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.

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.

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.