Study On The Biodegradation Mechanisms And Removal Routes Of 17?-estradiol In Aquaculture Wastewater

In order to meet the increasing demand for aquatic products,Chinese aquaculture industry has developed rapidly in recent years.The amount of aquaculture wastewater produced is extremely large,and high-density and intensive aquaculture may cause the accumulation of estrogens,e.g.17?-estradiol(E2).E2 has the strongest endocrine disrupting activity among natural estrogens.It has the characteristics of persistence and refractory degradation.At the level of ng/L,it can cause endocrine disorders and feminization of aquatic products.Therefore,it poses a huge threat to the quality and safety of aquatic products.Ammonia nitrogen and other pollutants in aquaculture wastewater are usually removed by biodegradation.Biodegradation is also an efficient,environmentally friendly,and economical method for the removal of E2,but the research on its removal mechanism is still very limited.In this paper,the biodegradation of E2 in aquaculture wastewater is studied from two levels: the bacteria and the reactor.In order to clarify the biodegradation mechanism of E2 by different microbial populations,two common functional microorganisms,ammonia oxidizing bacteria(AOB)and Sphingomonas species,are selected.The influence of E2 on AOB is explored from the perspective of the amount of AOB and the activity of functional enzyme.Metabolomics is innovatively used to explore the differential intracellular metabolites of Sphingomonas sp.under exposure to E2.In order to explore the removal of E2 by biofilm reactors,the common nitrifying moving bed biofilm reactor(MBBR)is selected.The removal routes of E2 by MBBRs are thus clarified.This is the first attempt to correlate the removal routes with the biofilm characteristics to provide a new idea for the efficient removal of estrogens.The main research contents and results of this work are as follows:(1)AOB are nitrifying bacteria that play an important role in the nitrification reactor and also have the ability to degrade E2.Taking the pure ammonia oxidizing bacteria Nitrosomonas europaea as the research object,it is found that the E2 degradation by N.europaea followed zero-order reaction kinetics(r² = 0.944,4.07 ?g L^-1 h^-1).There was a significant linear negative correlation between the decrease of E2 and the accumulation of nitrite nitrogen(r =-0.924,P = 0.001).However,E2 was harmful to N.europaea.The ammonia nitrogen oxidation rate(AOR),bacterial density and ammonia monooxygenase(AMO)activity of N.europaea were reduced under exposure to 50 ng/L E2.The inhibitory impact on N.europaea was enhanced with increasing E2 dosage.(2)Compared with nitrifying bacteria,heterotrophic bacteria often have an absolute advantage in quantity.To study the E2 degradation by heterotrophs,pure species of Sphingomonas sp.(MCCC 1A06484)was selected as the research object.Batch experiments showed that the bacterium preferentially utilized glucose,sodium succinate and sodium acetate over E2.Interestingly,the presence of these preferred carbons increased the E2 removal efficiency by 20.1%.Furthermore,a positive relation between the utilization of total organic carbon(TOC)and E2 was found(P < 0.05).Using the method of metabolomics,it was shown that E2 exposure contributed to metabolism changes of lipid,nucleotide,carbohydrate,amino acid and membrane transport,which were considered to play roles in the E2 degradation.In addition,by comparing the differential intracellular metabolites of Sphingomonas sp.with and without E2,it was speculated that the up-regulated phosphatidylcholine might act as an indicator of bacterial degradation of E2.(3)AOB and heterotrophic bacteria together constitute the microbial community of a nitrification reactor,so this paper takes nitrifying MBBR as the representative to further explore the degradation characteristics of E2 at the reactor level.In marine aquaculture systems,the slow start-up of nitrifying MBBR under high salt stress was firstly targeted.It was found that the start-up was shorted by 16-18 days with increasing inlet salinity compared with the other strategies.Next,E2 was supplied into the MBBR influent.It was found that the ammonia removal remarkably decreased from 94.7% ± 2.1% to 85.6% ± 2.1%(P < 0.05)with E2 increasing from 10 ?g/L to 1 mg/L.Yet,this interference was negligible at the reactor level given the low level of E2 in practical field conditions(ng/L).Besides,E2 removal by MBBR was 84.5 ± 2.0% under the E2 dosage of 10 ?g/L,while it significantly increased to 98.7 ± 0.4% with inlet E2 increasing to 1 mg/L.This result was likely related to bacterial stress response.(4)In order to further clarify the removal routes of E2 in nitrifying MBBRs,three MBBRs with different C/N ratios(0 for C/N0;2 for C/N2;and 5 for C/N5)were operated in continuous mode.A 65-day degradation demonstrated that the MBBRs had high potential to remove E2 regardless of the C/N(E2 removal greater than 99% for all MBBRs,P > 0.05).Further batch tests showed that the E2 removal mainly resulted from heterotrophic activities for all MBBRs,accounting for approximately 85% for all MBBRs(P > 0.05),followed by nitrification(10-11%)and adsorption(4-5%).The correlation analysis between the E2 removal routes and the biofilm characteristics showed that,lower adhesive force likely led to higher E2 adsorption onto biofilms.Besides,the enhanced AOR was consistent with the high contribution of nitrification to the E2 attenuation.Importantly,the heterotrophic activity was positively correlated with its contribution to E2 removal(r = 0.99,P < 0.05).