Isolation And Screening Of Benzo[a]pyrene Aerobic Degrading Bacteria Acinetobacter XS-4 And Study On Its Degradation Performance And Mechanism

In this paper,BaP,a typical pollutant in PAHs,was used as the research object,the microorganisms with the ability to degrade BaP were screened from the actual coking wastewater,and their degradation performance was studied,and the degradation parameters were optimized by the response surface method.Through genome-wide sequencing and analysis of microorganisms,interpreting the degradation characteristics of microorganisms,looking for degradation genes related to BaP degradation,using GC-MS to detect their degradation products,and speculating on the degradation pathway.Finally,the immobilized microbial technology is used to immobilize microorganisms,and the removal effects of free bacteria and immobilized pellets on BaP are compared and analyzed in the environment of single BaP simulated wastewater and actual coking wastewater,in order to provide a certain theoretical basis for the establishment of BaP contaminated microbial remediation system and the application of bioremediation technology.The main conclusions are as follows:(1)Screening and identification of BaP aerobic degrading bacteriaUsing BaP as the only carbon source,the concentration was gradually increased,and seven strains of BaP-degrading bacteria(XS-1～XS-7)with the ability to degrade BaP were successfully screened,belonging to Paenarthrobacter,Pseudomonas,Acinetobacter and Microbacterium.The degradation capacity of 7 degrading bacteria was measured,among which the degradation capacity of strain XS-4 was the strongest,and the degradation rate of BaP within 7 days could reach 53.84%,so the strain XS-4was selected for follow-up research.Strain XS-4 belongs to the gram-negative bacterium,pale white,short rod-shaped,about 1 μm long and about 0.5 to 0.7 μm wide.Strain identification showed that strain XS-4 was a new species under the phylum Proteus,γ-Proteobacterium,Pseudomonas,Moraxella,and Acinetobacter.(2)Parameter optimization of BaP degradation of strain XS-4The Plackett-Burman experiment demonstrated that pH,substrate concentration,and temperature were significant influencing factors for BaP degradation by strain XS-4.The results of the Box-Behnken experiment showed that the interaction between pH and substrate concentration was the strongest,and the effect on BaP degradation efficiency was significant.The predictive analysis of the response surface model was carried out by Design Expert8.0,and the optimized degradation conditions were: pH 8,substrate concentration of 10 μg/mL,temperature of 25°C,and BaP degradation rate of 62.73%,which was 16.51% higher than before optimization.(3)Analysis of the degradation pathway of BaP by strain XS-4Strain XS-4 has a relatively rich gene for degrading aromatic hydrocarbons.KEGG annotation analysis showed that strain XS-4 was involved in multiple metabolic pathways,such as benzoate degradation(ko00362)and polycyclic aromatic hydrocarbon degradation(ko00624),and identified hydroxylase salicylate,protocateic acid 3,4-bioxygenase(β subunit),and protocateic acid 3,4-bioxygenase(α subunit).GC-MS analysis showed that derivatives such as benzoic acid,salicylic acid and phthalic acid were present in the degradation products.Combined with the results of genomic analysis,it was inferred that there was both a salicylic acid pathway and a phthalic acid pathway in the biological process of BaP degradation by strain XS-4.(4)A preliminary study on the application of fixed and degraded coking wastewater of BaP degraded strainsImmobilized pellets made under 10% polyvinyl alcohol(PVA)+ 0.5% sodium alginate(SA)have a uniform appearance texture and a loose and porous surface,providing a suitable growth environment for microbial adsorption and growth.The immobilizing agent of directly adding strain XS-4 to the wastewater has more advantages than free bacteria,and after 7 days,the BaP removal rates of the immobilized XS-4 pellets in the simulated wastewater and coking wastewater were62.36% and 71.55%,respectively.Immobilized pellets work together to remove BaP from coking wastewater by adsorption and microbial utilization.