Michal Ziv-El

Systematic evaluation of nitrate and perchlorate bioreduction kinetics in groundwater using a hydrogen-based membrane biofilm reactor

To evaluate the simultaneous reduction kinetics of the oxidized compounds, we treated nitrate-contaminated groundwater (approximately 9.4 mg-N/L) containing low concentrations of perchlorate (approximately 12.5 microg/L) and saturated with dissolved oxygen (approximately 8 mg/L) in a hydrogen-based membrane biofilm reactor (MBfR). We systematically increased the hydrogen availability and simultaneously varied the surface loading of the oxidized compounds on the biofilm in order to provide a comprehensive, quantitative data set with which to evaluate the relationship between electron donor (H(2)) availability, surface loading of the electron acceptors (oxidized compounds), and simultaneous bioreduction of the electron acceptors. Increasing the H(2) pressure delivered more H(2) gas, and the total H(2) flux increased linearly from approximately 0.04 mg/cm(2)-d for 0.5 psig (0.034 atm) to 0.13 mg/cm(2)-d for 9.5 psig (0.65 atm). This increased rate of H(2) delivery allowed for continued reduction of the acceptors as their surface loading increased. The electron acceptors had a clear hydrogen-utilization order when the availability of hydrogen was limited: oxygen, nitrate, nitrite, and then perchlorate. Spiking the influent with perchlorate or nitrate allowed us to identify the maximum surface loadings that still achieved more than 99.5% reduction of both oxidized contaminants: 0.21 mg NO(3)-N/cm(2)-d and 3.4 microg ClO(4)/cm(2)-d. Both maximum values appear to be controlled by factors other than hydrogen availability.

Interactions between Nitrate-Reducing and Sulfate-Reducing Bacteria Coexisting in a Hydrogen-Fed Biofilm

To explore the relationships between denitrifying bacteria (DB) and sulfate-reducing bacteria (SRB) in H(2)-fed biofilms, we used two H(2)-based membrane biofilm reactors (MBfRs) with or without restrictions on H(2) availability. DB and SRB compete for H(2) and space in the biofilm, and sulfate (SO(4)(2-)) reduction should be out-competed when H(2) is limiting inside the biofilm. With H(2) availability restricted, nitrate (NO(3)(-)) reduction was proportional to the H(2) pressure and was complete at a H(2) pressure of 3 atm; SO(4)(2-) reduction began at H(2) ≥ 3.4 atm. Without restriction on H(2) availability, NO(3)(-) was the preferred electron acceptor, and SO(4)(2-) was reduced only when the NO(3)(-) surface loading was ≤ 0.13 g N/m(2)-day. We assayed DB and SRB by quantitative polymerase chain reaction targeting the nitrite reductases and dissimilatory sulfite reductase, respectively. Whereas DB and SRB increased with higher H(2) pressures when H(2) availability was limiting, SRB did not decline with higher NO(3)(-) removal flux when H(2) availability was not limiting, even when SO(4)(2-) reduction was absent. The SRB trend reflects that the SRBs metabolic diversity allowed them to remain in the biofilm whether or not they were reducing SO(4)(2-). In all scenarios tested, the SRB were able to initiate strong SO(4)(2-) reduction only when competition for H(2) inside the biofilm was relieved by nearly complete removal of NO(3)(-).

Effect of dechlorination and sulfate reduction on the microbial community structure in denitrifying membrane-biofilm reactors

Recent studies showed that the chlorinated solvents trichloroethene (TCE), 1,1,1-trichloroethane (TCA), and chloroform (CF) were reductively dehalogenated in a H(2)-based membrane biofilm reactor (MBfR) under denitrifying conditions. Here, we describe a detailed phylogenetic characterization of MBfR biofilm communities having distinctly different metabolic functions with respect to electron-acceptor reduction. Using massively parallel pyrosequencing of the V6 region of the 16S rRNA gene, we detected 312, 592, and 639 operational taxonomic units (OTU) in biofilms of three MBfRs that reduced nitrate; nitrate and TCE; or nitrate, sulfate, and all three chlorinated solvents. Comparative community analysis revealed that 13% of the OTUs were shared by all MBfRs, regardless of the feed, but 65% were unique to one MBfR. Pyrosequencing and real-time quantitative PCR showed that Dehalococcoides were markedly enriched in the TCE+nitrate biofilm. The input of a mixture of three chlorinated compounds, which coincided with the onset of sulfate reduction, led to a more diverse community that included sulfate-reducing bacteria (Desulfovibrio) and nitrate-reducing bacteria (Geothrix and Pseudomonas). Our results suggest that chlorinated solvents, as additional electron acceptors to nitrate and sulfate, increased microbial diversity by allowing bacteria with special metabolic capabilities to grow in the biofilm.

Managing methanogens and homoacetogens to promote reductive dechlorination of trichloroethene with direct delivery of H2 in a membrane biofilm reactor

A study with H2‐based membrane biofilm reactors (MBfRs) was undertaken to examine the effectiveness of direct H2 delivery in ex‐situ reductive dechlorination of chlorinated ethenes. Trichloroethene (TCE) could be reductively dechlorinated to ethene with up to 95% efficiency as long as the pH‐increase effects of methanogens and homoacetogens were managed and dechlorinators were selected for during start‐up by creating H2 limitation. Based on quantitative PCR, the dominant bacterial groups in the biofilm at the end of reactor operation were Dehalococcoides, Geobacter, and homoacetogens. Pyrosequencing confirmed the dominance of the dechlorinators and identified Acetobacterium as the key homoacetogen. Homoacetogens outcompeted methanogens for bicarbonate, based on the effluent concentration of acetate, by suppressing methanogens during batch start‐up. This was corroborated by the methanogenesis functional gene mcrA, which was 1–2 orders of magnitude lower than the FTHFS functional gene for homoacetogens. Imaging of the MBfR fibers using scanning electron microscopy showed a distinct Dehalococcoides‐like morphology in the fiber biofilm. These results support that direct addition of H2 can allow for efficient and complete reductive dechlorination, and they shed light into how H2‐fed biofilms, when operated to manage methanogenic and homoacetogenic activity, can be used for ex‐situ bioremediation of chlorinated ethenes. Biotechnol. Bioeng. 2012;109: 2200–2210.

A pH-control model for heterotrophic and hydrogen-based autotrophic denitrification

This work presents a model to predict the alkalinity, pH, and Langelier Saturation Index (LSI) in heterotrophic and H(2)-based autotrophic denitrification systems. The model can also be used to estimate the amount of acid, e.g. HCl, added to the influent (method 1) or the pH set point in the reactor (method 2: pH can be maintained stable by CO(2)-sparge using a pH-control loop) to prevent the pH from exceeding the optimal range for denitrification and to prevent precipitation from occurring. The model was tested with two pilot plants carrying out denitrification of groundwater with high hardness: a heterotrophic system using ethanol as the electron donor and an H(2)-based autotrophic system. The measured alkalinity, pH, and LSI were consistent with the model for both systems. This work also quantifies: (1) how the alkalinity and pH in Stage-1 significantly differ from those in Stage-2; (2) how the pH and LSI differ significantly in the two denitrification systems while the alkalinity increase is about the same; and (3) why CO(2) addition is the preferred method for autotrophic system, while HCl addition is the preferred method for the heterotrophic system.

Using electron balances and molecular techniques to assess trichoroethene-induced shifts to a dechlorinating microbial community

This study demonstrated the utility in correlating performance and community structure of a trichloroethene (TCE)‐dechlorinating microbial consortium; specifically dechlorinators, fermenters, homoacetogens, and methanogens. Two complementary approaches were applied: predicting trends in the microbial community structure based on an electron balance analysis and experimentally assessing the community structure via pyrosequencing and quantitative polymerase chain reaction (qPCR). Fill‐and‐draw reactors inoculated with the DehaloR⁁2 consortium were operated at five TCE‐pulsing rates between 14 and 168 µmol/10‐day‐SRT, amended with TCE every 2 days to give peak concentrations between 0.047 and 0.56 mM (6–74 ppm) and supplied lactate and methanol as sources of e‐ donor and carbon. The complementary approaches demonstrated the same trends: increasing abundance of Dehalococcoides and Geobacter and decreasing abundance of Firmicutes with increasing TCE pulsing rate, except for the highest pulsing rate. Based on qPCR, the abundance of Geobacter and Dehalococcoides decreased for the highest TCE pulsing rate, and pyrosequencing showed this same trend for the latter. This deviation suggested decoupling of Dehalococcoides growth from dechlorination. At pseudo steady‐state, methanogenesis was minimal for all TCE pulsing rates. Pyrosequencing and qPCR showed suppression of the homoacetogenic genera Acetobacterium at the two highest pulsing rates, and it was corroborated by a decreased production of acetate from lactate fermentation and increased propionate production. Suppression of Acetobacterium, which can provide growth factors to Dehalococcoides, may have contributed to the decoupling for the highest TCE‐pulsing rate. Biotechnol. Bioeng. 2012;109: 2230–2239.

Simultaneous Determination of Chlorinated Ethenes and Ethene in Groundwater Using Headspace Solid-Phase Microextraction with Gas Chromatography

Widespread contamination of groundwater by chlorinated ethenes and their biological dechlorination products necessitates the reliable monitoring of liquid matrices; current methods approved by the U.S. Environmental Protection Agency (EPA) require a minimum of 5 mL of sample volume and cannot simultaneously detect all transformative products. This paper reports on the simultaneous detection of six chlorinated ethenes and ethene itself, using a liquid sample volume of 1 mL by concentrating the compounds onto an 85-µm carboxen-polydimenthylsiloxane solid-phase microextraction fiber in 5 min and subsequent chromatographic analysis in 9.15 min. Linear increases in signal response were obtained over three orders of magnitude (∼0.05 to ∼50 µM) for simultaneous analysis with coefficient of determination (R(2)) values of ≥ 0.99. The detection limits of the method (1.3-6 µg/L) were at or below the maximum contaminant levels specified by the EPA. Matrix spike studies with groundwater and mineral medium showed recovery rates between 79-108%. The utility of the method was demonstrated in lab-scale sediment flow-through columns assessing the bioremediation potential of chlorinated ethene-contaminated groundwater. Owing to its low sample volume requirements, good sensitivity and broad target analyte range, the method is suitable for routine compliance monitoring and is particularly attractive for interpreting the bench-scale feasibility studies that are commonly performed during the remedial design stage of groundwater cleanup projects.