高效反硝化聚磷菌的筛选及其脱氮除磷条件和性能研究

doi:10.13560/j.cnki.biotech.bull.1985.2022-1522

摘要/Abstract

摘要：

筛选高效反硝化聚磷菌，探究其脱氮除磷条件及对模拟废水的处理性能。纯培养技术分离反硝化聚磷菌；基于16S rRNA基因序列分析和生理生化特征，完成菌株D4的初步鉴定；通过单因素和响应面法优化脱氮除磷条件，并应用于食品模拟废水的生物脱氮除磷。反硝化聚磷细菌D4为戈登氏菌（Gordonia sp.），脱氮除磷的最佳条件为乙酸钠（3.32 g/L）作碳源、接种量5%、初始磷含量18.61 mg/L、31.3℃、pH 7.9，该条件下菌株D4对总磷、硝氮和氨氮的去除量分别为14.18、39.67和69.71 mg/L，相应去除率为84.50%、97.67%和96.22%；应用于不同模拟废水的结果显示，菌株D4对多种废水均具有良好的脱氮除磷效果，其中对豆制品废水中的氮磷去除效果最好，脱氮量和除磷量分别达101.58 mg/L和10.40 mg/L。戈登氏菌D4具有较好的反硝化聚磷能力，是豆制品废水等高氮高磷类食品加工废水生物脱氮除磷的良好菌种资源。

关键词: 反硝化聚磷菌, 脱氮除磷, 条件优化, 废水处理

Abstract:

The objective is to explore the denitrification and phosphorus removal conditions and performance of a efficient denitrifying phosphorus-accumulating bacterium, and its treatment effect on simulated wastewater. Denitrifying phosphorus-accumulating strain D4 was isolated by pure culture technology. Preliminary identification of the strain was completed based on 16S rRNA gene sequence analysis and physiological and biochemical characteristics, and the denitrification and dephosphorization conditions were optimized by single factor and response surface method, so as to achieve efficient biological denitrification and dephosphorization of mimic food wastewater. As results，denitrifying phosphorus-accumulating bacterium D4 was Gordonia sp.. The optimal conditions for nitrogen and phosphorus removal were sodium acetate（3.32 g/L）as carbon source, 5% inoculum, initial phosphorus content of 18.61 mg/L, 31.3℃, and pH 7.9. Under these conditions, the removed amounts of total phosphorus, nitrate nitrogen and ammonia nitrogen by strain D4 were 14.18, 39.67 and 69.71 mg/L, respectively; and the corresponding removal rates were 84.50%, 97.67% and 96.22%. Results of treatment to different simulated wastewater showed that strain D4 had the best nitrogen and phosphorus removal effects in soybean product wastewater, with the removed amount of nitrogen and phosphorus being 101.58 mg/L and 10.40 mg/L, respectively. In conclusion, strain D4 has promising denitrification and phosphorus-accumulating ability and is a good strain resource for biological denitrification and phosphorus removal of food processing wastewater with high nitrogen and phosphorus such as soybean product wastewater.

Key words: denitrification and phosphorus-accumulating bacterium, denitrification and phosphorus removal, condition optimization, wastewater treatment

袁野, 周佳, 屈建航, 张博源, 罗宇, 李海峰. 高效反硝化聚磷菌的筛选及其脱氮除磷条件和性能研究[J]. 生物技术通报, 2023, 39(7): 266-276.

YUAN Ye, ZHOU Jia, QU Jian-hang, ZHANG Bo-yuan, LUO Yu, LI Hai-feng. Screening of an Efficient Denitrifying Phosphorus-accumulating Bacterium and Its Denitrification and Phosphorus Removal[J]. Biotechnology Bulletin, 2023, 39(7): 266-276.

图/表 12

表1 反硝化聚磷菌三因素三水平表

Table 1 Three factors and three levels of denitrifying phosphorus-accumulating bacteria

因素代码 Code of factor	因素 Factor	水平Level
因素代码 Code of factor	因素 Factor	1	0	1
A	温度 Temperature/℃	20	28	34
B	pH	7	8	9
C	初始磷含量Initial phosphorus content/（mg·L^-1）	10	15	20

图1 菌株D4的16S rRNA基因系统发育树括号内为菌株GenBank登录号；分支节点数字为Bootstrap值（低于70不显示）；标尺的数据为进化距离

Fig. 1 Phylogenetic tree based on the 16S rRNA genes of strain D4 Code in parentheses is GenBank accession number of strain.The number of branch nodes is bootstrap value（That of lower than 70 is not displayed）. The data of the scale is the evolutionary distance

图2 菌株D4的生长曲线和脱氮除磷曲线

Fig. 2 Growth and nitrogen and phosphorus removal curves of strain D4

图3 碳源及碳源浓度对菌株D4脱氮除磷效率的影响不同小写字母表示在P<0.05水平差异显著。下同

Fig. 3 Effects of carbon sources and carbon source concentration on the nitrogen and phosphorus removal efficiency of strain D4 Different lower letters indicate significant difference at P<0.05 level. The same below

图4 接种量对D4脱氮除磷效率的影响

Fig. 4 Effects of inoculum size on nitrogen and phosphorus removal efficiency of D4

图5 初始磷含量对D4脱氮除磷效率的影响

Fig. 5 Effects of initial phosphorus concentration on the nitrogen and phosphorus removal efficiency of D4

图6 温度对D4脱氮除磷效率的影响

Fig. 6 Effects of temperature on the nitrogen and phosphorus removal efficiency of D4

图7 pH对D4脱氮除磷效率的影响

Fig. 7 Effects of pH on the nitrogen and phosphorus removal efficiency of D4

表2 显著性分析结果

Table 2 Significance analysis results

因素Source	平方和Sum of squares	自由度Freedom	均方Mean square	F值F value	显著性Significance
碳源Carbon source	143.696	4	35.924	376.090	0.000**
乙酸钠浓度Sodium acetate concentration	43.381	4	10.845	583.658	0.000**
接种量Vaccination percent	0.064	4	0.016	0.535	0.714
初始磷含量Initial phosphorus content	49.980	4	12.495	114.939	0.000**
温度Temperature	75.747	4	18.937	288.322	0.000**
pH	150.384	4	37.593	1492.194	0.000**

表3 二次模型的方差分析

Table 3 Analysis of variance of quadratic model

来源Source	平方和Sum of squares	自由度Freedom	均方Mean square	F值F value	P值P value	显著性Significance
模型	139.61	9	15.51	22.05	0.0002	**
A	15.89	1	15.89	22.59	0.0021	**
B	2.02	1	2.02	2.88	0.1337	-
C	53.54	1	53.54	76.12	< 0.0001	**
AB	2.52	1	2.52	3.58	0.1002	-
AC	9.49	1	9.49	13.50	0.0079	**
BC	0.0038	1	0.0038	0.0055	0.9431	-
A²	13.90	1	13.90	19.75	0.0030	**
B²	8.09	1	8.09	11.51	0.0116	*
C²	8.81	1	8.81	12.53	0.0095	**
残值	4.92	7	0.7034
失拟项	3.68	3	1.23	3.96	0.1083	-
纯误差	1.24	4	0.3099
总计	144.54	16

图8 各因素交互作用对总磷去除量的响应曲面图 a：温度和pH的交互作用图；b：温度和初始磷含量的交互作用图；c： pH和初始磷含量的交互作用图

Fig. 8 Response surface diagram of interaction of various factors on total phosphorus removal a: The interaction of temperature and pH; b: the interaction of temperature and initial phosphorus concentration; c: the interaction of pH and initial phosphorus concentration

图9 菌株D4对不同模拟废水中总磷（a）、总氮（b）、氨氮（c）的去除量

Fig. 9 Removed amount of total phosphorus (a), total nitrogen (b), ammonia nitrogen (c) in different simulated wastewater by bacterial strain D4

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