River water influenced by shale gas wastewater discharge for paddy irrigation has limited effects on soil properties and microbial communities.

The processes of hydraulic fracturing to extract shale gas generate a large amount of wastewater, and the potential impacts of wastewater discharge after treatment are concerning. In this field study, we investigated the effects of the irrigation of paddy fields for 2 consecutive years by river water that has been influenced by shale gas wastewater discharge on soil physicochemical properties, microbial community structure and function, and rice grain quality. The results showed that conductivity, chloride and sulfate ions in paddy soils downstream of the outfall showed an accumulative trend after two years of irrigation, but these changes occurred on a small scale (<500 m). Two-year irrigation did not cause the accumulation of trace metals (barium, cadmium, chromium, copper, lead, strontium, zinc, nickel, and uranium) in soil and rice grains. Among all soil parameters, the accumulation of chloride ions was the most pronounced, with concentrations in the paddy soil at the discharge site 13.3 times higher than at the upstream control site. The use of influenced river water for paddy irrigation positively increased the soil microbial diversity, but these changes occurred after two years of irrigation and did not occur after one year of irrigation. Overall, the use of river water affected by shale gas wastewater discharge for agricultural irrigation has limited effects on agroecosystems over a short period. Nevertheless, the possible negative effects of contaminant accumulation in soil and rice caused by longer-term irrigation should be seriously considered.

Combining eDNA and morphological approaches to reveal the impacts of long-term discharges of shale gas wastewaters on receiving waters.

The potential threats of shale gas wastewater discharges to receiving waters is of great concern. In this study, chemical analyses and biomonitoring were performed three times in a small river that received treated wastewater over a two-year period. The results of chemical analyses showed that the concentrations of chloride, conductivity, barium, and strontium increased at the discharge site, but their concentrations decreased considerably farther downstream (≥500 m). The concentrations of toxic organic compounds (16 US EPA priority polycyclic aromatic hydrocarbons and 6 priority phthalates), trace metals (strontium, arsenic, zinc, copper, chromium, lead, cadmium, nickel, and neodymium), and natural radionuclides (40K, 238U, 226Ra, and 232Th) were comparable to the corresponding background values or did not exhibit obvious accumulation in sediments with continued discharge. Morphological and environmental DNA approaches were used to reveal the potential effects of wastewater discharges on aquatic ecosystems. The results showed that the community structure of benthic invertebrates was not altered by the long-term discharges of shale gas wastewaters. However, the biodiversity indices (richness and Shannon) from the two approaches showed inconsistencies, which were caused by multiple reasons, and that substrates had a strong influence on the morphological biodiversity indices. A multimetric index was proposed to further analyze morphological and environmental DNA data, and the results showed no significant difference between the upstream and downstream sites. Generally, the chemical and biological results both demonstrated that the discharges of shale gas wastewaters had limited impacts on river ecosystems within two years.

Structure and functional capacity of a benzene-mineralizing, nitrate-reducing microbial community.

AIMS: How benzene is metabolized by microbes under anoxic conditions is not fully understood. Here, we studied the degradation pathways in a benzene-mineralizing, nitrate-reducing enrichment culture. METHODS AND RESULTS: Benzene mineralization was dependent on the presence of nitrate and correlated to the enrichment of a Peptococcaceae phylotype only distantly related to known anaerobic benzene degraders of this family. Its relative abundance decreased after benzene mineralization had terminated, while other abundant taxa-Ignavibacteriaceae, Rhodanobacteraceae and Brocadiaceae-slightly increased. Generally, the microbial community remained diverse despite the amendment of benzene as single organic carbon source, suggesting complex trophic interactions between different functional groups. A subunit of the putative anaerobic benzene carboxylase previously detected in Peptococcaceae was identified by metaproteomic analysis suggesting that benzene was activated by carboxylation. Detection of proteins involved in anaerobic ammonium oxidation (anammox) indicates that benzene mineralization was accompanied by anammox, facilitated by nitrite accumulation and the presence of ammonium in the growth medium. CONCLUSIONS: The results suggest that benzene was activated by carboxylation and further assimilated by a novel Peptococcaceae phylotype. SIGNIFICANCE AND IMPACT OF THE STUDY: The results confirm the hypothesis that Peptococcaceae are important anaerobic benzene degraders.

Risk assessment of pollutants in flowback and produced waters and sludge in impoundments.

Due to the growing hydraulic fracturing (HF) practices in China, the environmental risks of pollutants in flowback and produced waters (FPW) and sludge in impoundments for FPW reserves have drawn increasing attention. In this context, we first characterized the comparative geochemical characteristics of the FPW and the sludge in impoundments that collected FPW from 75 shale gas wells, and then the risks associated with the pollutants were assessed. The results demonstrated that four organic compounds detected in the FPW, naphthalene, acenaphthene, dibutyl phthalate, and bis(2-ethylhexyl)phthalate, were potential threats to surface waters. The concentrations of trace metals (copper, cadmium, manganese, chromium, nickel, zinc, arsenic, and lead) in the FPW and sludge were low; however, those of iron, barium, and strontium were high. The accumulation of chromium, nickel, zinc, and lead in the sludge became more evident as the depth increased. The environmental risks from heavy metals in the one-year precipitated sludge were comparable to those reported in the environment. However, the radium equivalent activities were 10-41 times higher than the recommended value for human health safety, indicating potential radiation risks. Although hydrophobic organic compounds, such as high-molecular-weight polycyclic aromatic hydrocarbons (PAHs), phthalate esters (PAEs), benzene, ethylbenzene, toluene, and xylene (BTEX), tended to accumulate in the sludge, their environmental risks were within tolerable ranges after proper treatment. Multiple antibiotic resistance genes (ARGs), such as those for macrolide, lincosamide, streptogramin (MLS), tetracycline, and multidrug resistances, were detected in the shale gas wastewaters and sludge. Therefore, the environmental risks of these emerging pollutants upon being discharged or leaked into surface waters require further attention.

Characterization of microbial communities and functions in shale gas wastewaters and sludge: Implications for pretreatment.

As hydraulic fracturing (HF) practices keep expanding in China, a comparative understanding of biological characteristics of flowback and produced waters (FPW) and sludge in impoundments for FPW reserve will help propose appropriate treatment strategies. Therefore, in this study, the microbial communities and functions in impoundments that collected wastewaters from dozens of wells were characterized. The results showed that microbial richness and diversity were significantly increased in sludge compared with those in FPW. The vast majority of microorganisms found in FPW and sludge are organic degraders, providing the possibility of using these indigenous microorganisms to biodegrade organic compounds. Our laboratory findings first show that wastewater pretreatment using these microorganisms was effective, and organic compounds in FPW from different shale formations were removed by 35-68% within 72 h in a wide temperature range (8 - 30 â„ƒ). Meanwhile, highly toxic compounds such as phthalate esters (PAEs), polycyclic aromatic hydrocarbons (PAHs), and petroleum hydrocarbons were effectively eliminated in reactors. The main microorganisms, key functional genes, and putative pathways for alkanes, PAHs, and PAEs degradation were also identified.

Bioanalytical equivalents and relative potencies for predicting the biological effects of mixtures.

Bioanalytical equivalents (BEQs) of mixtures and environmental samples are widely used to reflect the potential threat of pollutants in the environment and can be obtained by bioassays or using chemical analysis combined with relative potencies (REPs). In this study, the relationships between bioassay-detected BEQs (Bio-BEQs) and chemically analyzed BEQs (Chem-BEQs) were studied. BEQs and REPs are correlated with effect level and the concentration-response curves of the reference standard and sample. Thus, effect level (e.g., EC10, EC25 and EC50) should be addressed for the BEQ values obtained from bioassays or chemical analyses. The previous prerequisites for REPs application (i.e., curves that are parallel and have the same maximum response) are redundant, and the use of REPs for the calculation of BEQs or in risk assessment should instead be based on the same effect level. For a complex mixture with many components, all active components can be regarded as dilutions of a standard compound for inducing a specific effect. Relative toxicity estimates based on EC50 ignore the contribution of weak-active components with maximum response below EC50 of the reference standard, especially in complex mixtures or environmental samples. REPs based on an effect level EC10 that can be clearly discriminated from background response are recommended for BEQ calculation. As an example, the aryl hydrocarbon receptor (AhR)-mediated activity of US EPA priority polycyclic aromatic hydrocarbons (PAHs) in RTL-W1 cells was used to assess the reliability of REPs for mixture toxicity prediction based on the effect level EC10.

Nitric oxide-dependent biodegradation of phenanthrene and fluoranthene: The co-occurrence of anaerobic and intra-aerobic pathways.

Polycyclic aromatic hydrocarbons (PAHs) pollution as well as the emissions of nitric oxide (NO) and greenhouse gas nitrous oxide (N2O) in denitrification processes are currently two environmental issues of great concern. Although bioremediation of PAHs under denitrification is considered a promising approach, denitrification was an important contributor to N2O and NO emissions. This long-term study confirmed for the first time that microorganisms could utilize NO to efficiently degrade phenanthrene and fluoranthene. When the two systems of NO-dependent phenanthrene and fluoranthene degradation were stable, the first-order rate constants of phenanthrene and fluoranthene in the two systems (0.1940 and 0.0825 day-1, respectively) were close to those values (0.2290 and 0.1085 day-1, respectively) observed at nitrate-reducing conditions. Further analysis of functional genes revealed that phenanthrene and fluoranthene might be degraded under the combined action of the anaerobic pathway mediated by NO reduction and intra-aerobic pathway mediated by NO dismutation. The genomic analysis showed that Nod genes had high diversity and most of them were similar to aquifer cluster group in the two systems. Microbial community structure analysis indicated that Pseudomonas and Ochrobactrum might be key participants in NO-dependent phenanthrene degradation system, and Azoarcus, Alicycliphilus and Moheibacter might play vital roles in NO-dependent fluoranthene degradation system. This study provides new perspective for anaerobic remediation of PAH pollution and simultaneously reducing NO and N2O emissions during bioprocesses, which has important ecological significance for amending sediment and soil PAHs contamination and potential application for the removal of PAHs in flue gas.

The Co-occurrence of DNRA and Anammox during the anaerobic degradation of benzene under denitrification.

It was revealed that Anammox process promotes the anaerobic degradation of benzene under denitrification. This study investigates the effect of dissimilatory nitrate reduction to ammonium (DNRA) and exogenous ammonium on anaerobic ammonium oxidation bacteria (AnAOB) during the anaerobic degradation of benzene under denitrification. The results indicate that anammox occurs synergistically with organisms using the DNRA pathway, such as Draconibacterium and Ignavibacterium. Phylogenetic analysis showed 64% (16/25) and 36% (5/25) hzsB gene sequences, a specific biomarker of AnAOB, belonged to Candidatus 'Brocadia fuldiga' and Candidatus 'Kuenenia', respectively. Exogenous ammonium addition enhanced the anammox process and accelerated benzene degradation at a 1.89-fold higher average rate compared to that in the absence of exogenous ammonium and AnAOB belonged to Ca. 'Kuenenia' (84%) and Ca. 'Brocadia fuldiga' (16%). These results indicate that Ca. 'Brocadia fuldiga' could also play a role in DNRA. However, the diversity of abcA and bamA, the key anaerobic benzene metabolism biomarkers, remained unchanged. These findings suggest that anammox occurrence may be coupled with DNRA or exogenous ammonium and that anammox promotes anaerobic benzene degradation under denitrifying conditions. The results of this study contribute to understanding the co-occurrence of DNRA and Anammox and help explore their involvement in degradation of benzene, which will be crucial for directing remediation strategies of benzene-contaminated anoxic environment.

Inhibition of benzene, toluene, phenol and benzoate in single and combination on Anammox activity: implication to the denitrification-Anammox synergy.

A previous study demonstrated that denitrification synergized with Anammox could accelerate the anaerobic degradation of benzene. The inhibitory effects of benzene, toluene, phenol and benzoate in single and combination on Anammox activity were investigated by short-term batch tests. The results indicated that the inhibition of single compounds on Anammox could be well fitted with the extended non-competitive and Luong inhibition kinetic models. The inhibitions of the individual compound were in order as follows: benzene > toluene > phenol > benzoate. The joint inhibitions of bi-component mixtures of benzene with toluene, benzene with phenol and benzene with benzoate on Anammox activity were additive; the joint inhibition of a tri-component mixture (benzene, toluene and phenol) was partly additive; and the joint inhibition of a multicomponent mixture (benzene, toluene, phenol and benzoate) was synergistic. The effect of benzoate on the denitrification-Anammox synergy for benzene degradation was evaluated using a long-term test. Although the average rate of benzene degradation decreased by 13% with the addition of 10 mg L-1 benzoate, the average rate of NO3- and NH4+ increased by approximately 1- and 0.56-fold, respectively, suggesting that benzoate favors the stability of the denitrification-Anammox synergy. The carboxylation of benzene would be a more favorable pathway for the anaerobic degradation of benzene under denitrification synergized with Anammox.

Denitrification synergized with ANAMMOX for the anaerobic degradation of benzene: performance and microbial community structure.

To evaluate the effect of anaerobic ammonium oxidation (ANAMMOX) on benzene degradation under denitrification, a sequencing batch reactor (SBR) under denitrification synergized with ANAMMOX (SBR-DenAna) for benzene degradation was established by inoculating anaerobic ammonium-oxidizing bacteria (AnAOB) into a SBR under denitrification reactor (SBR-Den) for benzene degradation. The average rate of benzene degradation and the maximum first-order kinetic constant in SBR-DenAna were 2.34- and 1.41-fold those in SBR-Den, respectively, indicating that ANAMMOX improved the degradation of benzene under denitrification synergized with ANAMMOX. However, the average rate of benzene degradation decreased by 35% in the denitrification-ANAMMOX synergistic reactor when 10 mg N L-1 NO2- was added; the rate recovered once NO2- was depleted, indicating that ANAMMOX might detoxify NO2-. Results from high-throughput sequencing analysis revealed that Azoarcus within the family Rhodocyclaceae might be associated with benzene degradation in the two SBRs. AnAOB affiliated with the family Candidatus Brocadiaceae were just detected in SBR-DenAna.

Stoichiometry and kinetics of the anaerobic ammonium oxidation (Anammox) with trace hydrazine addition.

Purpose of this study is to investigate the stoichiometry and kinetics of anaerobic ammonium oxidation (Anammox) with trace hydrazine addition. The stoichiometry was established based on the electron balance of Anammox process with trace N2H4 addition. The stoichiometric coefficients were determined by the proton consumption and the changes in substrates and products. It was found that trace N2H4 addition can increase the yield of Anammox bacteria (AnAOB) and reduce NO3(-) yield, which enhances the Anammox. Subsequently, kinetic model of Anammox with trace N2H4 addition was developed, and the parameters of the anaerobic degradation model of N2H4 were obtained for the first time. The maximum specific substrate utilization rate, half-saturation constant and inhibition constant of N2H4 were 25.09mgN/g VSS/d, 10.42mgN/L and 1393.88mgN/L, respectively. These kinetic parameters might provide important information for the engineering applications of Anammox with trace N2H4 addition.

Groundwater-surface water interactions in the hyporheic zone under climate change scenarios.

Slight changes in climate, such as the rise of temperature or alterations of precipitation and evaporation, will dramatically influence nearly all freshwater and climate-related hydrological behavior on a global scale. The hyporheic zone (HZ), where groundwater (GW) and surface waters (SW) interact, is characterized by permeable sediments, low flow velocities, and gradients of physical, chemical, and biological characteristics along the exchange flows. Hyporheic metabolism, that is biogeochemical reactions within the HZ as well as various processes that exchange substances and energy with adjoining systems, is correlated with hyporheic organisms, habitats, and the organic matter (OM) supplied from GW and SW, which will inevitably be influenced by climate-related variations. The characteristics of the HZ in acting as a transition zone and in filtering and purifying exchanged water will be lost, resulting in a weakening of the self-purification capacity of natural water bodies. Thus, as human disturbances intensify in the future, GW and SW pollution will become a greater challenge for mankind than ever before. Biogeochemical processes in the HZ may favor the release of carbon dioxide (CO2), nitrous oxide (N2O), and methane (CH4) under climate change scenarios. Future water resource management should consider the integrity of aquatic systems as a whole, including the HZ, rather than independently focusing on SW and GW.