Junko Munakata-Marr

Effects of Heterogeneity and Experimental Scale on the Biodegradation of Diesel

Biodegradation of petroleum hydrocarbon contamination is a common method forremediating soils and groundwater. Due to complexities with field-scale studies,biodegradation rates are typically evaluated at the bench-scale in laboratory studies.However, important field conditions can be difficult to mimic in the laboratory. Thisstudy investigates three scaling factors that can impact laboratory biodegradation ratesand that are frequently unaccounted for in typical laboratory experimental procedures.These factors are soil heterogeneity, morphology of petroleum hydrocarbon non-aqueous phase liquids (NAPLs) and soil moisture distribution. The effects of these factors on the biodegradation rate of diesel NAPL is tested under a variety of experimental procedures from well-mixed batch studies to four-foot static soil columns. The results indicate that a high degree of variability results from even small-scale heterogeneities. In addition, it appears that as the experimental scale increases, the measured biodegradation rates slow. The results indicate that diesel biodegradation rates derived from small-scale experiments are not necessarily representative of field-scale biodegradation rates.

Disturbance and temporal partitioning of the activated sludge metacommunity

The resilience of microbial communities to press disturbances and whether ecosystem function is governed by microbial composition or by the environment have not been empirically tested. To address these issues, a whole-ecosystem manipulation was performed in a full-scale activated sludge wastewater treatment plant. The parameter solids retention time (SRT) was used to manipulate microbial composition, which started at 30 days, then decreased to 12 and 3 days, before operation was restored to starting conditions (30-day SRT). Activated sludge samples were collected throughout the 313-day time series in parallel with bioreactor performance (‘ecosystem function’). Bacterial small subunit (SSU) rRNA genes were surveyed from sludge samples resulting in a sequence library of >417 000 SSU rRNA genes. A shift in community composition was observed for 12- and 3-day SRTs. The composition was altered such that r-strategists were enriched in the system during the 3-day SRT, whereas K-strategists were only present at SRTs⩾12 days. This shift corresponded to loss of ecosystem functions (nitrification, denitrification and biological phosphorus removal) for SRTs⩽12 days. Upon return to a 30-day SRT, complete recovery of the bioreactor performance was observed after 54 days despite an incomplete recovery of bacterial diversity. In addition, a different, yet phylogenetically related, community with fewer of its original rare members displaced the pre-disturbance community. Our results support the hypothesis that microbial ecosystems harbor functionally redundant phylotypes with regard to general ecosystem functions (carbon oxidation, nitrification, denitrification and phosphorus accumulation). However, the impacts of decreased rare phylotype membership on ecosystem stability and micropollutant removal remain unknown.

Anaerobic ammonium oxidation by Nitrosomonas spp. and anammox bacteria in a sequencing batch reactor.

A sequencing batch reactor (SBR) was inoculated with mixed nitrifying bacteria from an anoxic tank at the conventional activated sludge wastewater treatment plant in Nongkhaem, Bangkok, Thailand. This enriched nitrifying culture was maintained under anaerobic conditions using ammonium (NH(4)(+)) as an electron donor and nitrite (NO(2)(-)) as an electron acceptor. Autotrophic ammonium oxidizing bacteria survived under these conditions. The enrichment period for anammox culture was over 100 days. Both ammonium and nitrite conversion rates were proportional to the biomass of ammonium oxidizing bacteria; rates were 0.08 g N/gV SS/d and 0.05 g N/g VSS/d for ammonium and nitrite, respectively, in a culture maintained for 3 months at 42 mg N/L ammonium. The nitrogen transformation rate at a ratio of NH(4)(+)-N to NO(2)(-)-N of 1:1.38 was faster, and effluent nitrogen levels were lower, than at ratios of 1:0.671, 1:2.18, and 1:3.05. Fluorescent in situ hybridization (FISH) was used to identify specific autotrophic ammonium oxidizing bacteria (Nitrosomonas spp., Candidatus Brocadia anammoxidans, and Candidatus Kuenenia stuttgartiensis). The ammonium oxidizing culture maintained at 42 mg N/L ammonium was enriched for Nitrosomonas spp. (30%) over Candidati B. anammoxidans and K. stuttgartiensis (2.1%) while the culture maintained at 210 mg N/L ammonium was dominated by Candidati B. anammoxidans and K. stuttgartiensis (85.6%). The specific nitrogen removal rate of anammox bacteria (0.6 g N/g anammox VSS/d) was significantly higher than that of ammonium oxidizing bacteria (0.4 g N/g Nitrosomonas VSS/d). Anammox bacteria removed up to 979 mg N/L/d of total nitrogen (ammonium:nitrite concentrations, 397:582 mg N/L). These results suggest significant promise of this approach for application to wastewater with high nitrogen but low carbon content, such as that found in Bangkok.

The effect of system variables on in situ sweep-efficiency improvements via viscosity modification

Laboratory experiments and numerical simulations were performed to critically evaluate the utility of viscosity modification as a technique to improve injected fluid sweep efficiencies within texturally heterogeneous geomedia. The objective of this technique is to improve the subsurface distribution of fluids by mitigating the potential for preferential flow and bypassing of lower permeability media that can limit the effectiveness of in situ remediation applications. The results of two-dimensional sand tank experiments and numerical simulations demonstrate that viscosity modification, via polymer amendment, can improve sweep efficiencies within layered heterogeneous structures by up to 90%, relative to the no-polymer case. The amount of sweep efficiency improvement depended on a number of system variables, including: the degree of layering, the relative positioning of layers within the system, the permeability contrast between layers, fluid viscosity, and the rheological character of the fluid utilized. Although significant sweep-efficiency improvement was observed, achieving 100% sweep in one pore volume was only possible when the permeability contrast was less than a factor of four, regardless of the viscosity and the rheological character of the fluid.

Identifying well contamination through the use of 3-D fluorescence spectroscopy to classify coalbed methane produced water.

Production of unconventional gas resources commonly requires the use of hydraulic fracturing and chemical production well additives. Concern exists for the use of chemical compounds in gas wells due to the risk of groundwater contamination. This study focuses on a proposed method of identifying groundwater contamination from gas production. The method focuses on the classification of naturally occurring organic signatures of coalbed methane (CBM) produced water compared to anthropogenic organic compounds. The 3-D fluorescence excitation-emission matrix (EEM) spectra of coalbed methane produced water samples revealed four peaks characteristic of coalbed methane produced water: Peak P (aromatic proteins region), Peak M(1) (microbial byproducts region), Peak M(2) (microbial byproducts region), and Peak H (humic acid-like region). Peak H is characteristic of the coal-water equilibria present in all basins, while peaks P and M(2) correlate with microbial activity in basins with biogenic methane generation pathways. Anthropogenic well additives produce EEM signatures with notable flooding of peaks P, M(1), M(2), and H, relatively higher overall fluorescence intensity, and slightly higher DOC concentrations. Fluorescence spectroscopy has the potential to be used in conjunction with groundwater contamination studies to determine if detected organic compounds originate from naturally occurring sources or well production additives.

Directed bacterial surface attachment via optical trapping

Abstract We describe a method to direct bacterial surface attachment using an optical trapping approach. By using a moving trap we are able to ‘catch’ individual cells, move them to a desired location, and attach them to a surface. We also illustrate an extension of the single optical trap by rapidly moving the single laser beam in a desired pattern and manipulating multiple bacteria simultaneously. This rapidly scanning laser optical trap creates a time-averaged light intensity profile capable of concurrently controlling several cells, thereby allowing study of more elaborate bacterial arrays. We demonstrate the utility of optical trapping in investigating the influence of cell spatial distribution and cell orientation on biofilm development.

Stable carbon isotope fractionation of trans-1,2-dichloroethylene during co-metabolic degradation by methanotrophic bacteria

Abstract Changes in the carbon isotope ratio ( δ 13 C) of trans -1,2-dichloroethylene ( t -DCE) were measured during its co-metabolic degradation by Methylomonas methanica , a type I methanotroph, and Methylosinus trichosporium OB3b, a type II methanotroph. In closed-vessel incubation experiments with each bacterium, the residual t -DCE became progressively enriched in 13 C, indicating isotopic fractionation. From these experiments, the biological fractionation during t -DCE co-metabolism, expressed as e , was measured to be −3.5‰ for the type I culture and −6.7‰ for the type II culture. This fractionation effect and subsequent enrichment in the δ 13 C of the residual t -DCE can thus be applied to determine the extent of biodegradation of DCE by these organisms. Based on these results, isotopic fractionation clearly warrants further study, as measured changes in the δ 13 C values of chlorinated solvents could ultimately be used to monitor the extent of biodegradation in laboratory or field settings where co-metabolism by methanotrophs occurs.

Disturbance opens recruitment sites for bacterial colonization in activated sludge

Little is known about the role of immigration in shaping bacterial communities or the factors that may dictate success or failure of colonization by bacteria from regional species pools. To address these knowledge gaps, the influence of bacterial colonization into an ecosystem (activated sludge bioreactor) was measured through a disturbance gradient (successive decreases in the parameter solids retention time) relative to stable operational conditions. Through a DNA sequencing approach, we show that the most abundant bacteria within the immigrant community have a greater probability of colonizing the receiving ecosystem, but mostly as low abundance community members. Only during the disturbance do some of these bacterial populations significantly increase in abundance beyond background levels and in few cases become dominant community members post-disturbance. Two mechanisms facilitate the enhanced enrichment of immigrant populations during disturbance: (i) the availability of resources left unconsumed by established species and (ii) the increased availability of niche space for colonizers to establish and displace resident populations. Thus, as a disturbance decreases local diversity, recruitment sites become available to promote colonization. This work advances our understanding of microbial resource management and diversity maintenance in complex ecosystems.

Near‐infrared spectroscopy of lacustrine sediments in the Great Salt Lake Desert: An analog study for Martian paleolake basins

The identification and characterization of aqueous minerals within ancient lacustrine environments on Mars are a high priority for determining the past habitability of the red planet. Terrestrial analog studies are useful both for understanding the mineralogy of lacustrine sediments, how the mineralogy varies with location in a lacustrine environment, and for validating the use of certain techniques such as visible–near-infrared (VNIR) spectroscopy. In this study, sediments from the Pilot Valley paleolake basin of the Great Salt Lake desert were characterized using VNIR as an analog for Martian paleolake basins. The spectra and subsequent interpretations were then compared to mineralogical characterization by ground truth methods, including X-ray diffraction, automated scanning electron microscopy, and several geochemical analysis techniques. In general, there is good agreement between VNIR and ground truth methods on the major classes of minerals present in the lake sediments and VNIR spectra can also easily discriminate between clay-dominated and salt-dominated lacustrine terrains within the paleolake basin. However, detection of more detailed mineralogy is difficult with VNIR spectra alone as some minerals can dominate the spectra even at very low abundances. At this site, the VNIR spectra are dominated by absorption bands that are most consistent with gypsum and smectites, though the ground truth methods reveal more diverse mineral assemblages that include a variety of sulfates, primary and secondary phyllosilicates, carbonates, and chlorides. This study provides insight into the limitations regarding the use of VNIR in characterizing complex mineral assemblages inherent in lacustrine settings.

Effects of oxytetracycline on anammox activity

Batch experiments were conducted to investigate the effects of oxytetracycline on anaerobic ammonium oxidation (anammox) process. The short-and long-term effects on anammox activity were studied by measuring ammonium (NH4 +), nitrite (NO2 −), and nitrate (NO3 −) concentrations over time. Experiments were conducted at NH4 +and NO2 − concentrations of 60–90 mg N/L and 60–190 mg N/L, respectively (NH4 +:NO2 − ratio from 1:1–1:2.25), oxytetracycline concentrations of 10–100 mg/L, and biomass concentrations of 300–800 mg/L. In the short-term study, anammox activity was inhibited by all oxytetracycline concentrations studied. However, daily addition of oxytetracycline to a concentration of 5 ± 3.5 mg/L in the anammox sequencing batch reactor completely inhibited anammox activity in the fifth week. Fluorescent in situ hybridization was used to identify autotrophic ammonium oxidizing bacteria (Nitrosomonas spp., Nitrobacter spp., Nitrospira spp., Candidatus Brocadia anammoxidans, and Candidatus Kuenenia stuttgartiensis). The population of anammox culture was significantly decreased while Nitrosomonas spp. and Nitrospira spp. increased in the fifth week compared with the first week of experiment. A not-competitive model fit the anammox inhibition data at oxytetracycline concentrations of 0–100 mg/L quite well with Vmax of 0.0435 mg N/mg VSS-hr and Ki of 54.66 mg/L.