Reductive metabolism of the important atmospheric gas isoprene by homoacetogens.

Isoprene is the most abundant biogenic volatile organic compound (BVOC) in the Earth's atmosphere and plays important roles in atmospheric chemistry. Despite this, little is known about microbiological processes serving as a terrestrial sink for isoprene. While aerobic isoprene degrading bacteria have been identified, there are no known anaerobic, isoprene-metabolizing organisms. In this study an H2-consuming homoacetogenic enrichment was shown to utilize 1.6 µmoles isoprene h-1 as an electron acceptor in addition to HCO3-. The isoprene-reducing community was dominated by Acetobacterium spp. and isoprene was shown to be stoichiometrically reduced to three methylbutene isomers (2-methyl-1-butene (>97%), 3-methyl-1-butene (≤2%), 2-methyl-2-butene (≤1%). In the presence of isoprene, 40% less acetate was formed suggesting that isoprene reduction is coupled to energy conservation in Acetobacterium spp. This study improves our understanding of linkages and feedbacks between biogeochemistry and terrestrial microbial activity.

Establishment and convergence of photosynthetic microbial biomats in shallow unit process open-water wetlands.

The widespread adoption of engineered wetlands designed for water treatment is hindered by uncertainties in system reliability, resilience and management associated with coupled biological and physical processes. To better understand how shallow unit process open-water wetlands self-colonize and evolve, we analyzed the composition of the microbial community in benthic biomats from system establishment through approximately 3 years of operation. Our analysis was conducted across three parallel demonstration-scale (7500 m2) cells located within the Prado Constructed Wetlands in Southern California. They received water from the Santa Ana River (5.9 ±â€¯0.2 mg/L NO3-N), a water body where the flow is dominated by municipal wastewater effluent from May to November. Phylogenetic inquiry and microscopy confirmed that diatoms and an associated aerobic bacterial community facilitated early colonization. After approximately nine months of operation, coinciding with late summer, an anaerobic community emerged with the capability for nitrate attenuation. Varying the hydraulic residence time (HRT) from 1 to 4 days the subsequent year resulted in modest ecological changes across the three parallel cells that were most evident in the outlet regions of the cells. The community that established at this time was comparatively stable for the remaining years of operation and converged with one that had previously formed approximately 550 km (350 miles) away in a pilot-scale (400 m2) wetland in Northern California. That system received denitrified (20.7 ±â€¯0.7 mg/L NO3-N), secondary treated municipal wastewater for 5 years of operation. Establishment of a core microbiome between the two systems revealed a strong overlap of both aerobic and anaerobic taxa with approximately 50% of the analyzed bacterial sequences shared between the two sites. Additionally the same species of diatom, Stauirsa construens var. venter, was prolific in both systems as the putative dominant primary producer. Our results indicate that despite differences in scale, geographic location and source waters, the shallow open-water wetland design can select for a rapid convergence of microbial structure and functionality associated with the self-colonizing benthic biomat. This resulting biomat matures over the first growing season with operational parameters such as HRT further exerting a modest selective bias on community succession.

Sulfide-Induced Dissimilatory Nitrate Reduction to Ammonium Supports Anaerobic Ammonium Oxidation (Anammox) in an Open-Water Unit Process Wetland.

Open-water unit process wetlands host a benthic diatomaceous and bacterial assemblage capable of nitrate removal from treated municipal wastewater with unexpected contributions from anammox processes. In exploring mechanistic drivers of anammox, 16S rRNA gene sequencing profiles of the biomat revealed significant microbial community shifts along the flow path and with depth. Notably, there was an increasing abundance of sulfate reducers (Desulfococcus and other Deltaproteobacteria) and anammox microorganisms (Brocadiaceae) with depth. Pore water profiles demonstrated that nitrate and sulfate concentrations exhibited a commensurate decrease with biomat depth accompanied by the accumulation of ammonium. Quantitative PCR targeting the anammox hydrazine synthase gene, hzsA, revealed a 3-fold increase in abundance with biomat depth as well as a 2-fold increase in the sulfate reductase gene, dsrA These microbial and geochemical trends were most pronounced in proximity to the influent region of the wetland where the biomat was thickest and influent nitrate concentrations were highest. While direct genetic queries for dissimilatory nitrate reduction to ammonium (DNRA) microorganisms proved unsuccessful, an increasing depth-dependent dominance of Gammaproteobacteria and diatoms that have previously been functionally linked to DNRA was observed. To further explore this potential, a series of microcosms containing field-derived biomat material confirmed the ability of the community to produce sulfide and reduce nitrate; however, significant ammonium production was observed only in the presence of hydrogen sulfide. Collectively, these results suggest that biogenic sulfide induces DNRA, which in turn can explain the requisite coproduction of ammonium and nitrite from nitrified effluent necessary to sustain the anammox community.IMPORTANCE This study aims to increase understanding of why and how anammox is occurring in an engineered wetland with limited exogenous contributions of ammonium and nitrite. In doing so, the study has implications for how geochemical parameters could potentially be leveraged to impact nutrient cycling and attenuation during the operation of treatment wetlands. The work also contributes to ongoing discussions about biogeochemical signatures surrounding anammox processes and enhances our understanding of the contributions of anammox processes in freshwater environments.

rRNA Gene Expression of Abundant and Rare Activated-Sludge Microorganisms and Growth Rate Induced Micropollutant Removal.

The role of abundant and rare taxa in modulating the performance of wastewater-treatment systems is a critical component of making better predictions for enhanced functions such as micropollutant biotransformation. In this study, we compared 16S rRNA genes (rDNA) and rRNA gene expression of taxa in an activated-sludge-treatment plant (sequencing batch membrane bioreactor) at two solids retention times (SRTs): 20 and 5 days. These two SRTs were used to influence the rates of micropollutant biotransformation and nutrient removal. Our results show that rare taxa (<1%) have disproportionally high ratios of rRNA to rDNA, an indication of higher protein synthesis, compared to abundant taxa (≥1%) and suggests that rare taxa likely play an unrecognized role in bioreactor performance. There were also significant differences in community-wide rRNA expression signatures at 20-day SRT: anaerobic-oxic-anoxic periods were the primary driver of rRNA similarity. These results indicate differential expression of rRNA at high SRTs, which may further explain why high SRTs promote higher rates of micropollutant biotransformation. An analysis of micropollutant-associated degradation genes via metagenomics and direct measurements of a suite of micropollutants and nutrients further corroborates the loss of enhanced functions at 5-day SRT operation. This work advances our knowledge of the underlying ecosystem properties and dynamics of abundant and rare organisms associated with enhanced functions in engineered systems.

Nitrate removal in shallow, open-water treatment wetlands.

The diffuse biomat formed on the bottom of shallow, open-water unit process wetland cells contains suboxic zones that provide conditions conducive to NO3(-) removal via microbial denitrification, as well as anaerobic ammonium oxidation (anammox). To assess these processes, nitrogen cycling was evaluated over a 3-year period in a pilot-scale wetland cell receiving nitrified municipal wastewater effluent. NO3(-) removal varied seasonally, with approximately two-thirds of the NO3(-) entering the cell removed on an annual basis. Microcosm studies indicated that NO3(-) removal was mainly attributable to denitrification within the diffuse biomat (i.e., 80 ± 20%), with accretion of assimilated nitrogen accounting for less than 3% of the NO3(-) removed. The importance of denitrification to NO3(-) removal was supported by the presence of denitrifying genes (nirS and nirK) within the biomat. While modest when compared to the presence of denitrifying genes, a higher abundance of the anammox-specific gene hydrazine synthase (hzs) at the biomat bottom than at the biomat surface, the simultaneous presence of NH4(+) and NO3(-) within the biomat, and NH4(+) removal coupled to NO2(-) and NO3(-) removal in microcosm studies, suggested that anammox may have been responsible for some NO3(-) removal, following reduction of NO3(-) to NO2(-) within the biomat. The annual temperature-corrected areal first-order NO3(-) removal rate (k20 = 59.4 ± 6.2 m yr(-1)) was higher than values reported for more than 75% of vegetated wetlands that treated water in which NO3(-) was the primary nitrogen species (e.g., nitrified secondary wastewater effluent and agricultural runoff). The inclusion of open-water cells, originally designed for the removal of trace organic contaminants and pathogens, in unit-process wetlands may enhance NO3(-) removal as compared to existing vegetated wetland systems.

Biotransformation of trace organic contaminants in open-water unit process treatment wetlands.

The bottoms of shallow, open-water wetland cells are rapidly colonized by a biomat consisting of an assemblage of photosynthetic and heterotrophic microorganisms. To assess the contribution of biotransformation in this biomat to the overall attenuation of trace organic contaminants, transformation rates of test compounds measured in microcosms were compared with attenuation rates measured in a pilot-scale system. The biomat in the pilot-scale system was composed of diatoms (Staurosira construens) and a bacterial community dominated by ß- and γ-Proteobacteria. Biotransformation was the dominant removal mechanism in the pilot-scale system for atenolol, metoprolol, and trimethoprim, while sulfamethoxazole and propranolol were attenuated mainly via photolysis. In microcosm experiments, biotransformation rates increased for metoprolol and propranolol when algal photosynthesis was supported by irradiation with visible light. Biotransformation rates increased for trimethoprim and sulfamethoxazole in the dark, when microbial respiration depleted dissolved oxygen concentrations within the biomat. During summer, atenolol, metoprolol, and propranolol were rapidly attenuated in the pilot-scale system (t1/2 < 0.5 d), trimethoprim and sulfamethoxazole were transformed more slowly (t1/2 ≈ 1.5-2 d), and carbamazepine was recalcitrant. The combination of biotransformation and photolysis resulted in overall transformation rates that were 10 to 100 times faster than those previously measured in vegetated wetlands, allowing for over 90% attenuation of all compounds studied except carbamazepine within an area similar to that typical of existing full-scale vegetated treatment wetlands.

Unit Process Wetlands for Removal of Trace Organic Contaminants and Pathogens from Municipal Wastewater Effluents.

Treatment wetlands have become an attractive option for the removal of nutrients from municipal wastewater effluents due to their low energy requirements and operational costs, as well as the ancillary benefits they provide, including creating aesthetically appealing spaces and wildlife habitats. Treatment wetlands also hold promise as a means of removing other wastewater-derived contaminants, such as trace organic contaminants and pathogens. However, concerns about variations in treatment efficacy of these pollutants, coupled with an incomplete mechanistic understanding of their removal in wetlands, hinder the widespread adoption of constructed wetlands for these two classes of contaminants. A better understanding is needed so that wetlands as a unit process can be designed for their removal, with individual wetland cells optimized for the removal of specific contaminants, and connected in series or integrated with other engineered or natural treatment processes. In this article, removal mechanisms of trace organic contaminants and pathogens are reviewed, including sorption and sedimentation, biotransformation and predation, photolysis and photoinactivation, and remaining knowledge gaps are identified. In addition, suggestions are provided for how these treatment mechanisms can be enhanced in commonly employed unit process wetland cells or how they might be harnessed in novel unit process cells. It is hoped that application of the unit process concept to a wider range of contaminants will lead to more widespread application of wetland treatment trains as components of urban water infrastructure in the United States and around the globe.