Phillip B. Gedalanga

Influence of physicochemical and operational parameters on Nitrobacter and Nitrospira communities in an aerobic activated sludge bioreactor.

To understand how to optimize performance of a partially nitrifying plant, the dynamics of Nitrospira and Nitrobacter abundance were studied over a 1 year period using quantitative polymerase chain reaction (qPCR) and their relative contributions to nitrite oxidation assessed including the affects of temperature and dissolved oxygen (DO). Correlation coefficients linking shifts in the community composition of nitrite-oxidizing bacteria (NOB) to operational or environmental variables indicated Nitrospira was significantly and negatively correlated to nitrite concentrations (r = -0.45, P < 0.01) and DO (r = -0.46, P < 0.01), while temperature showed a strong positive correlation (r = 0.59, P < 0.0001). However, the Nitrobacter portion of the total NOB populations showed a positive correlations with DO (r = 0.38, P < 0.01) and hydraulic retention time (HRT) (r = 0.33, P < 0.05), as well as being negatively correlated with temperature (r = -0.49, P < 0.001) suggesting specific niche adaptations within the NOB community. Nitrospira was dominant being better adapted to the low DO and shorter sludge retention times (SRT) of this plant, while Nitrobacter increased in abundance during the winter months, when temperatures were lower and DO concentrations higher. Principal component analysis (PCA) results supported these findings by the close proximity of Nitrospira and temperature biplots of PC1 and PC2 as well as grouping Nitrobacter, NO(2)(-)-N, HRT, and DO in the loadings together. The clustering of samples from specific dates also exhibited a strong seasonality.

Identification of Biomarker Genes To Predict Biodegradation of 1,4-Dioxane

ABSTRACT Bacterial multicomponent monooxygenase gene targets in Pseudonocardia dioxanivorans CB1190 were evaluated for their use as biomarkers to identify the potential for 1,4-dioxane biodegradation in pure cultures and environmental samples. Our studies using laboratory pure cultures and industrial activated sludge samples suggest that the presence of genes associated with dioxane monooxygenase, propane monooxygenase, alcohol dehydrogenase, and aldehyde dehydrogenase are promising indicators of 1,4-dioxane biotransformation; however, gene abundance was insufficient to predict actual biodegradation. A time course gene expression analysis of dioxane and propane monooxygenases in Pseudonocardia dioxanivorans CB1190 and mixed communities in wastewater samples revealed important associations with the rates of 1,4-dioxane removal. In addition, transcripts of alcohol dehydrogenase and aldehyde dehydrogenase genes were upregulated during biodegradation, although only the aldehyde dehydrogenase was significantly correlated with 1,4-dioxane concentrations. Expression of the propane monooxygenase demonstrated a time-dependent relationship with 1,4-dioxane biodegradation in P. dioxanivorans CB1190, with increased expression occurring after over 50% of the 1,4-dioxane had been removed. While the fraction of P. dioxanivorans CB1190-like bacteria among the total bacterial population significantly increased with decrease in 1,4-dioxane concentrations in wastewater treatment samples undergoing active biodegradation, the abundance and expression of monooxygenase-based biomarkers were better predictors of 1,4-dioxane degradation than taxonomic 16S rRNA genes. This study illustrates that specific bacterial monooxygenase and dehydrogenase gene targets together can serve as effective biomarkers for 1,4-dioxane biodegradation in the environment.

Biodegradation Kinetics of 1,4-Dioxane in Chlorinated Solvent Mixtures

This study investigated the impacts of individual chlorinated solvents and their mixtures on aerobic 1,4-dioxane biodegradation by Pseudonocardia dioxanivorans CB1190. The established association of these co-occurring compounds suggests important considerations for their respective biodegradation processes. Our kinetics and mechanistic studies demonstrated that individual solvents inhibited biodegradation of 1,4-dioxane in the following order: 1,1-dichloroethene (1,1-DCE) > cis-1,2-diochloroethene (cDCE) > trichloroethene (TCE) > 1,1,1-trichloroethane (TCA). The presence of 5 mg L(-1) 1,1-DCE completely inhibited 1,4-dioxane biodegradation. Subsequently, we determined that 1,1-DCE was the strongest inhibitor of 1,4-dioxane biodegradation by bacterial pure cultures exposed to chlorinated solvent mixtures as well as in environmental samples collected from a site contaminated with chlorinated solvents and 1,4-dioxane. Inhibition of 1,4-dioxane biodegradation rates by chlorinated solvents was attributed to delayed ATP production and down-regulation of both 1,4-dioxane monooxygenase (dxmB) and aldehyde dehydrogenase (aldH) genes. Moreover, increasing concentrations of 1,1-DCE and cis-1,2-DCE to 50 mg L(-1) respectively increased 5.0-fold and 3.5-fold the expression of the uspA gene encoding a universal stress protein. In situ natural attenuation or enhanced biodegradation of 1,4-dioxane is being considered for contaminated groundwater and industrial wastewater, so these results will have implications for selecting 1,4-dioxane bioremediation strategies at sites where chlorinated solvents are present as co-contaminants.

Interaction of Operational and Physicochemical Factors Leading to Gordonia amarae-Like Foaming in an Incompletely Nitrifying Activated Sludge Plant

ABSTRACT The overgrowth of Gordonia amarae-like bacteria in the mixed liquor of an incompletely nitrifying water reclamation plant was inversely correlated with temperature (r = −0.78; P < 0.005) and positively correlated with the solids retention time (SRT) obtained a week prior to sampling (r = 0.67; P < 0.005). Drops followed by spikes in the food-to-mass ratio (0.18 to 0.52) and biochemical oxygen demand concentrations in primary effluent (94 to 298 mg liter−1) occurred at the initiation of G. amarae-like bacterial growth. The total bacterial concentration did not increase as concentrations of G. amarae-like cells increased, but total bacterial cell concentrations fluctuated in a manner similar to that of G. amarae-like bacteria in the pseudo-steady state. The ammonium ion removal rate (percent) was inversely related to G. amarae-like cell concentrations during accelerated growth and washout phases. The dissolved oxygen concentration decreased as the G. amarae-like cell concentration decreased. The concentrations of G. amarae-like cells peaked (2.47 × 109 cells liter−1) approximately 1.5 months prior to foaming. Foaming occurred during the late pseudo-steady-state phase, when temperature declines reversed. These findings suggested that temperature changes triggered operational and physicochemical changes favorable to the growth of G. amarae-like bacteria. Fine-scale quantitative PCR (qPCR) monitoring at weekly intervals allowed a better understanding of the factors affecting this organism and indicated that frequent sampling was required to obtain statistical significance with factors changing as the concentrations of this organism increased. Furthermore, the early identification of G. amarae-like cells when they are confined to mixed liquor (107 cells liter−1) allows management strategies to prevent foaming.

Advances in bioremediation of 1,4-dioxane-contaminated waters

1,4-Dioxane is a contaminant of emerging concern that has been found widespread in groundwater, surface water, and drinking water environments. Many states are implementing lower regulatory advisory levels based on the toxicological profile of 1,4-dioxane and the potential public health risks. However, the unique chemical properties of 1,4-dioxane, such as high water solubility, low Henrys law constant, and importantly, the co-occurrence with chlorinated solvents and other contaminants, increase the challenges to efficiently cleanup 1,4-dioxane. This review summarizes currently available chemical and physical 1,4-dioxane treatment technologies and focuses on recent advances in bioremediation and monitoring tools. We also include laboratory studies and field applications to propose the next steps in 1,4-dioxane bioremediation research. It is important to provide useful references to change the industrial and regulatory perception of 1,4-dioxane biodegradability, to understand treatment mechanisms especially in contaminant mixtures, and to direct research for meeting practical needs.

Synergistic Treatment of Mixed 1,4-Dioxane and Chlorinated Solvent Contaminations by Coupling Electrochemical Oxidation with Aerobic Biodegradation

Biodegradation of the persistent groundwater contaminant 1,4-dioxane is often hindered by the absence of dissolved oxygen and the co-occurrence of inhibiting chlorinated solvents. Using flow-through electrolytic reactors equipped with Ti/IrO2-Ta2O5 mesh electrodes, we show that combining electrochemical oxidation with aerobic biodegradation produces an overadditive treatment effect for degrading 1,4-dioxane. In reactors bioaugmented by Pseudonocardia dioxanivorans CB1190 with 3.0 V applied, 1,4-dioxane was oxidized 2.5 times faster than in bioaugmented control reactors without an applied potential, and 12 times faster than by abiotic electrolysis only. Quantitative polymerase chain reaction analyses of CB1190 abundance, oxidation-reduction potential, and dissolved oxygen measurements indicated that microbial growth was promoted by anodic oxygen-generating reactions. At a higher potential of 8.0 V, however, the cell abundance near the anode was diminished, likely due to unfavorable pH and/or redox conditions. When coupled to electrolysis, biodegradation of 1,4-dioxane was sustained even in the presence of the common co-contaminant trichloroethene in the influent. Our findings demonstrate that combining electrolytic treatment with aerobic biodegradation may be a promising synergistic approach for the treatment of mixed contaminants.

Novel applications of molecular biological and microscopic tools in environmental engineering

Molecular biological methods offer flexible and powerful tools to environmental practitioners and researchers interested in studying environmental challenges in natural and engineered systems. In recent years, these techniques have allowed investigators to connect the fate, transport, and transformation of environmental chemical contaminants and pathogens with biological processes of functionally diverse microorganisms or microbial communities. Indeed, the boundaries of microbial ecosystems are constantly refined as researchers discover new links that extend beyond ————————— 1 Department of Civil and Environmental Engineering, University of California, Los Angeles, CA 90095. *Corresponding author: Phone: 310-794-9850, E-mail: mahendra@seas.ucla.edu 2 Department of Civil and Environmental Engineering, University of Utah, Salt Lake City, UT 84112. 3 Department of Civil, Architectural, and Environmental Engineering, Drexel University, Philadelphia, PA 19104 4 Department of Civil and Environmental Engineering, University of Massachusetts, Amherst, MA 01003. Bacteria to include Archaea and unicellular Eukarya. Quantitative polymerase chain reaction (qPCR) provides a rapid and sensitive approach to determine gene abundance and expression from a wide range of microorganisms from complex environments. Whole genome arrays (WGA) and functional gene arrays (FGA) are being used to elucidate transcriptional changes in response to environmental parameters. Antibiotic resistance profiling and microbial source tracking studies continue to benefit from the information provided by a molecular-based experimental design. Quantitative fluorescent in situ hybridization (qFISH) and next generation sequencing technologies are changing the way we view suspended solids in wastewater treatment. Innovative sensors are being developed that couple molecular biological, chemical, or physical properties to improve the sensitivity and specificity for intended targets. Thus, advanced molecular analysis complements conventional approaches to provide a more

Abiotic and bioaugmented granular activated carbon for the treatment of 1,4-dioxane-contaminated water

1,4-Dioxane is a probable human carcinogen and an emerging contaminant that has been detected in surface water and groundwater resources. Many conventional water treatment technologies are not effective for the removal of 1,4-dioxane due to its high water solubility and chemical stability. Biological degradation is a potentially low-cost, energy-efficient approach to treat 1,4-dioxane-contaminated waters. Two bacterial strains, Pseudonocardia dioxanivorans CB1190 (CB1190) and Mycobacterium austroafricanum JOB5 (JOB5), have been previously demonstrated to break down 1,4-dioxane through metabolic and co-metabolic pathways, respectively. However, both CB1190 and JOB5 have been primarily studied in laboratory planktonic cultures, while most environmental microbes grow in biofilms on surfaces. Another treatment technology, adsorption, has not historically been considered an effective means of removing 1,4-dioxane due to the contaminants low Koc and Kow values. We report that the granular activated carbon (GAC), Norit 1240, is an adsorbent with high affinity for 1,4-dioxane as well as physical dimensions conducive to attached bacterial growth. In abiotic batch reactor studies, 1,4-dioxane adsorption was reversible to a large extent. By bioaugmenting GAC with 1,4-dioxane-degrading microbes, the adsorption reversibility was minimized while achieving greater 1,4-dioxane removal when compared with abiotic GAC (95-98% reduction of initial 1,4-dioxane as compared to an 85-89% reduction of initial 1,4-dioxane, respectively). Bacterial attachment and viability was visualized using fluorescence microscopy and confirmed by amplification of taxonomic genes by quantitative polymerase chain reaction (qPCR) and an ATP assay. Filtered samples of industrial wastewater and contaminated groundwater were also tested in the bioaugmented GAC reactors. Both CB1190 and JOB5 demonstrated 1,4-dioxane removal greater than that of the abiotic adsorbent controls. This study suggests that bioaugmented adsorbents could be an effective technology for 1,4-dioxane removal from contaminated water resources.

Microbial responses to combined oxidation and catalysis treatment of 1,4-dioxane and co-contaminants in groundwater and soil

Post-treatment impacts of a novel combined hydrogen peroxide (H2O2) oxidation and WOx/ZrO2 catalysis used for the removal of 1,4-dioxane and chlorinated volatile organic compound (CVOC) contaminants were investigated in soil and groundwater microbial community. This treatment train removed ~90% 1,4-dioxane regardless of initial concentrations of 1,4-dioxane and CVOCs. The Illumina Miseq platform and bioinformatics were used to study the changes to microbial community structure. This approach determined that dynamic shifts of microbiomes were associated with conditions specific to treatments as well as 1,4-dioxane and CVOCs mixtures. The biodiversity was observed to decrease only after oxidation under conditions that included high levels of 1,4-dioxane and CVOCs, but increased when 1,4-dioxane was present without CVOCs. WOx/ZrO2 catalysis reduced biodiversity across all conditions. Taxonomic classification demonstrated oxidative tolerance for members of the genera Massilia and Rhodococcus, while catalyst tolerance was observed for members of the genera Sphingomonas and Devosia. Linear discriminant analysis effect size was a useful statistical tool to highlight representative microbes, while the multidimensional analysis elucidated the separation of microbiomes under the low 1,4-dioxane-only condition from all other conditions containing CVOCs, as well as the differences of microbial population among original, post-oxidation, and post-catalysis states. The results of this study enhance our understanding of microbial community responses to a promising chemical treatment train, and the metagenomic analysis will help practitioners predict the microbial community status during the post-treatment period, which may have consequences for long-term management strategies that include additional biodegradation treatment or natural attenuation.