Effects of ZnO nanoparticles on flocculation and sedimentation of activated sludge in wastewater treatment process.

Despite the behaviors of ZnO nanoparticles (ZnO NPs) in wastewater treatment processes have been widely explored, the impacts of ZnO NPs on the activated sludge's flocculation and sedimentation performances for solid-liquid separation have rarely been involved yet. In this study, ZnO NPs were observed to exert a dose-dependent negative effect on the sludge's flocculation performance but did not significantly impact the sludge' sedimentation behaviors. Furthermore, it was NPs themselves rather than the dissolved Zn2+ who impaired on the sludge flocculation performance because the Zn2+ alone would not compromise the sludge's flocculation efficiency. In addition, the sludge flocculation performance was revealed to be inversely related to the extracellular polymeric substances (EPS) content in the sludge and the direct contacts between ZnO NPs and the cells in the sludge should be the prerequisite to stimulate the secretion of the sludge EPS. The poor sludge flocculation performance could also be caused by the reduced protein/polysaccharide (PN/PS) ratio and the zeta (ζ) potential in the loosely bound (LB-EPS) after the sludge exposure to ZnO NPs. Fourier transform-infrared spectra (FT-IR) and three dimensional - excitation emission fluorescence spectra (3D-EEM) analysis further revealed that the decrease of the tyrosine PN-like substance level in the LB-EPS was probably the key reason for the decreased PN/PS ratio and ζ potential in the LB-EPS, which eventually induced the decline of the sludge flocculation performance under the ZnO NP stress. These results could potentially expand the knowledge on sludge flocculation and sedimentation in the presence of ZnO NPs.

Regulation of membrane fixation and energy production/conversion for adaptation and recovery of ZnO nanoparticle impacted Nitrosomonas europaea.

The ZnO nanoparticle (NP) effects on typical ammonia-oxidizing bacteria, Nitrosomonas europaea in a chemostat bioreactor, and the cells' toxicity adaptation and recovery potentials were explored. Hardly any inhibition was observed when the NP concentration was high up to 10 mg/L. The cells exposed to 50 mg/L ZnO NPs displayed time-dependent impairment and recovery potentials in terms of cell density, membrane integrity, nitrite production rate, and ammonia monooxygenase activity. The 6-h NP stress impaired cells were nearly completely restored during a 12-h recovery incubation, while the longer exposure time would cause irretrievable cell damage. Microarray analysis further indicated the transcriptional adaptation of N. europaea to NP stress. The regulations of genes encoding for membrane permeability or osmoprotectant, membrane integrity preservation, and inorganic ion transport during NP exposure and cell recovery revealed the importance of membrane fixation and the associated metabolisms for cells' self-protection and the following recovery from NP stress. The oxidative phosphorylation, carbon assimilation, and tricarboxylic acid (TCA) cycling pathways involved in the cells' antitoxicity activities and would promote the energy production/conversion efficiency for cell recovery. The heavy metal resistance, histidine metabolism, toxin-antitoxin defense, glycolysis, and sulfate reduction pathways were also suggested to participate in the cell detoxication and recovery processes. All these findings provided valuable insights into the mechanisms of cell-mediated ZnO NP cytotoxicity and their potential impacts on wastewater nitrogen removal system.

Chronic effects of cerium dioxide nanoparticles on biological nitrogen removal and nitrous oxide emission: Insight into impact mechanism and performance recovery potential.

The influence of cerium dioxide nanoparticles (CeO2 NPs) on biological nitrogen removal and associated nitrous oxide (N2O) emission has seldom been addressed yet. Herein, the chronic effect of CeO2 NPs on the nitrogen transformation processes during wastewater treatment and the impacted system's self-recovery potential after CeO2 NP stress removal were investigated. CeO2 NP of 10-50 mg/L induced significant declines of the ammonia nitrogen (NH4+-N) and the total nitrogen removal efficiencies, but triggered the nitrite accumulation and the N2O emission. The N2O reductase (NOS) activity was negatively correlated with the N2O emission level, and the inhibition of NOS activity under CeO2 NP stress was probably due to the depressions of the sludge denitrifiers' metabolic activities. The NH4+-N removal efficiency was successfully regained after the recovery period although the N2O emission level was still higher than the pre-exposure period, which was probably due to the residual CeO2 NPs inside the activated sludge.

Enhanced Nutrient Removal in A2N Effluent by Reclaimed Biochar Adsorption.

The excessive nitrogen and phosphorus discharged into the water environment will cause water eutrophication and thus disrupt the water ecosystem and even exert biological toxicities. In this study, the absorption removal of nitrogen and phosphorus from the anaerobic tank in an anaerobic−anoxic/nitrifying system using four different kinds of biowaste-reclaimed biochars were investigated and compared. The effects of temperature and pH on nutrient adsorption removal were further investigated. The four kinds of biochar were successfully prepared and well characterized using a scanning electron microscope, fourier transform infrared spectroscopy, X-ray diffraction and Brunner−Emmet−Teller methods. Generally, there was no significant change in chemical oxygen demand (COD) and NH4+-N removal efficiencies when treated by the different biochars, while the activated sludge biochar (ASB) displayed the highest total phosphorus (TP) removal efficiency. The initial TP concentrations (<40 mg/L) displayed no remarkable effects on the TP adsorption removal, while the increase of temperature generally enhanced TP and NH4+-N adsorptions on the ASB. Besides, the increase of pH significantly promoted NH4+-N removal but depressed TP removal. Moreover, the adsorption process of TP by the ASB complies with the secondary kinetic model, suggesting the chemical precipitation and physical electrostatic interaction mechanisms of TP adsorption removal. However, the adsorption of NH4+-N conformed to the inner-particle diffusion model, indicating that the NH4+-N adsorption was mainly involved with pore diffusions in the particles.

Long-term exposure to nano-TiO2 interferes with microbial metabolism and electron behavior to influence wastewater nitrogen removal and associated N2O emission.

The extensive use of nano-TiO2 has caused concerns regarding their potential environmental risks. However, the stress responses and self-recovery potential of nitrogen removal and greenhouse gas N2O emissions after long-term nano-TiO2 exposure have seldom been addressed yet. This study explored the long-term effects of nano-TiO2 on biological nitrogen transformations in a sequencing batch reactor at four levels (1, 10, 25, and 50 mg/L), and the reactor's self-recovery potential was assessed. The results showed that nano-TiO2 exhibited a dose-dependent inhibitory effect on the removal efficiencies of ammonia nitrogen and total nitrogen, whereas N2O emissions unexpectedly increased. The promoted N2O emissions were probably due to the inhibition of denitrification processes, including the reduction of the denitrifying-related N2O reductase activity and the abundance of the denitrifying bacteria Flavobacterium. The inhibition of carbon source metabolism, the inefficient electron transfer efficiency, and the electronic competition between the denitrifying enzymes would be in charge of the deterioration of denitrification performance. After the withdrawal of nano-TiO2 from the influent, the nitrogen transformation efficiencies and the N2O emissions of activated sludge recovered entirely within 30 days, possibly attributed to the insensitive bacteria survival and the microbial community diversity. Overall, this study will promote the current understanding of the stress responses and the self-recovery potential of BNR systems to nanoparticle exposure.

Insights into chronic zinc oxide nanoparticle stress responses of biological nitrogen removal system with nitrous oxide emission and its recovery potential.

The nitrogen transformation performances and greenhouse gas nitrous oxide (N2O) emissions in a sequencing batch reactor under chronic exposure to zinc oxide nanoparticles (ZnO NPs) were quantified and the system's self-recovery potentials were assessed. ZnO NPs posed a dose-dependent depression effect on the removal efficiencies of ammonia nitrogen (NH4+-N) and total nitrogen (TN), and the N2O emissions. The suppressed N2O emissions had a positive relationship with the activity ratios of nitrite/NO reductases and N2O reductase, and were expected to be caused by the inhibited heterotrophic denitrification process. The inhibition of glucose metabolism key enzymes and electron transport chain activities would be responsible for the heterotrophic denitrification performances deterioration. Furthermore, the removal efficiencies of NH4+-N and TN were recovered to control levels through the nitrite-shunt. However, the N2O emission increased significantly above the control during the recovery period mainly due to the irreversibility of the depressed nitrite oxidation activities.

Responses of nitrogen transformation processes and N2O emissions in biological nitrogen removal system to short-term ZnO nanoparticle stress.

Although the adverse effects of ZnO nanoparticles (ZnO NPs) on biological nitrogen removal (BNR) processes have widely been reported, the impacts of ZnO NPs on the whole nitrogen transformation processes, especially the production of nitrous oxide (N2O), a typical greenhouse gas in a BNR system have rarely been systematically studied yet. In this study, the performances of both the nitrification and denitrification processes were investigated and the N2O emission was simultaneously monitored in a sequencing batch reactor (SBR) when exposed to 1, 25 or 50 mg/L ZnO NPs for one cycle. The dose-dependent ZnO NP depression effects were generally observed on denitrification processes, total nitrogen (TN) removal efficiency and N2O emissions but not nitrification process. Meanwhile, the N2O emission was positively correlated with NO2--N accumulation in the oxic stage. Further investigation showed that the expressions of nitrate (NO3-) reduction associated narG gene were down-regulated with the increase of NP stress, and the transcript ratios of NO2-/NO reduction gene to N2O reduction one (nirK/nosZ and norB/nosZ) decreased. The released Zn2+ from ZnO NPs took an important role in the inhibition of denitrification processes. ZnO NPs addition also induced the dose-dependent variations in the superoxide dismutase (SOD) and catalase (CAT) activities, which probably contributed to the suppression of the excess reactive oxygen species (ROS) generations to mitigate nanotoxicity. The excessive secretion of protein (PN) in tightly bound EPS (TB-EPS) when ZnO NPs were no <25 mg/L further supported the system's potential self-regulation mechanism for nanotoxicity resistance. CAPSULE: The effects of ZnO NPs on the whole nitrogen transformation processes in a biological nitrogen removal sequencing batch reactor, including the N2O emissions were investigated. The system's potential self-regulation mechanism for nanotoxicity resistance was addressed.

Effects of Exogenous N-Acyl-Homoserine Lactone as Signal Molecule on Nitrosomonas Europaea under ZnO Nanoparticle Stress.

Despite the adverse effects of emerging ZnO nanoparticles (nano-ZnO) on wastewater biological nitrogen removal (BNR) systems being widely documented, strategies for mitigating nanoparticle (NP) toxicity impacts on nitrogen removal have not been adequately addressed. Herein, N-acyl-homoserine lactone (AHL)-based quorum sensing (QS) was investigated for its effects against nano-ZnO toxicity to a model nitrifier, Nitrosomonas europaea. The results indicated that AHL-attenuated nano-ZnO toxicity, which was inversely correlated with the increasing dosage of AHL from 0.01 to 1 µM. At 0.01 µM, AHL notably enhanced the tolerance of N. europaea cells to nano-ZnO stress, and the inhibited cell proliferation, membrane integrity, ammonia oxidation rate, ammonia monooxygenase activity and amoA gene expression significantly increased by 18.2 ± 2.1, 2.4 ± 0.9, 58.7 ± 7.1, 32.3 ± 1.7, and 7.3 ± 5.9%, respectively, after 6 h of incubation. However, increasing the AHL dosage compromised the QS-mediated effects and even aggravated the NPs' toxicity effects. Moreover, AHLs, at all tested concentrations, significantly increased superoxide dismutase activity, indicating the potential of QS regulations to enhance cellular anti-oxidative stress capacities when facing NP invasion. These results provide novel insights into the development of QS regulation strategies to reduce the impact of nanotoxicity on BNR systems.

Adaption and recovery of Nitrosomonas europaea to chronic TiO2 nanoparticle exposure.

Although the adverse impacts of emerging nanoparticles (NPs) on the biological nitrogen removal (BNR) process have been broadly reported, the adaptive responses of NP-impaired nitrifiers and the related mechanisms have seldom been addressed to date. Here, we systematically explored the adaption and recovery capacities of the ammonia oxidizer Nitrosomonas europaea under chronic TiO2 NP exposure and different dissolved oxygen (DO) conditions at the physiological and transcriptional levels in a chemostat reactor. N. europaea cells adapted to 50 mg/L TiO2 NP exposure after 40-d incubation and the inhibited cell growth, membrane integrity, nitritation rate, and ammonia monooxygenase activity all recovered regardless of the DO concentrations. Transmission electron microscope imaging indicated the remission of the membrane distortion after the cells' 40-d adaption to the NP exposure. The microarray results further suggested that the metabolic processes associated with the membrane repair were pivotal for cellular adaption/recovery, such as the membrane efflux for toxicant exclusion, the structural preservation or stabilization, and the osmotic equilibrium adjustment. In addition, diverse metabolic and stress-defense pathways, including aminoacyl-tRNA biosynthesis, respiratory chain, ATP production, toxin-antitoxin 'stress-fighting', and DNA repair were activated for the cellular adaption coupled with the metabolic activity recovery, probably via recovering the energy production/conversion efficiency and mediating the non-photooxidative stress. Finally, low DO (0.5 mg/L) incubated cells were more susceptible to TiO2 NP exposure and required more time to adapt to and recover from the stress, which was probably due to the stimulation limitation of the oxygen-dependent energy metabolism with a lower oxygen supply. The findings of this study provide new insights into NP contamination control and management adjustments during the BNR process.

Responses and recovery assessment of continuously cultured Nitrosomonas europaea under chronic ZnO nanoparticle stress: Effects of dissolved oxygen.

Although the antibacterial performances of emerging nanoparticles (NPs) have been extensively explored in the nitrifying systems, the impacts of dissolved oxygen (DO) levels on their bio-toxicities to the nitrifiers and the impaired cells' recovery potentials have seldom been addressed yet. In this study, the physiological and transcriptional responses of the typical ammonia oxidizers - Nitrosomonas europaea in a chemostat to the chronic ZnO NP exposure under different DO conditions were investigated. The results indicated that the cells in steady-growth state in the chemostat were more persevering than batch cultured ones to resist ZnO NP stress despite the dose-dependent NP inhibitory effects were observed. In addition, the occurred striking over-expressions of amoA and hao genes at the initial NP exposure stage suggested the cells' self-regulation potentials at the transcriptional level. The low DO (0.5 mg/L) cultured cells displayed higher sensitivity to NP stress than the high DO (2.0 mg/L) cultured ones, probably owning to the inefficient oxygen-dependent electron transfer from ammonia oxidation for energy conversion/production. The following 12-h NP-free batch recovery assays revealed that both high and low DO cultured cells possessed the physiological and metabolic activity recovery potentials, which were in negative correlation with the NP exposure time. The duration of NP stress and the resulting NP dissolution were critical for the cells' damage levels and their performance recoverability. The membrane preservation processes and the associated metabolism regulations were expected to actively participate in the cells' self-adaption to NP stress and thus be responsible for their metabolic activities recovery.

Physiological and transcriptional responses of Nitrosomonas europaea to TiO2 and ZnO nanoparticles and their mixtures.

The short-term combined effects of two most extensively used nanoparticles (NPs) TiO2 NPs (n-TiO2) and ZnO NPs (n-ZnO) versus their individual cytotoxicities on a model ammonia-oxidizing bacterium, Nitrosomonas europaea, were investigated at both physiological and transcriptional levels. n-ZnO exerted more serious impairment effects on cell morphology, cell density, membrane integrity, and ammonia monooxygenase activity than n-TiO2. However, the co-existing n-TiO2 displayed a dose-dependent mitigation effect on n-ZnO cytotoxicity. Consistently, the n-TiO2 and n-ZnO mixture-impacted global transcriptional expression profile, obtained with the whole-genome microarray technique, was more comparable to the n-TiO2-impacted one than that impacted by n-ZnO. The expressions of numerous genes associated with heavy metal scavenging, DNA repair, and oxidative stress response were less up-regulated under the binary impacts of NP mixture than n-ZnO. Moreover, only n-ZnO alone stimulated the up-regulations of heavy metal resistance genes, which further implied the capacity of co-existing n-TiO2 to alleviate n-ZnO cytotoxicity. In addition, the damage of cell membrane structures and the suppression of cell membrane biogenesis-related gene expressions under the influence of either individual NPs or their combinations strongly suggested that the interruption of cell membranes and the associated metabolic activities would probably be one of NPs' critical cytotoxicity mechanisms.

Toxicity of binary mixtures of metal oxide nanoparticles to Nitrosomonas europaea.

Although the widely used metal oxide nanoparticles (NPs) titanium dioxide NPs (n-TiO2), cerium dioxide NPs (n-CeO2), and zinc oxide NPs (n-ZnO) have been well known for their potential cytotoxicities to environmental organisms, their combined effects have seldom been investigated. In this study, the short-term binary effect of n-CeO2 and n-TiO2 or n-ZnO on a model ammonia oxidizing bacterium, Nitrosomonas europaea were evaluated based on the examinations of cells' physiological, metabolic, and transcriptional responses. The addition of n-TiO2 mitigated the negative effect of more toxic n-CeO2 and the binary toxicity (antagonistic toxicity) of n-TiO2 and n-CeO2 was generally lower than the single NPs induced one. While the n-CeO2/n-ZnO mixture exerted higher cytotoxicity (synergistic cytotoxicity) than that from single NPs. The increased addition of the less toxic n-CeO2 exaggerated the binary toxicity of n-CeO2/n-ZnO mixture although the solubility of n-ZnO was not significantly affected, which excluded the contribution of the dissolved Zn ions to the enhancement of the combined cytotoxicity. The cell membrane disturbances and NP internalizations were detected for all the NP impacted cultures and the electrostatic interactions among the two distinct NPs and the cells were expected to play a key role in mediating their direct contacts and the eventual binary nanotoxicity to the cells.