Isolation Of Aromatic Pollutants Degrading Bacteria And Investigation Of Degradation Mechanism

Aromatic compounds,containing benzene ring structure,are common in the natural environment.Many aromatic compounds are synthetic chemical raw materials and widely used in industrial manufacture,agricultural production and daily life.Some of aromatic compounds are widely used with a large amount.A part of these aromatic compounds were long-term residues in the environment,which has brought a huge threat to the ecological environment and human health.Microbial degradation is one of the main ways for the degradation of aromatic compounds in the environment,and it has attracted much attention because of its high efficiency,low cost and safety.In this study,a strain capable of degrading p-nitrophenol was isolated from sludge in a long-term polluted wastewater treatment plant of aromatic compounds,which was named as YC-RL1.The degradation rate of p-nitrophenol was 96.2% in 60 hours,.The strain YC-RL1 was identified as Arthrobacter sp.by colony and cell morphology observation,Biolog analysis and phylogenetic analysis of 16 S rDNA sequence.The effects of different environmental conditions on the degradation of p-nitrophenol were studied by single factor analysis.The results showed that strain YC-RL1 had good tolerance on the culture temperature(20-40),initial pH(6-9)and salinity(lower than 70g/L).Appropriate addition of glucose could significantly improve the degradation ability of the strain(≤ 7.5g / L),and the rotation speed and the inoculation amount of the strain had no significant effect on degradation.The optimum conditions for the degradation of the strain were optimized as pH 7.5,30 ℃ and the addition of 5.3 g/L glucose by response surface methodology(RSM).Substrate profile analysis showed that strain YC-RL1 could efficiently degrade p-xylene,biphenyl,bisphenol A,phenanthrene and naphthalene.The metabolic intermediates produced by the strain YC-RL1 during the degradation of p-nitrophenol were detected by high performance liquid chromatography tandem mass spectrometry.It was confirmed that the strain YC-RL1 could completely degrade p-nitrophenol via hydroxyquinol.The practical application potential of the strain was explored by simulating the soil remediation and the simulated wastewater treatment in the bioreactor.The results showed that the strain had good application potential.The structural characteristics of the strain showed that YC-RL1 had structural basis(terminal flagellum)for chemotaxis,and the chemotaxis towards p-nitrophenol was confirmed by plate chemotaxis and soil chemotaxis analysis.The whole genome sequence of the strain YC-RL1 was obtained by the third-generation sequencing technology(PacBio sequencing platform),and the whole genome sequence of strain YC-RL1 was submitted to GenBank(CP013297).Strain YC-RL1 contained a circular genome(3.8Mb,G + C content 64.26%)and two plasmids(plasmid01,112 Kb,G + C content 57.81%;plasmid02,59 Kb,G + C content 58.96%).The coding sequence(CDS)in the genome was predicted to yield 3,578 CDSs,of which 3,469 CDSs were annotated to public databases(COG,GO,KEGG,etc.).The genes involved in the degradation of aromatic compounds,especially the ring-cleavage dioxygenase genes,were analyzed by comparative genomics analysis.Five genes encoding ring-cleavage dioxygenase were obtained,which involved the degradation of catechol,biphenyl,protocatechuic acid and p-nitrophenol.The degrading gene cluster of biphenyl was further analyzed,and gene clusters involved the complete degradation of biphenyl were obtained.The bphC and bphD genes of biphenyl were cloned,expressed and verified,and the enzymological characteristics of BphD were analyzed.BphD showed high activity in wide ranges of pH and temperature,but most metal irons inhibited the enzyme activity.Further,the primary bioinformatics analysis of BphD was performed to a better understanding of amino acid sequences.