The Microbial Ecology of a Hyper-Alkaline Spring,and Impacts of an Alkali-Tolerant Community During Sandstone Batch and Column Experiments Representative of a Geological Disposal Facility for Intermediate-Level Radioactive Waste

Naturally occurring hyper-alkaline springs and associated hyper-alkaline environments may have components that are analogous to a cement-based deep geological disposal facility (GDF) for intermediate level radioactive waste (ILW). Such high pH environments could give insights into the biogeochemical processes that could occur in the region of a GDF environment after the ingress of GDF-derived groundwater leads to the formation of a hyper-alkaline plume in the surrounding rock mass. This study focuses on the microbial community composition found at a highly alkaline spring near Buxton, Derbyshire, England, and the variation in community structure across spatially separated sample points of contrasting pH values (ranging from pH 7.5–13). Communities containing alkaliphilic and alkalitolerant bacteria were observed across the site by PCR amplification and 16S rRNA gene pyrosequencing and included members of the families Comamonadaceae and Xanthomonadaceae. At pH 13, the sequence library was dominated by Gammaproteobacteria of the families Pseudomonadaceae and Enterobacteriaceae. Bacterial communities from the site demonstrated the ability to reduce Fe(III) in microcosm experiments up to pH 11.5, suggesting the potential to reduce other metals and radionuclides of relevance to cement-encapsulated intermediate level radioactive waste (ILW) disposal. In laboratory column flow-through experiments, microbial communities present at the field site were also able to colonize crushed sandstone. Bacterial community composition varied between columns that had been supplied with alkali surface waters from the site amended with carbon (lactate and acetate, as proxies for products of cellulose degradation from ILW), and control columns that were not supplied with added carbon. Members of the family Clostridiaceae dominated the sequence library obtained from the carbon amended column inlet (45.8% of library), but became less dominant at the outlet (20.8%). Members of the family Sphingomonadaceae comprised 11.8% of the sequence library obtained from the control column inlet, but were not present in sediments collected from the column outlet, whereas the relative abundance of members of the family Comamonadaceae increased from the column inlet (35.2%) to the column outlet (57.2%). The spatial variation in community composition within the columns is indicative of discrete biogeochemical zonation in these flow-through systems.     

Anoxic Biodegradation of Isosaccharinic Acids at Alkaline pH by Natural Microbial Communities

One design concept for the long-term management of the UK’s intermediate level radioactive wastes (ILW) is disposal to a cementitious geological disposal facility (GDF). Under the alkaline (10.0<pH>13.0) anoxic conditions expected within a GDF, cellulosic wastes will undergo chemical hydrolysis. The resulting cellulose degradation products (CDP) are dominated by α- and β-isosaccharinic acids (ISA), which present an organic carbon source that may enable subsequent microbial colonisation of a GDF. Microcosms established from neutral, near-surface sediments demonstrated complete ISA degradation under methanogenic conditions up to pH 10.0. Degradation decreased as pH increased, with β-ISA fermentation more heavily influenced than α-ISA. This reduction in degradation rate was accompanied by a shift in microbial population away from organisms related to Clostridium sporosphaeroides to a more diverse Clostridial community. The increase in pH to 10.0 saw an increase in detection of Alcaligenes aquatilis and a dominance of hydrogenotrophic methanogens within the Archaeal population. Methane was generated up to pH 10.0 with acetate accumulation at higher pH values reflecting a reduced detection of acetoclastic methanogens. An increase in pH to 11.0 resulted in the accumulation of ISA, the absence of methanogenesis and the loss of biomass from the system. This study is the first to demonstrate methanogenesis from ISA by near surface microbial communities not previously exposed to these compounds up to and including pH 10.0.     

Distribution and activity of microorganisms in the deep repository for liquid radioactive waste at the Siberian Chemical Combine

The physicochemical conditions, composition of microbial communities, and the rates of anaerobic processes in the deep sand horizons used as a repository for liquid radioactive wastes (LRW) at the Siberian Chemical Combine (Seversk, Tomsk oblast), were studied. Formation waters from the observation wells drilled into the horizons used for the radioactive waste disposal were found to be inhabited by microorganisms of different physiological groups, including aerobic organotrophs, anaerobic fermentative, denitrifying, sulfate-reducing, and methanogenic bacteria. The density of microbial population, as determined by cultural methods, was low and usually did not exceed 10⁴ cells/ml. Enrichment cultures of microorganisms producing gases (hydrogen, methane, carbon dioxide, and hydrogen sulfide) and capable of participation in the precipitation of metal sulfides were obtained from the waters of the disposal site. The contemporary processes of sulfate reduction and methanogenesis were assayed; the rates of these terminal processes of organic matter destruction were found to be low. The denitrifying bacteria from the deep repository were capable of reducing the nitrates contained in the wastes, provided sources of energy and biogenic elements were available. Biosorption of radionuclides by the biomass of aerobic bacteria isolated from groundwater was demonstrated. The results obtained give us insight into the functional structure of the microbial community inhabiting the waters of repository horizons. This study indicates that the numbers and activity of microbial cells are low both inside and outside the zone of radioactive waste dispersion, in spite of the long period of waste discharge.     

Microbiological processes in the Severnyi deep disposal site for liquid radioactive wastes

Local monitoring of physicochemical, radiochemical, and microbiological parameters was performed in the deep horizons of the Severnyi site used for disposal of liquid radioactive waste (LRW). Analysis of the chemical and radiochemical composition of the wastes and formation fluid revealed that the boundary for migration of radionuclides lagged behind that for nonradioactive waste components (sodium nitrate) and tritium. The physicochemical and radiochemical conditions in deep horizons did not prevent microbial growth. The numbers of microorganisms (aerobic organotrophs, denitrifying, fermentative, sulfate-reducing, and methanogenic) were low, as were the rates of sulfate reduction and methanogenesis; they increased in the waste dispersion zone. The microorganisms from deep horizons were able to produce gases (CH₄, CO₂, N₂, and H₂S) from possible waste components. Denitrifying bacteria belonged to different Pseudomonas species and reduced nitrate to dinitrogen under the conditions of pH, salinity, temperature, and radioactivity found in the disposal site. These results suggest the need for control of microbiological processes in deep disposal site for liquid RW.     

Distribution,diversity and activity of microorganisms in the hyper-alkaline spring waters of Maqarin in Jordan

The hyper-alkaline, high-Ca²⁺ springs of Maqarin, Jordan, were investigated as an analogue for various microbial processes at the extremely high pH generated by cement and concrete in some underground radioactive waste repositories. Leaching of metamorphic, cementitious phases in Maqarin has produced current, hyper-alkaline groundwater with a maximum pH of 12.9. Six consecutive expeditions were undertaken to the area during 1994–2000. The total number of microorganisms in the alkaline waters was 10³–10⁵ cells/ml. Analysis of the 16S-ribosomal ribonucleic acid (rRNA) diversity revealed microorganisms mainly belonging to the Proteobacteria. Obvious similarities between the obtained sequences and sequences from other alkaline sites could not be found. Numerous combinations of culture media compositions were inoculated with spring, seepage and groundwaters and incubated under aerobic and anaerobic conditions with various carbon sources. Assimilation studies were performed using identical radio-labeled carbon sources. Glucose seemed to be the preferred carbon source for assimilation, followed by acetate, lactate, and leucine. The results demonstrate that microorganisms from the hyper-alkaline springs of Maqarin could grow and be metabolically active under aerobic and anaerobic hyper-alkaline conditions. However, the growth and activity found were not vigorous; instead, slow growth, low numbers, and a generally low metabolic activity were found. This suggests that microbial activity will be low during the hyper-alkaline phase of cementitious repositories.Communicated by W.D. Grant     

Biodegradation of the Alkaline Cellulose Degradation Products Generated during Radioactive Waste Disposal

The anoxic, alkaline hydrolysis of cellulosic materials generates a range of cellulose degradation products (CDP) including α and β forms of isosaccharinic acid (ISA) and is expected to occur in radioactive waste disposal sites receiving intermediate level radioactive wastes. The generation of ISA''s is of particular relevance to the disposal of these wastes since they are able to form complexes with radioelements such as Pu enhancing their migration. This study demonstrates that microbial communities present in near-surface anoxic sediments are able to degrade CDP including both forms of ISA via iron reduction, sulphate reduction and methanogenesis, without any prior exposure to these substrates. No significant difference (n = 6, p = 0.118) in α and β ISA degradation rates were seen under either iron reducing, sulphate reducing or methanogenic conditions, giving an overall mean degradation rate of 4.7×10⁻² hr⁻¹ (SE±2.9×10⁻³). These results suggest that a radioactive waste disposal site is likely to be colonised by organisms able to degrade CDP and associated ISA''s during the construction and operational phase of the facility.     

Distribution and activity of microorganisms in the deep repository for liquid radioactive waste at the Siberian Chemical Combine

The physicochemical conditions, composition of microbial communities, and the rates of anaerobic processes in the deep sandy horizons used as a repository for liquid radioactive wastes (LRW) at the Siberian Chemical Combine (Seversk, Tomsk oblast), were studied. Formation waters from the observation wells drilled into the production horizons of the radioactive waste disposal site were found to be inhabited by microorganisms of different physiological groups, including aerobic organotrophs, anaerobic fermentative, denitrifying, sulfate-reducing, and methanogenic bacteria. The density of microbial population, as determined by cultural methods, was low and usually did not exceed 10(4) cells/ml. Enrichment cultures of microorganisms producing gases (hydrogen, methane, carbon dioxide, and hydrogen sulfide) and capable of participation in the precipitation of metal sulfides were obtained from the waters of production horizons. The contemporary processes of sulfate reduction and methanogenesis were assayed; the rates of these terminal processes of organic matter destruction were found to be low. The denitrifying bacteria from the underground repository were capable of reducing the nitrates contained in the wastes, provided sources of energy and biogenic elements were available. Biosorption of radionuclides by the biomass of aerobic bacteria isolated from groundwater was demonstrated. The results obtained give us insight into the functional structure of the microbial community inhabiting the waters of repository production horizons. This study indicates that the numbers and activity of microbial cells are low both inside and outside the zone of radioactive waste dispersion, in spite of the long period of waste discharge.     

Microorganisms in a Disposal Site for Liquid Radioactive Wastes and Their Influence on Radionuclides

Deep subsurface horizons used for the disposal of liquid low- and intermediate-level radioactive wastes of the Siberian Chemical Complex (SCC, Russia) were studied by microbiological, radioisotope, and molecular biological methods. It was shown that a diverse microbial community inhabited the groundwater. The cell numbers of microorganisms of the major metabolic groups and the rates of sulfate reduction, denitrification, and methanogenesis in natural groundwater were low and increased in the zone of wastes dispersion. More than 40 strains belonging to the genera Kocuria, Microbacterium, Pseudomonas, Pantoea, Acinetobacter, Enterobacter, Klebsiella, Stenotrophomonas, Sphingomonas, Staphylococcus, Acidivorax, Shewanella, and Desulfosporosinus were isolated from the disposal sites. Among the isolates, the microorganisms were found that were able to concentrate actinides and other transuranium elements. Aerobic bacteria were able to sorb various radionuclides in laboratory experiments; however, biosorption was low in sample of groundwater and in carbonate solutions containing several radionuclides. Reduction of U(VI) by a sulfate-reducing enrichment culture from the site and reduction of U(VI) and Np(V) by an isolate Shewanella were observed in the presence of various organic substrates. These results show the necessity of further ecosystem characterization based on microbiological and radiochemical studies and modeling of biogeochemical processes at the deep disposal sites for liquid radioactive wastes.     

Microbial Diversity at Iron-Clay Interfaces after 10 Years of Interaction Inside a Deep Argillite Geological Formation (Tournemire,France)

In the context of a geological disposal of radioactive waste in clayey formations, the consequences of microbial activity are of concern regarding the corrosion of metallic components. The purpose of the present work was to characterize the microbial diversity that may have impacted corrosion processes at the interface between re-compacted argillite and steel coupons after 10 years of interaction inside the Toarcian argillite layer in Tournemire (France) under in situ conditions. Several types of steel coupons were introduced in 1999 in two boreholes (so-called CR6 and CR8) drilled in the geological formation and filled with re-compacted argillite. CR6 borehole was drilled horizontally from a century-old railway tunnel and coupons were placed in the undisturbed argillite. CR8 borehole was drilled vertically under the tunnel, in conditions influenced by the draining of the Cernon fault water. CR6 and CR8 boreholes were overcored 10 years later and steel coupons as well as re-compacted argillite samples were analyzed separately. The characterization of their microbial diversity was carried out using culture media and molecular methods using 16S rRNA genes cloning. Data resulting from both approaches were complementary. Isolates and clone sequences were affiliated to only 3 bacterial phyla: Firmicutes, Actinobacteria and Proteobacteria. The biodiversities differed depending on the steel type and the borehole considered, indicating the influence of both iron-clay interactions and in situ environmental conditions. This analysis has highlighted the presence of sulphate-reducing bacteria, iron-reducing bacteria and isolates capable to develop at high temperatures. These microorganisms can grow at the interfaces between materials in a very short period of time compared with planned durations of disposal. Thus, these results should be considered to assess the consequences of microbial activities on the evolution of the metallic components like overpacks.     

Evidence of the Generation of Isosaccharinic Acids and Their Subsequent Degradation by Local Microbial Consortia within Hyper-Alkaline Contaminated Soils,with Relevance to Intermediate Level Radioactive Waste Disposal

The contamination of surface environments with hydroxide rich wastes leads to the formation of high pH (>11.0) soil profiles. One such site is a legacy lime works at Harpur Hill, Derbyshire where soil profile indicated in-situ pH values up to pH 12. Soil and porewater profiles around the site indicated clear evidence of the presence of the α and β stereoisomers of isosaccharinic acid (ISA) resulting from the anoxic, alkaline degradation of cellulosic material. ISAs are of particular interest with regards to the disposal of cellulosic materials contained within the intermediate level waste (ILW) inventory of the United Kingdom, where they may influence radionuclide mobility via complexation events occurring within a geological disposal facility (GDF) concept. The mixing of uncontaminated soils with the alkaline leachate of the site resulted in ISA generation, where the rate of generation in-situ is likely to be dependent upon the prevailing temperature of the soil. Microbial consortia present in the uncontaminated soil were capable of surviving conditions imposed by the alkaline leachate and demonstrated the ability to utilise ISAs as a carbon source. Leachate-contaminated soil was sub-cultured in a cellulose degradation product driven microcosm operating at pH 11, the consortia present were capable of the degradation of ISAs and the generation of methane from the resultant H₂/CO₂ produced from fermentation processes. Following microbial community analysis, fermentation processes appear to be predominated by Clostridia from the genus Alkaliphilus sp, with methanogenesis being attributed to Methanobacterium and Methanomassiliicoccus sp. The study is the first to identify the generation of ISA within an anthropogenic environment and advocates the notion that microbial activity within an ILW-GDF is likely to influence the impact of ISAs upon radionuclide migration.     

Microbial Investigations in Opalinus Clay,an Argillaceous Formation under Evaluation as a Potential Host Rock for a Radioactive Waste Repository

Various deep, compact, sedimentary formations have been studied in recent years as potential host rock for a repository for high-level, long-lived radioactive waste. Considering that microbial activities may influence radionuclide chemistry and migration in such environments, we investigated the potential presence of microorganisms in the Opalinus Clay formation, from unperturbed sediment samples (i.e., not affected by gallery excavation and borehole drilling) recovered under aseptic conditions in the Mont Terri Underground Rock Laboratory (Switzerland). A combination of molecular biology techniques and a cultivation-based approach suggested the presence of a few sparse autochthonous microbial cells in the Opalinus Clay. For the first time, ribosomal RNA (rRNA) genes were sequenced from enrichment cultures from such samples. The results suggested that at least two of the bacterial strains isolated were likely unknown species of the Sphingomonas and Alicyclobacillus genera, as their fully-sequenced 16S-rRNA genes shared less than 97% similarity with validly published sequences. Early genetic divergence occurring after physical isolation of bacterial ancestors in the geosphere by the sedimentation process or following later geological events may have resulted in the generation of particular taxa in the subsurface.     

Microbial Community Evolution Is Significantly Impacted by the Use of Calcium Isosaccharinic Acid as an Analogue for the Products of Alkaline Cellulose Degradation

Diasteriomeric isosaccharinic acid (ISA) is an important consideration within safety assessments for the disposal of the United Kingdoms’ nuclear waste legacy, where it may potentially influence radionuclide migration. Since the intrusion of micro-organisms may occur within a disposal concept, the impact of ISA may be impacted by microbial metabolism. Within the present study we have established two polymicrobial consortia derived from a hyperalkaline soil. Here, α-ISA and a diatereomeric mix of ISAs’ were used as a sole carbon source, reflecting two common substrates appearing within the literature. The metabolism of ISA within these two consortia was similar, where ISA degradation resulted in the acetogenesis and hydrogenotrophic methanogenesis. The chemical data obtained confirm that the diastereomeric nature of ISA is likely to have no impact on its metabolism within alkaline environments. High throughput sequencing of the original soil showed a diverse community which, in the presence of ISA allowed for the dominance the Clostridiales associated taxa with Clostridium clariflavum prevalent. Further taxonomic investigation at the genus level showed that there was in fact a significant difference (p = 0.004) between the two community profiles. Our study demonstrates that the selection of carbon substrate is likely to have a significant impact on microbial community composition estimations, which may have implications with respect to a safety assessment of an ILW-GDF.     

Niche Partitioning of Microbial Communities at an Ancient Vitrified Hillfort: Implications for Vitrified Radioactive Waste Disposal

Abstract

Because microbes cannot be eliminated from radioactive waste disposal facilities, the consequences of bio-colonization must be understood. At a pre-Viking era vitrified hillfort, Broborg, Sweden, anthropogenic glass has been subjected to bio-colonization for over 1,500?years. Broborg is used as a habitat analogue for disposed radioactive waste glass to inform how microbial processes might influence long-term glass durability. Electron microscopy and DNA sequencing of surficial material from the Broborg vitrified wall, adjacent soil, and general topsoil show that the ancient glass supports a niche microbial community of bacteria, fungi, and protists potentially involved in glass alteration. Communities associated with the vitrified wall are distinct and less diverse than soil communities. The vitrified niche of the wall and adjacent soil are dominated by lichens, lichen-associated microbes, and other epilithic, endolithic, and epigeic organisms. These organisms exhibit potential bio-corrosive properties, including silicate dissolution, extraction of essential elements, and secretion of geochemically reactive organic acids, that could be detrimental to glass durability. However, long-term biofilms can also possess a homeostatic function that could limit glass alteration. This study documents potential impacts that microbial colonization and niche partitioning can have on glass alteration, and subsequent release of radionuclides from a disposal facility for vitrified radioactive waste.     

Investigation of Microorganisms in Bentonite Deposits

Microbiological characteristics of bentonite deposits were investigated as a natural analogue of microbial behavior in the buffer material for geological disposal of radioactive waste. Distributions of microorganisms in bentonite were examined at four sites in two different bentonite deposits in Japan. The sites included pond bottom, wetland, and wet mine gallery environments where bentonite layers have been left undisturbed for 2 to 30 years. Excavation was performed without using drilling water and the center parts of the cores were used for microbial examination. Plate counts with R2A medium of aerobic and anaerobic bacteria at the drilling mouth ranged from 10⁵ to 10⁷/g DW (dry weight) and from 10³ to 10⁶/g DW, respectively. The CFDA-AM (Carboxyfluorescein diacetate acetoxymethyl ester) cell counts ranged from 10⁶ to 10⁹/g DW. Bacterial numbers in the bentonite layers declined with distance from the drilling mouth; both aerobes and anaerobes were less than 10²/g DW and CFDA-AM cell counts less than 10⁶/g DW for core samples taken from approximately 1 m depth, except at the pond bottom. These results suggest that microbial activity in natural bentonite is lower than in typical soils and aquatic sediments and does not spread easily.     

Microbial degradation of isosaccharinic acid at high pH

Intermediate-level radioactive waste (ILW), which dominates the radioactive waste inventory in the United Kingdom on a volumetric basis, is proposed to be disposed of via a multibarrier deep geological disposal facility (GDF). ILW is a heterogeneous wasteform that contains substantial amounts of cellulosic material encased in concrete. Upon resaturation of the facility with groundwater, alkali conditions will dominate and will lead to the chemical degradation of cellulose, producing a substantial amount of organic co-contaminants, particularly isosaccharinic acid (ISA). ISA can form soluble complexes with radionuclides, thereby mobilising them and posing a potential threat to the surrounding environment or ‘far field''. Alkaliphilic microorganisms sampled from a legacy lime working site, which is an analogue for an ILW-GDF, were able to degrade ISA and couple this degradation to the reduction of electron acceptors that will dominate as the GDF progresses from an aerobic ‘open phase'' through nitrate- and Fe(III)-reducing conditions post closure. Furthermore, pyrosequencing analyses showed that bacterial diversity declined as the reduction potential of the electron acceptor decreased and that more specialised organisms dominated under anaerobic conditions. These results imply that the microbial attenuation of ISA and comparable organic complexants, initially present or formed in situ, may play a role in reducing the mobility of radionuclides from an ILW-GDF, facilitating the reduction of undue pessimism in the long-term performance assessment of such facilities.     

Microbial communities from weathered outcrops of a sulfide-rich ultramafic intrusion,and implications for mine waste management

The Duluth Complex (DC) contains sulfide-rich magmatic intrusions that represent one of the largest known economic deposits of copper, nickel, and platinum group elements. Previous work showed that microbial communities associated with experimentally-weathered DC waste rock and tailings were dominated by uncultivated taxa and organisms not typically associated with mine waste. However, those experiments were designed for kinetic testing and do not necessarily represent the conditions expected for long-term environmental weathering. We used 16S rRNA gene methods to characterize the microbial communities present on the surfaces of naturally-weathered and historically disturbed outcrops of DC material. Rock surfaces were dominated by diverse uncultured Ktedonobacteria, Acetobacteria, and Actinobacteria, with abundant algae and other phototrophs. These communities were distinct from microbial assemblages from experimentally-weathered DC rocks, suggesting different energy and nutrient resources in environmental samples. Sulfide mineral incubations performed with and without algae showed that photosynthetic microorganisms could have an inhibitory effect on autotrophic populations, resulting in slightly lower sulfate release and differences in dominant microorganisms. The microbial assemblages from these weathered outcrops show how communities develop during weathering of sulfide-rich DC rocks and represent baseline data that could evaluate the effectiveness of future reclamation of waste produced by large-scale mining operations.     

Treatment of Alkaline Cr(VI)-Contaminated Leachate with an Alkaliphilic Metal-Reducing Bacterium

Chromium in its toxic Cr(VI) valence state is a common contaminant particularly associated with alkaline environments. A well-publicized case of this occurred in Glasgow, United Kingdom, where poorly controlled disposal of a cementitious industrial by-product, chromite ore processing residue (COPR), has resulted in extensive contamination by Cr(VI)-contaminated alkaline leachates. In the search for viable bioremediation treatments for Cr(VI), a variety of bacteria that are capable of reduction of the toxic and highly soluble Cr(VI) to the relatively nontoxic and less mobile Cr(III) oxidation state, predominantly under circumneutral pH conditions, have been isolated. Recently, however, alkaliphilic bacteria that have the potential to reduce Cr(VI) under alkaline conditions have been identified. This study focuses on the application of a metal-reducing bacterium to the remediation of alkaline Cr(VI)-contaminated leachates from COPR. This bacterium, belonging to the Halomonas genus, was found to exhibit growth concomitant to Cr(VI) reduction under alkaline conditions (pH 10). Bacterial cells were able to rapidly remove high concentrations of aqueous Cr(VI) (2.5 mM) under anaerobic conditions, up to a starting pH of 11. Cr(VI) reduction rates were controlled by pH, with slower removal observed at pH 11, compared to pH 10, while no removal was observed at pH 12. The reduction of aqueous Cr(VI) resulted in the precipitation of Cr(III) biominerals, which were characterized using transmission electron microscopy and energy-dispersive X-ray analysis (TEM-EDX) and X-ray photoelectron spectroscopy (XPS). The effectiveness of this haloalkaliphilic bacterium for Cr(VI) reduction at high pH suggests potential for its use as an in situ treatment of COPR and other alkaline Cr(VI)-contaminated environments.     

Metal Sorption to Pseudomonas fluorescens: Influence of pH,Ionic Strength and Metal Concentrations

Migration behavior of radionuclides should be understood in order to estimate the impact of high-level radioactive waste (HLW) disposal on the environment. Bacteria, one of the major organic sorbents in solid and aquatic environments, can affect the fate of actinides and lanthanides by sorption onto their cell surfaces. In this study, the sorption of the radionuclide Americium (Am(III)) and several metal ions (Eu(III), Cu(II) and Ca(II)) to Pseudomonas fluorescens were measured under various conditions. It was revealed that as pH decreased, the sorption of Eu(III) and Am(III) increased when the metals were at relatively low concentrations but decreased at higher metal concentrations. On the other hand, sorption of Cu(II) followed the opposite trend. In the case of calcium, an increase in calcium ions was observed due to release from the cells. These findings suggest that the sorption mechanisms for low levels of Eu(III) and Am(III) on the cells of Pseudomonas fluorescens are different from those of Cu(II), Ca(II), and high concentrations of Eu(III) (> 10 ^{? 5} M).     

The Synergistic Effects of High Nitrate Concentrations on Sediment Bioreduction

Groundwaters at nuclear sites can be characterized by low pH and high nitrate concentrations (10–100 mM). These conditions are challenging for bioremediation, often inhibiting microbial Fe(III)-reduction which can limit radionuclide migration. Here, sediment microcosms representative of the UK Sellafield site were used to study the influence of variable pH and nitrate concentrations on microbially-mediated TEAP (terminal electron accepting processes) progression. The rate of reduction through the terminal electron accepting cascade NO^? ₃ > NO^? ₂ > Mn(IV)/Fe(III) > SO^2? ₄ at low pH (～5.5) was slower than that in bicarbonate buffered systems (pH ～ 7.0), but in the low pH systems, denitrification and associated pH buffering resulted in conditioning of the sediments for subsequent Fe(III) and sulfate-reduction. Under very high nitrate conditions (100 mM), bicarbonate buffering (pH ～ 7.0) was necessary for TEAP progression beyond denitrification and the reduction of 100 mM nitrate created alkaline conditions (pH 9.5). 16S rRNA gene analysis showed that close relatives of known nitrate reducers Bacillus niacini and Ochrobactrum grignonense dominated the microbial communities in this reduced sediment. In Fe(III)-reducing enrichment cultures from the 100 mM nitrate system, close relatives of the Fe(III)-reducing species Alkaliphilus crotonatoxidans and Serratia liquifaciens were observed. These results highlight that under certain conditions and contrary to expectations, denitrification may support bioreduction via pH conditioning for optimal metal reduction and radionuclide immobilization.     

Microbial corrosion monitoring by an amperometric microbial biosensor developed using whole cell of Pseudomonas sp.

A microbial biosensor was developed for monitoring microbiologically influenced corrosion (MIC) of metallic materials in industrial systems. The Pseudomonas sp. isolated from corroded metal surface was immobilized on acetylcellulose membrane and its respiratory activity was estimated by measuring oxygen consumption. The microbial biosensor was used for the measurement of sulfuric acid in a batch culture medium contaminated by microorganisms. A linear relationship between the microbial sensor response and the concentration of sulfuric acid was observed. The response time of biosensor was 5 min and was dependent on the immobilized cell loading of Pseudomonas sp., pH, temperature and corrosive environments. The microbial biosensor response was stable, reproducible and specific for sensing of sulfur oxidizing bacterial activity.