Roger L. Ely

An Evaluation of Aerobic Trichloroethene Attenuation Using First-Order Rate Estimation

Natural attenuation of trichloroethene (TCE) was evaluated for a groundwater plume at the Idaho National Engineering and Environmental Laboratory. Significant evidence demonstrated that reductive dechlori-nation is occurring, but is limited to a small area around the contamination source. In spite of this, the plume is relatively stable. Three first-order rate estimation methods were used to help understand transport processes affecting TCE in the large, aerobic portion of the plume. Two of the methods gave attenuation half-life estimates for TCE of approximately 8 years; however, these methods do not adequately distinguish between degradation and dispersion. The third method showed TCE attenuation relative to the co-contaminants, tritium and tetrachloroethene (PCE), and used these “tracers” to distinguish between dispersion and degradation. The estimated aerobic degradation half-life for TCE was between 13 and 21 years. Aerobic cometabolism of TCE has been identified as a potential mechanism for the apparent degradation. The importance of distinguishing between dispersion and degradation was shown using an analytical model. The model demonstrated that, in general, the rate of contaminant concentration decrease due to dispersion is not constant with time after the source is removed. This has important implications for predicting the long-term effectiveness of natural attenuation for groundwater restoration.

Silica sol‐gel encapsulation of cyanobacteria: lessons for academic and applied research

Cyanobacteria inhabit nearly every ecosystem on earth, play a vital role in nutrient cycling, and are useful as model organisms for fundamental research in photosynthesis and carbon and nitrogen fixation. In addition, they are important for several established biotechnologies for producing food additives, nutritional and pharmaceutical compounds, and pigments, as well as emerging biotechnologies for biofuels and other products. Encapsulation of living cyanobacteria into a porous silica gel matrix is a recent approach that may dramatically improve the efficiency of certain production processes by retaining the biomass within the reactor and modifying cellular metabolism in helpful ways. Although encapsulation has been explored empirically in the last two decades for a variety of cell types, many challenges remain to achieving optimal encapsulation of cyanobacteria in silica gel. Recent evidence with Synechocystis sp. PCC 6803, for example, suggests that several unknown or uncharacterized proteins are dramatically upregulated as a result of encapsulation. Also, additives commonly used to ease stresses of encapsulating living cells, such as glycerol, have detrimental impacts on photosynthesis in cyanobacteria. This mini-review is intended to address the current status of research on silica sol-gel encapsulation of cyanobacteria and research areas that may further the development of this approach for biotechnology applications.

High-throughput screening assay for biological hydrogen production.

This paper describes a screening assay, compatible with high-throughput bioprospecting or molecular biology methods, for assessing biological hydrogen (H2) production. While the assay is adaptable to various physical configurations, we describe its use in a 96-well, microtiter plate format with a lower plate containing H2-producing cyanobacteria strains and controls and an upper, membrane-bottom plate containing a color indicator and a catalyst. H2 produced by cells in the lower plate diffuses through the membrane into the upper plate, causing a color change that can be quantified with a microplate reader. The assay is reproducible, semiquantitative, sensitive down to at least 20 nmol of H2, and largely unaffected by oxygen, carbon dioxide, or volatile fatty acids at levels appropriate to biological systems.

Optimization of pH and nitrogen for enhanced hydrogen production by Synechocystis sp. PCC 6803 via statistical and machine learning methods.

The nitrogen (N) concentration and pH of culture media were optimized for increased fermentative hydrogen (H2) production from the cyanobacterium, Synechocystis sp. PCC 6803. The optimization was conducted using two procedures, response surface methodology (RSM), which is commonly used, and a memory‐based machine learning algorithm, Q2, which has not been used previously in biotechnology applications. Both RSM and Q2 were successful in predicting optimum conditions that yielded higher H2 than the media reported by Burrows et al., Int J Hydrogen Energy. 2008;33:6092–6099 optimized for N, S, and C (called EHB‐1 media hereafter), which itself yielded almost 150 times more H2 than Synechocystis sp. PCC 6803 grown on sulfer‐free BG‐11 media. RSM predicted an optimum N concentration of 0.63 mM and pH of 7.77, which yielded 1.70 times more H2 than EHB‐1 media when normalized to chlorophyll concentration (0.68 ± 0.43 μmol H2 mg Chl−1 h−1) and 1.35 times more when normalized to optical density (1.62 ± 0.09 nmol H2 OD730−1 h−1). Q2 predicted an optimum of 0.36 mM N and pH of 7.88, which yielded 1.94 and 1.27 times more H2 than EHB‐1 media when normalized to chlorophyll concentration (0.77 ± 0.44 μmol H2 mg Chl−1 h−1) and optical density (1.53 ± 0.07 nmol H2 OD730−1 h−1), respectively. Both optimization methods have unique benefits and drawbacks that are identified and discussed in this study.

Whole-genome transcriptional and physiological responses of Nitrosomonas europaea to cyanide: identification of cyanide stress response genes.

Nitrosomonas europaea (ATCC 19718) is one of several nitrifying species that participate in the biological removal of nitrogen from wastewater by oxidizing ammonia to nitrite, the first step in nitrification. Because nitrification is quite sensitive to cyanide, a compound often encountered in wastewater treatment plants, we characterized the physiological and transcriptional responses of N. europaea cells to cyanide. The cells were extremely sensitive to low concentrations of cyanide, with

Thermotolerant Hydrogenases: Biological Diversity, Properties, and Biotechnological Applications

{\rm NO}_2^ -

Effects of selected electron transport chain inhibitors on 24-h hydrogen production by Synechocystis sp. PCC 6803.

production and ammonia‐dependent oxygen uptake rates decreasing by 50% within 30 min of exposure to 1 µM NaCN. Whole‐genome transcriptional responses of cells exposed to 1 µM NaCN were examined using Affymetrix microarrays to identify stress‐induced genes. The transcript levels of 35 genes increased more than 2‐fold while transcript levels of 29 genes decreased more than 20‐fold. A gene cluster that included moeZ (NE2353), encoding a rhodanese homologue and thought to be involved in detoxification of cyanide, showed the highest up‐regulation (7‐fold). The down‐regulated genes included genes encoding proteins involved in the sulfate reduction pathway, signal transduction mechanisms, carbohydrate transport, energy production, coenzyme metabolism, and amino acid transport. Biotechnol. Bioeng. 2009;102: 1645–1653.

Diffusion of dissolved ions from wet silica sol–gel monoliths: Implications for biological encapsulation

Hydrogenases are metalloproteins that catalyze the oxidation and reduction of molecular hydrogen and play a crucial role in many microbial metabolic processes. A subset of hydrogenases capable of functioning at temperatures from 50 to 125°C is found in thermophilic microorganisms. Most known thermotolerant hydrogenases contain a [NiFe] active site and are either bidirectional or uptake type. Although no exhaustive survey has been done of the ecological diversity of thermophilic hydrogen-reducing or oxidizing bacteria, they appear to exist in virtually every thermophilic environment examined to date. Thermotolerant hydrogenases share many similarities with their mesophilic counterparts, but they have several features in addition to thermotolerance that make them especially well suited for biotechnological applications. Ongoing research is focused on potential applications of thermotolerant H2ases in biosynthesis, H2 production, bioremediation, and biosensors.

Transcriptional and Physiological Responses of Nitrosococcus mobilis to Copper Exposure

One factor limiting biosolar hydrogen (H(2)) production from cyanobacteria is electron availability to the hydrogenase enzyme. In order to optimize 24-h H(2) production this study used Response Surface Methodology and Q2, an optimization algorithm, to investigate the effects of five inhibitors of the photosynthetic and respiratory electron transport chains of Synechocystis sp. PCC 6803. Over 3 days of diurnal light/dark cycling, with the optimized combination of 9.4 mM KCN (3.1 μmol 10(10) cells(-1)) and 1.5 mM malonate (0.5 μmol 10(10) cells(-1)) the H(2) production was 30-fold higher, in EHB-1 media previously optimized for nitrogen (N), sulfur (S), and carbon (C) concentrations (Burrows et al., 2008). In addition, glycogen concentration was measured over 24 h with two light/dark cycling regimes in both standard BG-11 and EHB-1 media. The results suggest that electron flow as well as glycogen accumulation should be optimized in systems engineered for maximal H(2) output.

Comparison of Artificial Neural Network, Genetic Programming, and Mechanistic Modeling of Complex Biological Processes

Divalent nickel (Ni(2+)), Cu(II)EDTA, methyl orange, and dichromate were used to investigate diffusion from hydrated silica sol-gel monoliths. The objective was to examine diffusion of compounds on a size regime relevant to supporting biological components encapsulated within silica gel prepared in a biologically compatible process space with no post-gelation treatments. With an initial sample set, gels prepared from tetraethoxysilane were explored in a factorial design with Ni(2+) as the tracer, varying water content during hydrolysis, acid catalyst present during hydrolysis, and the final concentration of silica. A second sample set explored diffusion of all four tracers in gels prepared with aqueous silica precursors and a variety of organically modified siloxanes. Excluding six outliers which displayed significant syneresis, the mean diffusion constant (D(gel)) across the entire process space of sample set 1 was 2.42×10(-10) m(2) s(-1); approximately 24% of the diffusion coefficient of Ni(2+) in unconfined aqueous solution. In sample set 2, the tracer size and not gel hydrophobicity was the primary determinant of changes in diffusion rates. A strong linear inverse correlation was found between tracer size and the magnitude of D(gel). Based on correlation with the tracers used in this investigation, the characteristic 1-h diffusion distance for carbonate species relevant to supporting active phototrophic organisms was approximately 1.5mm. These results support the notion that silica sol-gel formulations may be optimized for a given biological entity of interest with manageable impact to the diffusion of small ions and molecules.