异养硝化-好氧反硝化菌Acinetobacter johnsonii sp. N26的脱氮性能及代谢途径

Optimization of nitrogen removal performance and metabolic pathway of a heterotrophic nitrifying-aerobic denitrifying bacterial strain Acinetobacter johnsonii sp. N26

摘要:

【背景】水体中含氮物质的大量累积会造成水体富营养化、水生生物死亡等问题，严重威胁水生态环境，制约我国环境保护的持续发展。【目的】为去除生活污水中的含氮污染物，从羊粪堆肥中筛选出了一株具有异养硝化-好氧反硝化功能的细菌——约氏不动杆菌Acinetobacter johnsonii sp. N26，研究其脱氮性能和代谢途径。【方法】测定菌株N26在氨氮和硝态氮中的生长和脱氮曲线，通过单因素试验对其脱氮性能进行优化，通过氮平衡分析和功能基因鉴定研究其脱氮代谢途径。【结果】生长和脱氮曲线表明，菌株N26对初始浓度均为50 mg/L的氨氮和硝态氮的去除速度快、效率高，其中9 h内对氨氮的去除效率为95.5%，最大去除速率为5.330 mg/(L·h)；15 h内对硝态氮的去除效率为93.6%，最大去除速率为3.147 mg/(L·h)，且最终仅有少量硝酸盐、亚硝酸盐积累。脱氮性能优化结果表明，该菌株的最适氮源为氯化铵，最适碳源为丁二酸钠，最适温度为30 ℃，最适接种量为15%，最适pH值为8.0-9.0，最适碳氮比为15，最适转速为120 r/min，最适氮负荷≤300 mg/L (氨氮)。氮平衡分析和功能基因鉴定结果表明，该菌种的脱氮不仅是异养硝化-好氧反硝化过程，还是一个短程硝化-反硝化过程，其脱氮代谢途径为NH₄⁺-N→NO₂^--N→ NO→N₂O→N₂和NO₃^--N→NO₂^--N→NO→N₂O→N₂。【结论】Acinetobacter johnsonii sp. N26菌株具有优异的脱氮特性，在治理水体氨氮污染方面具备应用潜力，该结果可为异养硝化-好氧反硝化微生物应用于生活污水的生物脱氮处理提供理论依据。

关键词:异养硝化-好氧反硝化;约氏不动杆菌;生长曲线;碳源;脱氮途径;功能基因

Abstract:

[Background] The accumulation of nitrogenous substances in water bodies will cause problems such as eutrophication and death of aquatic organisms, which seriously threatens the aquatic environment and restricts the environmental protection for the sustainable development of China. [Objective] To study the nitrogen removal performance and metabolic pathway of Acinetobacter johnsonii sp. N26, a bacterial strain with heterotrophic nitrifying-aerobic denitrifying function and screened out from sheep manure compost, for the removal of nitrogen pollutants in domestic sewage. [Methods] The growth and nitrogen removal curves of N26 in the media with ammonia nitrogen and nitrate nitrogen were established. The nitrogen removal performance of the strain was optimized by single factor experiment, and the metabolic pathway was studied by nitrogen balance analysis and functional gene identification. [Results] The growth and nitrogen removal curves illustrated that N26 rapidly removed ammonia nitrogen and nitrate nitrogen (initial concentration of 50 mg/L) with high efficiency. For the removal of ammonia nitrogen, the strain showed the removal efficiency of 95.5% and the maximum removal rate of 5.330 mg/(L·h) within 9 h. For the removal of nitrate nitrogen, the strain showed the removal efficiency of 93.6% and the maximum removal rate of 3.147 mg/(L·h) within 15 h, and only a small amount of nitrate and nitrite were accumulated in the end. The strain had the optimum nitrogen removal performance in the medium with ammonium chloride as the nitrogen source and sodium succinate as the carbon source at 30 ℃, the inoculum amount of 15%, pH 8.0–9.0, C/N ratio of 15, rotating speed of 120 r/min, and nitrogen load ≤300 mg/L (ammonia nitrogen). The results of nitrogen balance analysis and functional gene identification indicated that the nitrogen removal process of this strain was not only in line with heterotrophic nitrification-aerobic denitrification but also a short-range nitrification-denitrification process. The metabolic pathways of this strain for nitrogen removal were NH₄⁺-N→NO₂^--N→NO→N₂O→N₂ and NO₃^--N→NO₂^--N→NO→ N₂O→N₂. [Conclusion] A. johnsonii sp. N26 has excellent performance of heterotrophic nitrification-aerobic denitrification and application potential in the treatment of ammonia nitrogen pollution in water bodies. The results can provide a theoretical basis for the application of heterotrophic nitrifying-aerobic denitrifying microorganisms to the biological removal of nitrogen in domestic sewage.

Key words:heterotrophic nitrification-aerobic denitrification;Acinetobacter johnsonii;growth curve;carbon source;nitrogen removal pathway;functional genes