研究报告
秦雯,骆阳,王志红,宋阳,刘立凡,李布林,李伟光.滤池滤料上细菌数的预处理方法优化[J].环境科学学报,2020,40(10):3726-3734
滤池滤料上细菌数的预处理方法优化
- The optimized pretreatment method for flow-cytometric quantification of microbial cells on activated carbon from BAC filters
- 基金项目：国家自然科学基金面上项目（No.51578178）；广东省自然科学基金面上项目（No.2020A1515010719）；国家自然科学基金青年基金项目（No.51908143）；广东省教育厅创新强校项目-青年创新人才类项目（No.2018KQNCX194）
- 作者
- 单位
- 秦雯
- 广东工业大学土木与交通工程学院, 广州 510006
- 骆阳
- 广东工业大学土木与交通工程学院, 广州 510006
- 王志红
- 广东工业大学土木与交通工程学院, 广州 510006
- 宋阳
- 广东工业大学土木与交通工程学院, 广州 510006
- 刘立凡
- 广东工业大学土木与交通工程学院, 广州 510006
- 李布林
- 广东工业大学土木与交通工程学院, 广州 510006
- 李伟光
- 1. 哈尔滨工业大学环境学院, 哈尔滨 150090;2. 城市水资源与水环境国家重点实验室(哈尔滨工业大学), 哈尔滨 150090
- 摘要：为优化生物活性炭滤池活性炭上细菌数的流式细胞术定量分析的预处理方法，本研究考察了4种物理预处理方式（机械振荡、低能量超声、高能量超声、机械振荡耦合高能量超声）以及4种化学预处理方式（含10%的柠檬酸（Citric Acid，CA）和2.5%的羟基乙酸（Glycolic acid，GA）混合液（CA+GA）、溶菌酶、吐温20、乙二胺四乙酸）对流式细胞术测定活性炭滤料上细菌数的影响，结果表明，在无菌超纯水为提取液时，机械振荡耦合高能量超声预处理后测定的细菌数最高（9.7（±1.2）×107 cells·g-1（以WW活性炭计，下同）），是最佳的物理预处理方式.0.1% CA+GA为最优的化学分散剂，耦合机械振荡与1次高能量超声预处理后细胞累积回收率较空白组（无菌超纯水）提高了21%±17%，其活细菌数也最高（约1.0×107 cells·g-1）；耦合机械振荡与3次高能量超声预处理后，分离的生物膜细菌量最高（1.5（±0.2）×108 cells·g-1），证明0.1% CA+GA耦合机械振荡与3次高能量超声是利用流式细胞术测定活性炭生物膜的细菌数的最佳预处理方式.
- Abstract：To optimize the pretreatment method of flow cytometric quantification of microbial cells on activated carbon from biological activated carbon filters, four physical pretreatment methods, including mechanical oscillation, low energy sonication, high energy sonication, mechanical oscillation and high energy sonication coupling, and four chemical pretreatments (four dispersants, i.e., CA+GA, containing 10% citric acid (CA) and 2.5% glycolic acid (GA), lysozyme, Tween 20 and ethylenediaminetetraacetic acid) were investigated in this study. The results showed that mechanical oscillation and high energy sonication coupling pretreatment was the best physical pretreatment method, which obtained the highest biomass (9.7(±1.2)×107 cells·g-1 WW activated carbon) by flow cytometric quantification when the sterile ultrapure water was the extraction solution. 0.1% CA+GA was the best chemical dispersant because the cumulative recovery rate of cells after 1 cycle of mechanical oscillation and 1 cycle of high energy sonication pretreatment was 21%±17% higher than that of the blank group (sterile ultrapure water), and the intact bacteria count was also the highest (about 1.0×107 Cells·g-1). 0.1% CA+GA as the extraction solution, coupling with one cycle of mechanical oscillation and 3 cycles of high energy sonication pretreatment which performed the highest amount of bacteria (1.5(±0.2)×108 cells·g-1), was consequently the optimal pretreatment method for flow cytometric quantification of microbial cells on activated carbon from biological activated carbon filters in this study.
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