The Characteristics Of PAHs Contamination In The WLFZ Of Three Gorges Reservoir And The Mechanisms Of Model Compoud Biodegradation Under Denitrification

The polycyclic aromatic hydrocarbons(PAHs)in the environment pose a serious threat to human health and the safety of the ecosystem due to the carcinogenicity,teratogenicity and mutagenicity as well as biological amplification and cumulative effect.PAHs are the priority organic pollutants and tend to concentrate in anoxic/anaerobic soil and sediment since the low water solubility in the Three Gorges Reservoir(TGR).Although the PAHs pollution has attracted widespread attention,the main biotransformation/degradation pathways are still not well understood in water-level-fluctuation-zone(WLFZ)of TGR.The anaerobic degradation of PAHs under denitrification has been studied for many years,but the reaction mechanism is not fully clear due to slow reaction and difficult microorganism enrichment,which hindered the development of bioremediation.Therefore,in this thesis,firstly the pollution characteristics and biodegradation pathway of 16 priority PAHs were surveyed based on GC-MS,microcosms experiment and molecular biology techniques in WLFZ of TGR.Further,phenanthrene and fluoranthene were used as the model PAHs due to the highest concentration in WLFZ,the mechanism of anaerobic degradation of phenanthrene and fluoranthene under denitrification synergistic anaerobic ammonium oxidation(Anammox),the mechanism of anaerobic degradation of phenanthrene and fluoranthene with nitric oxide(NO)as electron acceptor and the effect of ammonium on Anammox process and benzene degradation under denitrification were explored.These studies will provide important theoretical basis for the remediation of PAH contamination in the WLFZ of the TGR.The results were given as follows:The results were given as follows:(1)The PAHs were detected in all top soils from 16 samping sites.The concentration of 16 priority PAHs were 43.75-228.24 ng/g and 30.75-206.26 ng/g soil(dry weight)in high-water-level(HWL)and low-water-level(LWL)samples,respectively.The concentration of PAHs in urban was higher than that in rural section.The abundance of bam A gene,a bio-marker for anaerobic PAHs biodegradation,was significant positive correlated with the concentration of PAHs(R²=0.8)and significantly higher than that of the RHD gene that is a bio-marker for aerobic PAHs biodegradation.The co-occurrence network analysis based on 16S r RNA high-throughput sequencing suggested the Anaerolineaceae,Dechloromonas,Bacteroidetes?vadin HA17 and Geobacter which involved in PAHs biodegradation under nitrate and iron reduction were the keystone species in the WLFZ of the TGR.The diversity analysis of functional bacteria based on bam A gene further demonstrated that nitrate was the major electron acceptor for PAHs biodegradaiton.In addition,the microcosms experiment indicated that more model PAHs were degraded in anaerobic rather than aerobic condition.Hence,The anaerobic biodegradation with nitrate as electron acceptor was the main way of the PAHs contaminants removal in the WLFZ.These results would provide the basic guide for anaerobic bioremediation of PAHs in WLFZ of TGR.(2)To evaluate the effect of anaerobic ammonium oxidation(Anammox)on PAHs degradation under denitrification,the microcosms experiment under denitrification synergized with Anammox for phenanthrene and fluoranthene degradation were established by inoculating Anammox sludge.The average degradation rate of phenanthrene and fluoranthene were 11.77±0.31?mol/g VSS·d and 6.07±0.42?mol/g VSS·d,which about 1.82-and 1.95-fold those before addition of Anammox sludge,respectively.Meanwhile,the abundance of bam A were 2.26-and 16.78-foid those in phenanthrene and fluoranthene degradation under denitrification,respectively.The community structure analysis showed that Moheibacter,Hydrogenophaga,Thauera and Azoarcus were the key participators for the biodegradation of phenanthrene and fluoranthene under denitrification.In addition,Pseudomonas might also play vital role in the degradation of the phenanthrene.This research would contribute to enhacing the anaerobic bioremediation of PAHs under denitrification.(3)To explore the role and mechanism of the nitric oxide(NO)in biodegradation of phenanthrene and fluoranthene,the microcosms experiments were established with the addition of NO rather than nitrate.The long-term operation performance suggested that microorganism could degrade phenanthrene and fluoranthene with NO as electron acceptor in anaerobic conditions.After the two systems of NO-dependent phenanthrene and fluoranthene degradation operated stable,the first-order rate constants were 0.1940and 0.0825 day^-1,these close to that values(0.2290 and 0.1085,day^-1)with nitrate-reducing,respectively.The Q-PCR results suggested that the abundances of c Nor B,q Nor B and Nod gene were increased significantly in the biodegradation systems of phenanthrene and fluoranthene,respectively.Further,the abundances of RHD-GP gene were increased 314.77 and 93.32 times,while the abundances of bam A were decreased 102.33 and 102.27 times in the biogradation systems of phenanthrene and fluoranthene after adding NO as electron acceptor.Hence,the degradation of PAHs might be driven by the combine action of NO reduction and NO dismutation pathway under anaerobic condition.In this process,Pseudomonas and Ochrobactrum as well as Azoarcus,Moheibacter and Alicycliphilus may be the key participators for the degradation of phenanthrene and fluoranthene,respectively.The NO-dependent anaerobic degradation of PAHs can reduce the emission of NO and N₂O and has important ecological significance.(4)Ammonium is indispensable substance in Anammox process.To evaluate the effect of ammonium on enrichment of anaerobic ammonium oxidation bacteria(An AOB)and degradation of aromatic hydrocarbon,the microcosms experiment for benzene biodegradation were established with and without addition of ammonium,respectively.The results indicated that Anammox occured synergistically with organisms using the(dissimilatory nitrate reduction to ammonium)DNRA pathway,such as Draconibacterium and Ignavibacteriumn under the condition without addition of ammonium.Phylogenetic analysis showed 64%(16/25)and 36%(5/25)hzs B gene sequences,a specific biomarker of An AOB,belonged to Candidatus‘Brocadia fuldiga'and Candidatus‘Kuenenia',respectively.The addition of ammonium 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 An AOB belonged to Ca.‘Kuenenia'(84%)and Ca.‘Brocadia fuldiga'(16%).These results indicated that Ca.‘Brocadia fuldiga'could also play a role in DNRA.Hence,the DNRA and ammonium can promoted An AOB enrichment and accelerated the anaerobic degradation of benzene under denitrification.The study was helpful to further understanding the transformation mechanism of nitrogen and enhancing the remediation of aromatic hydrocarbon pollution.