Influence of Carbon and Microbial Community Priming on the Attenuation of Uranium in a Contaminated Floodplain Aquifer.

The capacity for subsurface sediments to sequester radionuclide contaminants, such as uranium (U), and retain them after bioremediation efforts are completed is critical to the long-term stewardship of re-mediated sites. In U bioremediation strategies, carbon amendment stimulates bioreduction of U(VI) to U(IV), immobilizing it within the sediments. Sediments enriched in natural organic matter are naturally capable of sequestering significant U, but may serve as sources to the aquifer, contributing to plume persistence. Two types of organic-rich sediments were compared to better understand U release mechanisms. Sediments that were artificially primed for U removal were retrieved from an area previously biostimulated while detrital-rich sediments were collected from a location never subject to amendment. Batch incubations demonstrated that primed sediments rapidly removed uranium from the groundwater, whereas naturally reduced sediments released a sizeable portion of U before U(VI)-reduction commenced. Column experiments confirmed that U release persisted for 65 pore volumes in naturally reduced sediments, demonstrating their sink-source behavior. Acetate addition to primed sediments shifted the microbial community from sulfate-reducing bacteria within Desulfobacteraceae to the iron-reducing Geobacteraceae and Firmicutes, associated with efficient U(VI) removal and retention, respectively. In contrast, Geobacteraceae communities in naturally reduced sediments were replaced by sequences with similarity to Pseudomonas spp. during U release, while U(VI) removal only occurred with enrichment of Firmicutes. These investigations stress the importance of characterizing zones with heterogeneous carbon pools at U-contaminated sites prior to the determination of a remedial strategy to identify areas, which may contribute to long-term sourcing of the contaminants.

Microbial reduction of uranium under iron- and sulfate-reducing conditions: Effect of amended goethite on microbial community composition and dynamics.

There is a growing need for a better understanding of the biogeochemical dynamics involved in microbial U(VI) reduction due to an increasing interest in using biostimulation via electron donor addition as a means to remediate uranium contaminated sites. U(VI) reduction has been observed to be maximized during iron-reducing conditions and to decrease upon commencement of sulfate-reducing conditions. There are many unknowns regarding the impact of iron/sulfate biogeochemistry on U(VI) reduction. This includes Fe(III) availability as well as the microbial community changes, including the activity of iron-reducers during the uranium biostimulation period even after sulfate reduction becomes dominant. Column experiments were conducted with Old Rifle site sediments containing Fe-oxides, Fe-clays, and sulfate rich groundwater. Half of the columns had sediment that was augmented with small amounts of Fe(III) in the form of (57)Fe-goethite, allowing for a detailed tracking of minute changes of this added phase to study the effects of increased Fe(III) levels on the overall biostimulation dynamics. Mössbauer spectroscopy showed that the added (57)Fe-goethite was bioreduced only during the first thirty days of biostimultuion, after which it remained constant. Augmentation with Fe(III) had a significant effect on the total flux of electrons towards different electron acceptors; it suppressed the degree of sulfate reduction, had no significant impact on Geobacter-type bacterial numbers but decreased the bacterial numbers of sulfate reducers and affected the overall microbial community composition. The addition of Fe(III) had no noticeable effect on the total uranium reduction.

Polar lipid fatty acids, LPS-hydroxy fatty acids, and respiratory quinones of three Geobacter strains, and variation with electron acceptor.

The polar lipid fatty acids, lipopolysaccharide hydroxy-fatty acids, and respiratory quinones of Geobacter metallireducens str. GS-15, Geobacter sulfurreducens str. PCA, and Geobacter bemidjiensis str. Bem are reported. Also, the lipids of G. metallireducens were compared when grown with Fe(3+) or nitrate as electron acceptors and G. sulfurreducens with Fe(3+) or fumarate. In all experiments, the most abundant polar lipid fatty acids were 14:0, i15:0, 16:1 omega 7c, 16:1 omega 5c, and 16:0; lipopolysaccharide hydroxy-fatty acids were dominated by 3oh16:0, 3oh14:0, 9oh16:0, and 10oh16:0; and menaquinone-8 was the most abundant respiratory quinone. Some variation in lipid profiles with strain were observed, but not with electron acceptor.

Characterization of microbial activities and U reduction in a shallow aquifer contaminated by uranium mill tailings.

A characterization of the Shiprock, NM, uranium mill tailing site focused on the geochemical and microbiological factors governing in-situ uranium-redox reactions. Groundwater and aqueous extracts of sediment samples contained a wide concentration range of sulfate, nitrate, and U(VI) with median values of 21.2 mM, 16.1 micro M, and 2.7 micro M, respectively. Iron(III) was not detected in groundwater, but a median value of 0.3 mM in sediment extracts was measured. Bacterial diversity down gradient from the disposal pile reflected the predominant geochemistry with relatively high numbers of sulfate- and nitrate-reducing microorganisms, and smaller numbers of acetogenic, methanogenic, nitrate-dependent Fe(II)-oxidizing, Fe(III)-reducing, and sulfide-oxidizing bacteria. In aquifer slurry incubations, nitrate reduction was always preferred and had a negative impact on sulfate-, Fe(III)-, and U-reduction rates. We also found that sulfate-reduction rates decreased sharply in the presence of clay, while Fe(III)-reduction increased with no clear impact on U reduction. In the absence of clay, iron and sulfate reduction correlated with concentrations of Fe(III) and sulfate, respectively. Rates of U(VI) loss did not correlate with the concentration of any electron acceptor. With the exception of Fe(III), electron donor amendment was largely unsuccessful in stimulating electron acceptor loss over a 2-week incubation period, suggesting that endogenous forms of organic matter were sufficient to support microbial activity. Our findings suggest that efforts to accelerate biological U reduction should initially focus on stimulating nitrate removal.

Diversity and characterization of sulfate-reducing bacteria in groundwater at a uranium mill tailings site.

Microbially mediated reduction and immobilization of U(VI) to U(IV) plays a role in both natural attenuation and accelerated bioremediation of uranium-contaminated sites. To realize bioremediation potential and accurately predict natural attenuation, it is important to first understand the microbial diversity of such sites. In this paper, the distribution of sulfate-reducing bacteria (SRB) in contaminated groundwater associated with a uranium mill tailings disposal site at Shiprock, N.Mex., was investigated. Two culture-independent analyses were employed: sequencing of clone libraries of PCR-amplified dissimilatory sulfite reductase (DSR) gene fragments and phospholipid fatty acid (PLFA) biomarker analysis. A remarkable diversity among the DSR sequences was revealed, including sequences from delta-Proteobacteria, gram-positive organisms, and the Nitrospira division. PLFA analysis detected at least 52 different mid-chain-branched saturate PLFA and included a high proportion of 10me16:0. Desulfotomaculum and Desulfotomaculum-like sequences were the most dominant DSR genes detected. Those belonging to SRB within delta-Proteobacteria were mainly recovered from low-uranium (< or =302 ppb) samples. One Desulfotomaculum-like sequence cluster overwhelmingly dominated high-U (>1,500 ppb) sites. Logistic regression showed a significant influence of uranium concentration over the dominance of this cluster of sequences (P = 0.0001). This strong association indicates that Desulfotomaculum has remarkable tolerance and adaptation to high levels of uranium and suggests the organism's possible involvement in natural attenuation of uranium. The in situ activity level of Desulfotomaculum in uranium-contaminated environments and its comparison to the activities of other SRB and other functional groups should be an important area for future research.

A survey of 16S rRNA and amoA genes related to autotrophic ammonia-oxidizing bacteria of the beta-subdivision of the class proteobacteria in contaminated groundwater.

In this study, we investigated the size and structure of autotrophic ammonia oxidizer (AAO) communities in the groundwater of a contamination plume originating from a mill-tailings disposal site. The site has high levels of dissolved N from anthropogenic sources, and exhibited wide variations in the concentrations of NO3- and NH3 + NH4+. Community structures were examined by PCR-DGGE targeting 16S rDNA with band excision and sequence analysis, and by analysis of amoA fragment clone libraries. AAO population sizes were estimated by competitive PCR targeting the gene amoA, and correlated significantly with nitrate concentration. Most samples revealed novel diversity in AAO 16S rDNA and amoA gene sequences. Both 16S rDNA and amoA analyses suggested that all samples were dominated by Nitrosomonas sp., Nitrosospira sp. being detected in only 3 of 15 samples. This study indicated numerical dominance of Nitrosomonas over Nitrosospira in groundwater, and suggests that groundwater ammonia oxidizers are more similar to those dominating freshwater sediments than bulk soil.

Microbiology of vadose zone paleosols in south-central Washington State.

Three unsaturated subsurface paleosols influenced by moisture recharge, including a highly developed calcic paleosol, were studied to investigate the microbiology of paleosols. Two near-surface paleosols, one impacted by moisture recharge and the other beyond the influence of recharge, were also sampled to directly assess the effect of moisture recharge on the activity and composition of the microbial community associated with paleosols. The highly developed paleosol had a higher population of culturable heterotrophs, a greater glucose mineralization potential, a higher microbial diversity based on colony morphology, and a more than 20-fold higher concentration of ATP than the two weakly developed paleosols. The recharged near-surface paleosol, as compared to the near-surface paleosol unaffected by recharge, had a lower population of culturable heterotrophs, smaller mineralization rate constant, and lower richness based on colony morphology. The recharged paleosols contained predominantly gram-negative isolates, whereas the paleosol unaffected by recharge contained predominantly gram-positive isolates. Storage at 4°C of subsurface and near-surface paleosol samples containing high water potential increased the population of culturable aerobic heterotrophs, decreased diversity in colony morphology, and increased first-order rate constants and decreased lag times for glucose mineralization. These results indicate that aerobic heterotrophs are present in deep vadose zone paleosols and that there is potential for stimulation of their in situ growth and activity.

Dispersion of a fluorescein dye-ampicillin mixture after intratracheal administration in calves.

The pulmonary and systemic dispersion of a dye mixture after intratracheal (i.t.) administration was characterized in young dairy calves. Five calves were given i.t. injections of a fluorescein and ampicillin mixture, and were then killed at 2, 5 or 15 min after treatment. The respiratory system, nasal pharynx area, nose, liver, kidney, stomach and esophagus were removed and examined. Sections were taken from lung areas for histopathological examinations and all tissues were photographed, using floodlight and ultraviolet light. There was uniform low-grade fluorescence throughout the lung, but the greatest fluorescence followed an anterior ventral dispersion. The dye was readily absorbed from the lung into the bloodstream, causing fluorescence in the liver, bile, kidney and urine. Coughing of the calves during the injection caused some of the mixture to be expectorated and swallowed, resulting in fluorescence in the esophagus, stomach contents, and nasal pharanges, turbinates and nostrils.

Depletion of antibiotics from the mammary gland of goats.

Four intramammary infusion products were tested in 10 normal goats to determine their rates of depletion from milk. The products tested, which are marketed for treatment of mastitis in the bovine, contained the single active ingredient erythromycin, oxytetracycline, penicillin, or cephapirin. Each mammary gland was infused, after the goats were milked, with the maximum recommended dose of test product (dosing frequency and quantity) for administration to lactating dairy cattle. With one exception, no antibiotics could be detected in the milk by the end of the bovine milk-discard period. Penicillin was detectable in the milk of one goat for 72 h after the last dose of product containing this antibiotic was given (60-h withdrawal period). Only the product containing oxytetracycline produced significant adverse reactions in the mammary gland. The applicator tips of the products were too large for atraumatic insertion into the teat opening of some goats. Overall, results of this limited study indicated that some intramammary infusion products can be used to treat mastitis in the goat if instructions for use in the bovine are followed.

Disposition of ampicillin administered intravenously and intratracheally to young calves.

The profile of the blood concentration of ampicillin vs time was compared in 24 calves after ampicillin administration (10 mg/kg body weight) by the intratracheal (i.t.) and the intravenous (i.v.) routes in a crossover study. In a subsequent study, 12 of the calves were given ampicillin i.v. and 12 were given ampicillin i.t.; at 15 min and 2 h after drug administration, the calves were killed and lungs, kidneys, liver, urine and plasma were collected for ampicillin assay. The mean ampicillin concentration in blood after i.v. treatment was significantly (P less than 0.01) higher than after i.t. treatment. The mean ampicillin level in lungs after i.t. treatment was significantly higher than the level after i.v. treatment, but concentrations in kidneys, liver, urine and plasma after i.v. treatment were higher (but not significantly) than those after i.t. treatment.