Biodegradation mechanisms and microbial functional diversity during coupled pnitrophenol(PNP)and p-aminophenol(PAP)degradation were studied in a bioelectrochemical system.PNP in the biocathode and PAP in the bioanode ...Biodegradation mechanisms and microbial functional diversity during coupled pnitrophenol(PNP)and p-aminophenol(PAP)degradation were studied in a bioelectrochemical system.PNP in the biocathode and PAP in the bioanode were almost completely removed within 28 hr and 68 hr respectively.The degradation followed the steps including hydrating hydroxyalkylation,dehydrogenating carbonylation,and hydrolating ring cleavage,etc.Metagemomic analysis based on the KEGG and egg NOG database annotations revealed the microbial composition and functional genes/enzymes related to phenol degradation in the system.The predominant bacteria genera were Lautropia,Pandoraea,Thiobacillus,Ignavibacterium,Truepera and Hyphomicrobium.The recognized biodegradation genes/enzymes related to pollutant degradation were as follows:pmo,hbd,&ppo for phenol degradation,nzba,amie,&badh for aromatic degradation,and CYP&p450 for xenobiotics degradation,etc.The co-occurrence of ARGs(antibiotic resistant genes),such as ade F,Mex J,Erm F,PDC-93 and Escherichiacolimdf A,etc.,were annotated in CARD database during the biodegradation process.The Proteobacteria&Actinobacteria phylum was the primary host of both the biodegradation genes&ARGs in this system.The microbial functional diversity ensured the effective biodegradation of the phenol pollutants in the bioelectrochemical system.展开更多
A recombinant strain, Escherichia coli JM109-AN1, was obtained by constructing of a genomic library of the total DNA of Delftia sp. AN3 in E. coli JM109 and screening for catechol 2,3-dioxygenase activity. This recomb...A recombinant strain, Escherichia coli JM109-AN1, was obtained by constructing of a genomic library of the total DNA of Delftia sp. AN3 in E. coli JM109 and screening for catechol 2,3-dioxygenase activity. This recombinant strain could grow on aniline as sole carbon, nitrogen and energy source. Enzymatic assays revealed that the exogenous genes including aniline dioxygenase （AD） and catechol 2,3-dioxygenase （C230） genes could well express in the recombinant strain with the activities of AD and C230 up to 0.31 U/mg wet cell and 1.92 U/mg crude proteins, respectively. The AD or C23O of strain AN3 could only catalyze aniline or catechol but not any other substituted substrates. This recombinant strain contained a recombinant plasmid, pKC505-AN1, in which a 29.7-kb DNA fragment from Delftia sp. AN3 was inserted. Sequencing and open reading frame （orfs） analysis of this 29.7 kb fragment revealed that it contained at least 27 orfs, among them a gene cluster （consisting of at least 16 genes, named danQTA1A2BRDCEFG1HIJKG2） was responsible for the complete metabolism of aniline to TCA-cycle intermediates. This gene cluster could be divided into two main parts, the upper sequences consisted of 7 genes （danQTA1A2BRD） were predicted to encode a multi-component aniline dioxygenase and a LysR-type regulator, and the central genes （danCEFG1HIJKG2） were expected to encode meta-cleavage pathway enzymes for catechol degradation to TCA-cycle intermediates. Unlike clusters tad from Delftia tsuruhatensis AD9 and tdn from Pseudomonas putida UCC22, in this gene cluster, all the genes were in the same transcriptional direction. There was only one set of C230 gene （danC） and ferredoxin-like protein gene （danD）. The presence of only one set of these two genes and specificity of AD and C230 might be the reason for strain AN3 could only degrade aniline. The products of danQTA1A2BRDC showed 99%-100% identity to those from Delftia acidovorans 7N, and 50%-85% identity to those of tad cluster from D. tsuruhatensis AD9 in amino acid residues. Besides this dan cluster, the 29.7 kb fragment also contained genes encoding the trans-membrane transporter and transposases which might be needed for transposition of the gene cluster. Pulsed-field gel electrophoresis （PFGE） and plasmid curing experiments suggested that the dan cluster might be encoded on the chromosome of strain AN3. The GenBank accession number for the dan cluster of Delftia sp. AN3 is DQ661649.展开更多
基金supported by the National Natural Science Foundation of China(No.51508275)the Natural Science Foundation of Jiangsu Province(No.BK20201310)。
文摘Biodegradation mechanisms and microbial functional diversity during coupled pnitrophenol(PNP)and p-aminophenol(PAP)degradation were studied in a bioelectrochemical system.PNP in the biocathode and PAP in the bioanode were almost completely removed within 28 hr and 68 hr respectively.The degradation followed the steps including hydrating hydroxyalkylation,dehydrogenating carbonylation,and hydrolating ring cleavage,etc.Metagemomic analysis based on the KEGG and egg NOG database annotations revealed the microbial composition and functional genes/enzymes related to phenol degradation in the system.The predominant bacteria genera were Lautropia,Pandoraea,Thiobacillus,Ignavibacterium,Truepera and Hyphomicrobium.The recognized biodegradation genes/enzymes related to pollutant degradation were as follows:pmo,hbd,&ppo for phenol degradation,nzba,amie,&badh for aromatic degradation,and CYP&p450 for xenobiotics degradation,etc.The co-occurrence of ARGs(antibiotic resistant genes),such as ade F,Mex J,Erm F,PDC-93 and Escherichiacolimdf A,etc.,were annotated in CARD database during the biodegradation process.The Proteobacteria&Actinobacteria phylum was the primary host of both the biodegradation genes&ARGs in this system.The microbial functional diversity ensured the effective biodegradation of the phenol pollutants in the bioelectrochemical system.
文摘A recombinant strain, Escherichia coli JM109-AN1, was obtained by constructing of a genomic library of the total DNA of Delftia sp. AN3 in E. coli JM109 and screening for catechol 2,3-dioxygenase activity. This recombinant strain could grow on aniline as sole carbon, nitrogen and energy source. Enzymatic assays revealed that the exogenous genes including aniline dioxygenase （AD） and catechol 2,3-dioxygenase （C230） genes could well express in the recombinant strain with the activities of AD and C230 up to 0.31 U/mg wet cell and 1.92 U/mg crude proteins, respectively. The AD or C23O of strain AN3 could only catalyze aniline or catechol but not any other substituted substrates. This recombinant strain contained a recombinant plasmid, pKC505-AN1, in which a 29.7-kb DNA fragment from Delftia sp. AN3 was inserted. Sequencing and open reading frame （orfs） analysis of this 29.7 kb fragment revealed that it contained at least 27 orfs, among them a gene cluster （consisting of at least 16 genes, named danQTA1A2BRDCEFG1HIJKG2） was responsible for the complete metabolism of aniline to TCA-cycle intermediates. This gene cluster could be divided into two main parts, the upper sequences consisted of 7 genes （danQTA1A2BRD） were predicted to encode a multi-component aniline dioxygenase and a LysR-type regulator, and the central genes （danCEFG1HIJKG2） were expected to encode meta-cleavage pathway enzymes for catechol degradation to TCA-cycle intermediates. Unlike clusters tad from Delftia tsuruhatensis AD9 and tdn from Pseudomonas putida UCC22, in this gene cluster, all the genes were in the same transcriptional direction. There was only one set of C230 gene （danC） and ferredoxin-like protein gene （danD）. The presence of only one set of these two genes and specificity of AD and C230 might be the reason for strain AN3 could only degrade aniline. The products of danQTA1A2BRDC showed 99%-100% identity to those from Delftia acidovorans 7N, and 50%-85% identity to those of tad cluster from D. tsuruhatensis AD9 in amino acid residues. Besides this dan cluster, the 29.7 kb fragment also contained genes encoding the trans-membrane transporter and transposases which might be needed for transposition of the gene cluster. Pulsed-field gel electrophoresis （PFGE） and plasmid curing experiments suggested that the dan cluster might be encoded on the chromosome of strain AN3. The GenBank accession number for the dan cluster of Delftia sp. AN3 is DQ661649.
