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 H₂-fed biofilms, we used two H₂-based membrane biofilm reactors (MBfRs) with or without restrictions on H₂ availability. DB and SRB compete for H₂ and space in the biofilm, and sulfate (SO₄^2–) reduction should be out-competed when H₂ is limiting inside the biofilm. With H₂ availability restricted, nitrate (NO₃^–) reduction was proportional to the H₂ pressure and was complete at a H₂ pressure of 3 atm; SO₄^2– reduction began at H₂ ≥ 3.4 atm. Without restriction on H₂ availability, NO₃^– was the preferred electron acceptor, and SO₄^2– was reduced only when the NO₃^– surface loading was ≤0.13 g N/m²-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 H₂ pressures when H₂ availability was limiting, SRB did not decline with higher NO₃^– removal flux when H₂ availability was not limiting, even when SO₄^2– reduction was absent. The SRB trend reflects that the SRB’s metabolic diversity allowed them to remain in the biofilm whether or not they were reducing SO₄^2–. In all scenarios tested, the SRB were able to initiate strong SO₄^2– reduction only when competition for H₂ inside the biofilm was relieved by nearly complete removal of NO₃^–

Effects of Multiple Electron Acceptors on Microbial Interactions in a Hydrogen-Based Biofilm

To investigate interactions among multiple electron acceptors in a H₂-fed biofilm, we operated a membrane biofilm reactor with H₂-delivery capacity sufficient to reduce all acceptors. ClO₄^– and O₂ were input electron acceptors in all stages at surface loadings of 0.08 ± 0.006 g/m²-d (1.0 ± 0.7 e^– meq/m²-d) for ClO₄^– and 0.51 g/m²-d (76 e^– meq/m²-d) for O₂. SO₄^2– was added in Stage 2 at 3.77 ± 0.39 g/m²-d (331 ± 34 e^– meq/m²-d), and NO₃^– was further added in Stage 3 at 0.72 ± 0.03 g N/m²-d (312 ± 13 e^– meq/m²-d). At steady state for each stage, ClO₄^–, O₂, and NO₃^– (when present in the influent) were completely reduced; measured SO₄^2– reduction decreased from 78 ± 4% in Stage 2 to 59 ± 4% in Stage 3, when NO₃^– was present. While perchlorate-reducing bacteria (PRB), assayed by qPCR targeting the <i>pcrA</i> gene, remained stable throughout, sulfate-reducing bacteria (SRB), assayed by the <i>dsrA</i> gene, increased almost 3 orders of magnitude when significant SO₄^2– reduction occurred in stage 2. The abundance of denitrifying bacteria (DB), assayed by the <i>nirK</i> and <i>nirS</i> genes, increased in Stage 3, while SRB remained at high numbers, but did not increase. Based on pyrosequencing analyses, <i>β-Proteobacteria</i> dominated in Stage 1, but <i>ε-Proteobacteria</i> became more important in Stages 2 and 3, when the input of multiple electron acceptors favored genera with broader electron-accepting capabilities. <i>Sulfuricurvum</i> (a sulfur oxidizer and NO₃^– reducer) and <i>Desulfovibrio</i> (a SO₄^2– reducer) become dominant in Stage 3, suggesting redox cycling of sulfur in the biofilm

Palladium Recovery in a H₂‑Based Membrane Biofilm Reactor: Formation of Pd(0) Nanoparticles through Enzymatic and Autocatalytic Reductions

Recovering palladium (Pd) from waste streams opens up the possibility of augmenting the supply of this important catalyst. We evaluated Pd reduction and recovery as a novel application of a H₂-based membrane biofilm reactor (MBfR). At steady states, over 99% of the input soluble Pd­(II) was reduced through concomitant enzymatic and autocatalytic processes at acidic or near neutral pHs. Nanoparticulate Pd(0), at an average crystallite size of 10 nm, was recovered with minimal leaching and heterogeneously associated with microbial cells and extracellular polymeric substances in the biofilm. The dominant phylotypes potentially responsible for Pd­(II) reduction at circumneutral pH were denitrifying β-proteobacteria mainly consisting of the family <i>Rhodocyclaceae</i>. Though greatly shifted by acidic pH, the biofilm microbial community largely bounced back when the pH was returned to 7 within 2 weeks. These discoveries infer that the biofilm was capable of rapid adaptive evolution to stressed environmental change, and facilitated Pd recovery in versatile ways. This study demonstrates the promise of effective microbially driven Pd recovery in a single MBfR system that could be applied for the treatment of the waste streams, and it documents the role of biofilms in this reduction and recovery process

Using a Two-Stage Hydrogen-Based Membrane Biofilm Reactor (MBfR) to Achieve Complete Perchlorate Reduction in the Presence of Nitrate and Sulfate

We evaluated a strategy for achieving complete reduction of perchlorate (ClO₄^–) in the presence of much higher concentrations of sulfate (SO₄^2–) and nitrate (NO₃^–) in a hydrogen-based membrane biofilm reactor (MBfR). Full ClO₄^– reduction was achieved by using a two-stage MBfR with controlled NO₃^– surface loadings to each stage. With an equivalent NO₃^– surface loading larger than 0.65 ± 0.04 g N/m²-day, the lead MBfR removed about 87 ± 4% of NO₃^– and 30 ± 8% of ClO₄^–. This decreased the equivalent surface loading of NO₃^– to 0.34 ± 0.04–0.53 ± 0.03 g N/m²-day for the lag MBfR, in which ClO₄^– was reduced to nondetectable. SO₄^2– reduction was eliminated without compromising full ClO₄^– reduction using a higher flow rate that gave an equivalent NO₃^– surface loading of 0.94 ± 0.05 g N/m²-day in the lead MBfR and 0.53 ± 0.03 g N/m²-day in the lag MBfR. Results from qPCR and pyrosequencing showed that the lead and lag MBfRs had distinctly different microbial communities when SO₄^2– reduction took place. Denitrifying bacteria (DB), quantified using the <i>nirS</i> and <i>nirK</i> genes, dominated the biofilm in the lead MBfR, but perchlorate-reducing bacteria (PRB), quantified using the <i>pcrA</i> gene, became more important in the lag MBfR. The facultative anaerobic bacteria <i>Dechloromonas</i>, <i>Rubrivivax</i>, and <i>Enterobacter</i> were dominant genera in the lead MBfR, where their main function was to reduce NO₃^–. With a small NO₃^– surface loading and full ClO₄^– reduction, the dominant genera shifted to ClO₄^–-reducing bacteria <i>Sphaerotilus</i>, <i>Rhodocyclaceae</i>, and <i>Rhodobacter</i> in the lag MBfR

Pyrosequencing Analysis Yields Comprehensive Assessment of Microbial Communities in Pilot-Scale Two-Stage Membrane Biofilm Reactors

We studied the microbial community structure of pilot two-stage membrane biofilm reactors (MBfRs) designed to reduce nitrate (NO₃^–) and perchlorate (ClO₄^–) in contaminated groundwater. The groundwater also contained oxygen (O₂) and sulfate (SO₄^2–), which became important electron sinks that affected the NO₃^– and ClO₄^– removal rates. Using pyrosequencing, we elucidated how important phylotypes of each “primary” microbial group, i.e., denitrifying bacteria (DB), perchlorate-reducing bacteria (PRB), and sulfate-reducing bacteria (SRB), responded to changes in electron-acceptor loading. UniFrac, principal coordinate analysis (PCoA), and diversity analyses documented that the microbial community of biofilms sampled when the MBfRs had a high acceptor loading were phylogenetically distant from and less diverse than the microbial community of biofilm samples with lower acceptor loadings. Diminished acceptor loading led to SO₄^2– reduction in the lag MBfR, which allowed <i>Desulfovibrionales</i> (an SRB) and <i>Thiothrichales</i> (sulfur-oxidizers) to thrive through S cycling. As a result of this cooperative relationship, they competed effectively with DB/PRB phylotypes such as <i>Xanthomonadales</i> and <i>Rhodobacterales</i>. Thus, pyrosequencing illustrated that while DB, PRB, and SRB responded predictably to changes in acceptor loading, a decrease in total acceptor loading led to important shifts within the “primary” groups, the onset of other members (e.g., <i>Thiothrichales)</i>, and overall greater diversity

FINAL REPORT Perchlorate Destruction and Potable Water Production Using Membrane Biofilm Reduction and Membrane Filtration

Form Approve