameters from progress curve analysis of biologically catalyzed reactions that reduce to forms analogous to the Michaelis-Menten equation. Specifically, the Lambert W function is used to obtain explicit, closed-form solutions to differential rate expressions that describe the dynamics of substrate depletion. The explicit nature of the new solutions greatly simplifies nonlinear estimation of the kinetic parameters since numerical techniques such as the Runge-Kutta and Newton-Raphson methods used to solve the differential and integral forms of the kinetic equations, respectively, are replaced with a simple algebraic expression. The applicability of this approach for estimating Vmax and Km in the Michaelis-Menten equation was verified using a combination of simulated and experimental progress curve data. For simulated data, final estimates of Vmax and Km were close to the actual values of 1 microM/h and 1 microM, respectively, while the standard errors for these parameter estimates were proportional to the error level in the simulated data sets. The method was also applied to hydrogen depletion experiments by mixed cultures of bacteria in activated sludge resulting in Vmax and Km estimates of 6.531 microM/h and 2.136 microM, respectively. The algebraic nature of this solution, coupled with its relatively high accuracy, makes it an attractive candidate for kinetic parameter estimation from progress curve data.

Algorithms↗

PubMed2004

Periplasmic cytochrome c3 of Desulfovibrio vulgaris is directly involved in H2-mediated metal but not sulfate reduction.

Kinetic parameters and the role of cytochrome c(3) in sulfate, Fe(III), and U(VI) reduction were investigated in Desulfovibrio vulgaris Hildenborough. While sulfate reduction followed Michaelis-Menten kinetics (K(m) = 220 micro M), loss of Fe(III) and U(VI) was first-order at all concentrations tested. Initial reduction rates of all electron acceptors were similar for cells grown with H(2) and sulfate, while cultures grown using lactate and sulfate had similar rates of metal loss but lower sulfate reduction activities. The similarities in metal, but not sulfate, reduction with H(2) and lactate suggest divergent pathways. Respiration assays and reduced minus oxidized spectra were carried out to determine c-type cytochrome involvement in electron acceptor reduction. c-type cytochrome oxidation was immediate with Fe(III) and U(VI) in the presence of H(2), lactate, or pyruvate. Sulfidogenesis occurred with all three electron donors and effectively oxidized the c-type cytochrome in lactate- or pyruvate-reduced, but not H(2)-reduced cells. Correspondingly, electron acceptor competition assays with lactate or pyruvate as electron donors showed that Fe(III) inhibited U(VI) reduction, and U(VI) inhibited sulfate loss. However, sulfate reduction was slowed but not halted when H(2) was the electron donor in the presence of Fe(III) or U(VI). U(VI) loss was still impeded by Fe(III) when H(2) was used. Hence, we propose a modified pathway for the reduction of sulfate, Fe(III), and U(VI) which helps explain why these bacteria cannot grow using these metals. We further propose that cytochrome c(3) is an electron carrier involved in lactate and pyruvate oxidation and is the reductase for alternate electron acceptors with higher redox potentials than sulfate.

Cytochrome c Group↗

PubMed2003

Anaerobic oxidation of crude oil hydrocarbons by the resident microorganisms of a contaminated anoxic aquifer.

The biodegradation of two crude oils by microorganisms from an anoxic aquifer previously contaminated by natural gas condensate was examined under methanogenic and sulfate-reducing conditions. Artificially weathered Alaska North Slope crude oil greatly stimulated both methanogenesis and sulfate reduction. Gas chromatographic analysis revealed the entire n-alkane fraction of this oil (C13-C34) was consumed under both conditions. Naphthalene, 2-methylnaphthalene, and 2-ethylnaphthalene were also biodegraded but only in the presence of sulfate. Alba crude oil, which is naturally depleted in n-alkanes, resulted in a relatively modest stimulation of methanogenesis and sulfate reduction. Polycyclic aromatic hydrocarbon biodegradation was similar to that found for the Alaska North Slope crude oil, but a broader range of compounds was metabolized, including 2,6-dimethylnaphthalene and 2,7-dimethylnaphthalene in the presence of sulfate. These results indicate that n-alkanes are relatively labile, and their biodegradation in terrestrial environments is not necessarily limited by electron acceptor availability. Polycyclic aromatic hydrocarbons are relatively more recalcitrant, and the biodegradation of these substrates appeared to be sulfate-dependent and homologue-specific. This information should be useful for assessing the limits of in situ crude oil biodegradation in terrestrial environments and for making decisions regarding risk-based corrective actions.

Anaerobiosis↗

PubMed2003

Sources of sulfate supporting anaerobic metabolism in a contaminated aquifer.

Field and laboratory techniques were used to identify the biogeochemical factors affecting sulfate reduction in a shallow, unconsolidated alluvial aquifer contaminated with landfill leachate. Depth profiles of 35S-sulfate reduction rates in aquifer sediments were positively correlated with the concentration of dissolved sulfate. Manipulation of the sulfate concentration in samples revealed a Michaelis-Menten-like relationship with an apparent Km and Vmax of approximately 80 and 0.83 microM SO4(-2) x day(-1), respectively. The concentration of sulfate in the core of the leachate plume was well below 20 microM and coincided with very low reduction rates. Thus, the concentration and availability of this anion could limit in situ sulfate-reducing activity. Three sulfate sources were identified, including iron sulfide oxidation, barite dissolution, and advective flux of sulfate. The relative importance of these sources varied with depth in the alluvium. The relatively high concentration of dissolved sulfate at the water table is attributed to the microbial oxidation of iron sulfides in response to fluctuations of the water table. At intermediate depths, barite dissolves in undersaturated pore water containing relatively high concentrations of dissolved barium (approximately 100 microM) and low concentrations of sulfate. Dissolution is consistent with the surface texture of detrital barite grains in contact with leachate. Laboratory incubations of unamended and barite-amended aquifer slurries supported the field observation of increasing concentrations of barium in solution when sulfate reached low levels. At a deeper highly permeable interval just above the confining bottom layer of the aquifer, sulfate reduction rates were markedly higher than rates at intermediate depths. Sulfate is supplied to this deeper zone by advection of uncontaminated groundwater beneath the landfill. The measured rates of sulfate reduction in the aquifer also correlated with the abundance of accumulated iron sulfide in this zone. This suggests that the current and past distributions of sulfate-reducing activity are similar and that the supply of sulfate has been sustained at these sites.

Bacteria, Anaerobic↗

PubMed2003

Biodegradation of an alicyclic hydrocarbon by a sulfate-reducing enrichment from a gas condensate-contaminated aquifer.

We used ethylcyclopentane (ECP) as a model alicyclic hydrocarbon and investigated its metabolism by a sulfate-reducing bacterial enrichment obtained from a gas condensate-contaminated aquifer. The enrichment coupled the consumption of ECP with the stoichiometrically expected amount of sulfate reduced. During ECP biodegradation, we observed the transient accumulation of metabolite peaks by gas chromatography-mass spectrometry, three of which had identical mass spectrometry profiles. Mass-spectral similarities to analogous authentic standards allowed us to identify these metabolites as ethylcyclopentylsuccinic acids, ethylcyclopentylpropionic acid, ethylcyclopentylcarboxylic acid, and ethylsuccinic acid. Based on these findings, we propose a pathway for the degradation of this alicyclic hydrocarbon. Furthermore, a putative metabolite similar to ethylcyclopentylsuccinic acid was also found in samples of contaminated groundwater from the aquifer. However, no such finding was evident for samples collected from wells located upgradient of the gas condensate spill. Microbial community analysis of the ECP-degrading enrichment by denaturing gradient gel electrophoresis revealed the presence of at least three different organisms using universal eubacterial primers targeting 550 bp of the 16S rRNA gene. Based on sequence analysis, these organisms are phylogenetically related to the genera Syntrophobacter and Desulfotomaculum as well as a member of the Cytophaga-Flexibacter-Bacteroides group. The evidence suggests that alicyclic hydrocarbons such as ECP can be anaerobically activated by the addition to the double bond of fumarate to form alkylsuccinate derivatives under sulfate-reducing conditions and that the reaction occurs in the laboratory and in hydrocarbon-impacted environments.

Anaerobiosis↗

PubMed2002

Detection of anaerobic metabolites of saturated and aromatic hydrocarbons in petroleum-contaminated aquifers.

Recent investigations have demonstrated that several classes of petroleum hydrocarbons are susceptible to anaerobic decay, including alkanes and mono- and polycyclic aromatic compounds. In previous work, benzylsuccinates were shown to be useful indicators of in situ anaerobic alkylbenzene metabolism. In the present study, we sought to determine whether metabolites of alkanes and naphthalenes could similarly be used as indicators of the intrinsic decomposition of these compounds in petroleum-contaminated aquifers. Such metabolites include succinate derivatives of n-alkanes, cyclic alkanes, and alkylaromatic hydrocarbons as well as naphthoic acids. Using gas chromatography-mass spectrometry (GC-MS), we analyzed trimethylsilyl-derivatized organic extracts from six hydrocarbon-contaminated groundwaters for MS fragment ions indicative of such anaerobic metabolites. Geochemical indicators in these aquifers suggested the prevalence of anaerobic processes. In the groundwaters of the contaminated sites, we found compounds whose MS profiles suggested that they were indeed alkylsuccinic acids, ranging from C3 to C11 succinates. Propyl-, hexyl-, octyl-, and decylsuccinic acids were positively identified in the groundwaters by GC-MS matches with chemical or biologically produced standards. In two of the aquifers, we also detected components whose MS profiles matched with authentic standards of naphthoic acids and tetrahydronaphthoic acids. Metabolites were detected in nanomolar concentrations. The finding of these putative anaerobic metabolites of alkanes and naphthalenes signifies the in situ biodegradation of these hydrocarbons and attests to their value as indicators of intrinsic bioremediation.

Anaerobiosis↗

PubMed2002

In-situ evidence for uranium immobilization and remobilization.

The in-situ microbial reduction and immobilization of uranium was assessed as a means of preventing the migration of this element in the terrestrial subsurface. Uranium immobilization (putatively identified as reduction) and microbial respiratory activities were evaluated in the presence of exogenous electron donors and acceptors with field push-pull tests using wells installed in an anoxic aquifer contaminated with landfill leachate. Uranium(VI) amended at 1.5 microM was reduced to less than 1 nM in groundwater in less than 8 d during all field experiments. Amendments of 0.5 mM sulfate or 5 mM nitrate slowed U(VI) immobilization and allowed for the recovery of 10% and 54% of the injected element, respectively, as compared to 4% in the unamended treatment. Laboratory incubations confirmed the field tests and showed that the majority of the U(VI) immobilized was due to microbial reduction. In these tests, nitrate treatment (7.5 mM) inhibited U(VI) reduction, and nitrite was transiently produced. Further push-pull tests were performed in which either 1 or 5 mM nitrate was added with 1.0 uM U(VI) to sediments that already contained immobilized uranium. After an initial loss of the amendments, the concentration of soluble U(VI) increased and eventually exceeded the injected concentration, indicating that previously immobilized uranium was remobilized as nitrate was reduced. Laboratory experiments using heat-inactivated sediment slurries suggested that the intermediates of dissimilatory nitrate reduction (denitrification or dissimilatory nitrate reduction to ammonia), nitrite, nitrous oxide, and nitric oxide were all capable of oxidizing and mobilizing U(IV). These findings indicate that in-situ subsurface U(VI) immobilization can be expected to take place under anaerobic conditions, but the permanence of the approach can be impaired by disimilatory nitrate reduction intermediates that can mobilize previously reduced uranium.

Nitrates↗