Effects Of Zn-NPs,ZnO-NPs And Zn²⁺ On Nitrifying Bacteria And Microbial Communities And Their Microbiological Mechanisms

In this study,nitrifying bacteria enrichment culture was used as the research subject to study effects of short-term and long-term exposure of Zn-NPs,ZnO-NPs and Zn2+ with different concentrations on the physiology,biochemistry and activity of nitrifying bacteria,and analyze the microbial community structure and succession of nitrifying bacteria in the long-term exposure experiment.The affected distribution characteristics of AOB in nitrifying bacteria group were investigated.The main research contents and results are summarized as follows:(1)This study investigated the effects of short-term(12h)exposure of Zn-NPs,ZnO-NPs and Zn2+ on nitrifying bacteria.Zn2+ showed higher inhibitory effects on the growth of nitrifying bacteria than the two NPs under high zinc concentrations(>1 mg L-1).However,under low zinc concentration(0.1 mg L-1),SOUR was promoted by Zn2+ but inhibited by the two NPs.Nitrifying bacteria could produce ROS significantly under the two NPs with 0.1 mg L-1 and 1 mg L-1 zinc concentration,while Zn2+ could not induce oxidative damage to nitrifying bacteria until 10 mg L-1.Transcriptional analyses showed that total zinc concentration was strongly correlated with amoA gene expression but not with the expression of hao and nxrA genes.In addition,NOB showed a stronger stress response than AOB to the short-term exposure to Zn-NPs,ZnO-NPs and Zn2+.(2)This study investigated the effects of long-term gradient exposure of Zn-NPs,ZnO-NPs and Zn2+ on nitrifying bacteria.The results showed that the three materials posed no effect on the removal rate of ammonia nitrogen in nitrifying bacteria group at the concentrations of 0.1 mg L-1 and 1 mg L-1,even could promote the increase in biomass.Zn2+ first showed inhibition to nitrifying bacteria as the zinc concentration increased to 10 mg L-1,and 50 mg L-1 Zn2+ almost completely inhibited the transformation of nitrogen,and the recovery process was very slow.However,the inhibition of NPs to nitrifying bacteria temporarily appeared under the zinc concentration of 50 mg L-1.Overall,long-term exposure of the three materials has the less inhibition than short-term exposure.It was found that the SOUR inhibition rate and the expression of amoA gene in nitrifying bacteria were positively correlated with the dissolved zinc ion concentration in each reactor,while the expression of nxrA gene was less affected.Additionally,the toxicity of Zn-NPs and ZnO-NPs long-term stress to nitrifying bacteria is mainly due to the dissolution of zinc ions,rather than the nature of NPs.(3)This study investigated the microbiological mechanism of the effect of long-term gradient exposure to Zn-NPs,ZnO-NPs and Zn2+ on nitrifying bacteria.The results indicated that the microbial structure had no significant changes under the 0.1 mg L-1 and 1 mg L-1 zinc concentration of the three materials exposure.However,the distribution of microbial community were significantly different under high zinc concentrations(>10 mg L-1),which showed that the total abundance of AOB and NOB was declined,and AOB was more sensitive than NOB.It was also found that the AOB of Nitrosomonas genus decreased gradually with the increase of the stress concentration of the three materials,while the Nitrosospira genus appeared only in the nitrifying bacteria after 20 mg L-1 Zn2+ stress,and its abundance rised with the increase of the zinc concentration.Moreover,Nitrosospira was found to be the major genus of the NOB,whose abundance showed increasing trends after the exposure.RDA analysis showed that total zinc concentration,dissolved zinc ion concentration,effluent ammonia and nitrate nitrogen,MLSS and EPS all significantly affected the microbial distribution,and the difference of stress materials was the key factor leading to the change of nitrifying bacteria.It was found in AOB polymorphism analysis that the stress of the three materials changed the distribution of AOB in the nitrifying bacteria grotp conpared to the control.Nitrosomonas.nitrosa was the dominant AOB in control,while Nitrosospira dominated under Zn2+ stress,and Nitrosomonas oligotropha became the prevailing species.