一株高效异养硝化菌的选育、鉴定及其硝化条件

Isolation, identification and characterization of a high-efficient heterotrophic nitrifier

摘要:

摘要：【目的】针对现阶段异养硝化菌硝化速率较低的问题，选育更高效的异养硝化菌，进而鉴定该菌株的种属，了解其硝化特性和硝化条件。【方法】分别从污水处理厂活性污泥、化肥厂土壤以及农田土壤中取样，以柠檬酸钠为碳源，NH4Cl为氮源，采用污泥驯化、驯化过程中驯化液连续梯度稀释、平板划线分离及颜色指示剂快速硝化效果检测等步骤，筛得一株高效的异养硝化菌。经生理生化和16S rDNA序列的系统发育分析鉴定其种属；将该菌接入人工氨氮废水，定时检测水中含氮化合物的变化，了解其硝化特性；通过改变培养基碳源、溶氧量、C/N比、温度和pH考察其硝化条件。【结果】获得的高效异养硝化菌为革兰氏阴性杆菌，不利用葡萄糖发酵，氧化酶、接触酶阳性，不产吲哚，能由有机酸盐产碱；其与产碱菌属菌株Alcaligenes sp. ES-SDK-3的16S rDNA同源性高达99.7%。用该菌株处理初始氨氮浓度为182.30 mg/L的废水，30 h后氨氮去除率为99.8%，指数期平均氨氮去除速率为9.61 mg-N/L/h，其在硝化过程中几乎没有亚硝酸盐氮和硝酸盐氮产生；最佳碳源为柠檬酸钠；高的溶氧量和高的C/N比有利于其降解氨氮，当C/N比为12时即可达到较好的效果；该菌株在温度为30℃－35℃，pH为5.0－9.0范围内均能较彻底地降解氨氮。【结论】该菌株为产碱菌属，命名为Alcaligenes sp. HN-S；其在硝化速率与处理的氨氮浓度方面均高于目前国内外筛出的大多数异养硝化菌；通过考察其硝化条件，为其走向实际污水脱氮工艺提供了依据。

Abstract:

Abstract: [Objective] In order to improve the rate of the heterotrophic nitrification, we screened and identified a high-efficient heterotrophic nitrifier, as well as studied its nitrification characteristics and nitrification conditions.[Methods] We obtained activated sludge samples from sewage and chemical fertilizer factories and farmland. We then utilized sodium citrate and ammonium chloride as carbon and nitrogen source. We used methods including domestication, gradient dilution of domestication liquid, isolation from streaking plate and color indicator as rapid nitrification detection. Finally a high-efficient heterotrophic nitrifier was obtained. We identified this strain according to its physiological, biochemical properties and the sequence analysis of 16S rDNA. After inoculating the strain into artificial ammonia-nitrogen wastewater, changes of nitrogen compounds were measured in order to understand the nitrification characteristics. Nitrification condition was also optimized by changing the carbon source, dissolved oxygen, C/N ratio, temperature and pH of the medium. [Results] The heterotrophic nitrifier was a gram-negative bacilli. It neither fermented glucose, nor produced indole. Oxidase and catalase tests were positive. It could produce alkali if organic salt was provided. The strain shared 99.7% sequence identity of its 16S rDNA with ES-SDK-3 of Alcaligenes sp. In the artificial wastewater with 182.30 mg/L ammonia nitrogen as initial concentration, the removal efficiency by the strain was 99.8% after 30h cultivation. The average nitrogen removal rate was 9.61mg-N/L/h in its exponential phase. It produced almost no NO2―-N and NO3―-N in the entire nitrification process. The optimal carbon source is sodium citrate. Higher dissolved oxygen and C/N ratio favor its nitrification. When temperature is ranged from 30℃ to 35℃ and pH is ranged from 5.0 to 9.0, it can completely remove ammonia nitrogen. [Conclusions] The strain was identified as Alcaligenes genus, and named as Alcaligenes sp. HN-S. Our research confirmed that the Alcaligenes sp. HN-S had significant advantages over heterotrophic nitrifiers that were screened previously with aspect of ammonia nitrogen removal rate. The research of its nitrification condition definitely provided necessary theory support for a new biology process to remove nitrogen with high efficiency.