Isolation of a p-Nitrophenol-Degrading Bacterium and Investigation of Its Degrading Mechanism

doi:10.13560/j.cnki.biotech.bull.1985.2019-0286

Abstract

Abstract: This work is designed to isolate marine microbes that can efficiently degrade p-nitrophenol and to systematically elucidate related mechanism. The sole carbon source method was employed for the enrichment,domestication and isolation of marine microbes degrading p-nitrophenol. The adaptability of isolated strains to different environmental factors was investigated by single factor experiment. The metabolic pathway of p-nitrophenol was deduced by mass spectrometry analysis of metabolic intermediates. Genes involved in the metabolism of p-nitrophenol were obtained by gene cloning. One marine bacterium RL-JY1,which efficiently degraded p-nitrophenol,was isolated. 100 mg/L p-nitrophenol was completely degraded by the strain RL-JY1 within 72 h. Strain RL-JY1 was identified as Pseudomonas putida via the analysis of morphological,physiological and biochemical characteristics and 16S rRNA gene sequence. The degradation rates of 100 mg/L p-nitrophenol were 97.2%,100% and 100% when RL-JY1 was incubated under 20,30 and 40℃,respectively. The degradation rates of 100 mg/L p-nitrophenol were 64.7%,100%,97.2% and 84.2% within 72 h when the initial pH was 6.0,7.0,8.0 and 9.0,respectively. When NaCl concentration was under 0-8%（W/V）,all the degradation rates of 100 mg/L p-nitrophenol within 72 h were 100%. When NaCl concentration was 10% and 12%,the degradation rates of 100 mg/L p-nitrophenol within 72 h were 81.3% and 50.6%,respectively. The metabolic pathway of p-nitrophenol in strain RL-JY1 was deduced via metabolic intermediates identification through mass spectrometry analysis,which was a typical 1,2,4-benzenetriol pathway. Gene pnpABC,involved in the transformation of p-nitrophenol to maleacetic acid,was cloned from the genome of strain RL-JY1 according to known p-nitrophenol degrading related genes. The sequence alignments indicated that the obtained pnpABC gene showed high similarity（all above 99%）with the sequences reported in P. putida DLL-E4. In conclusion,p-Nitrophenol degrading bacterium RL-JY1 was identified as P. putida,and showed promising tolerance to environmental temperature,pH and salinity. p-Nitrophenol was utilized via 1,2,4-benzenetriol pathway by strain RL-JY1. p-Nitrophenol degrading related genes pnpABC were identified in the genome of strain RL-JY1.

Key words: p-Nitrophenol, Pseudomonas putida, biodegradation, salt tolerance, molecular mechanism

REN Lei, LIU Bin, LIN Zhong, ZHEN Zhen, LIU Yue-lian, HU Han-qiao, YAN Yan-chun. Isolation of a p-Nitrophenol-Degrading Bacterium and Investigation of Its Degrading Mechanism[J]. Biotechnology Bulletin, 2019, 35(9): 184-193.

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