A flow cytometer based protocol for quantitative analysis of bloom-forming cyanobacteria(Microcystis) in lake sediments  

作　　者：Quan Zhou Wei Chen Huiyong Zhang Liang Peng Liming Liu Zhiguo Han Neng Wan Lin Li Lirong Song 

机构地区：[1]State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China [2]Graduate School of Chinese Academy of Sciences, Beijing 100039, China [3]Zealquest Laboratory for Ecological Research, Zealquest Scientific Technology Co., Ltd., Shanghai 200333, China [4]Changshu Institute of Technology, Changshu 215500, China

出　　处：《Journal of Environmental Sciences》2012年第9期1709-1716,共8页环境科学学报（英文版）

基　　金：supported by the National Basic Research Program (973) of China (No. 2008CB418006);the National Special Program of Water Environment (No.2009ZX07106-001-002);the National Natural Science Foundation of China (No. 31070355);the National Major Science and Technology Program for Water Pollution Control and Treatment (No. 2009ZX07101-013)

摘　　要：A quantitative protocol for the rapid analysis of Microcystis cells and colonies in lake sediment was developed using a modified flow cytometer, the CytoSense. For cell enumeration, diluted sediment samples containing Microcystis were processed with sonication to disintegrate colonies into single cells. An optimized procedure suggested that 5 mg dw （dry weight）/mL dilution combined with 200 W x 2 min sonication yielded the highest counting efficiency. Under the optimized determination conditions, the quantification limit of this protocol was 3.3x104 cells/g dw. For colony analysis, Microcystis were isolated from the sediment by filtration. Colony lengths measured by flow cytometry were similar to those measured by microscopy for the size range of one single cell to almost 400 ~tm in length. Moreover, the relationship between colony size and cell number was determined for three Microcystis species, including Microcystisflos-aquae, M. aeruginosa and M. wessenbergii. Regression formulas were used to calculate the cell numbers in different- sized colonies. The developed protocol was applied to field sediment samples from Lake Taihu. The results indicated the potential and applicability of flow cytometry as a tool for the rapid analysis of benthic Microcystis. This study provided a new capability for the high frequency monitoring of benthic overwintering and population dynamics of this bloom-forming cyanobacterium.A quantitative protocol for the rapid analysis of Microcystis cells and colonies in lake sediment was developed using a modified flow cytometer, the CytoSense. For cell enumeration, diluted sediment samples containing Microcystis were processed with sonication to disintegrate colonies into single cells. An optimized procedure suggested that 5 mg dw （dry weight）/mL dilution combined with 200 W x 2 min sonication yielded the highest counting efficiency. Under the optimized determination conditions, the quantification limit of this protocol was 3.3x104 cells/g dw. For colony analysis, Microcystis were isolated from the sediment by filtration. Colony lengths measured by flow cytometry were similar to those measured by microscopy for the size range of one single cell to almost 400 ~tm in length. Moreover, the relationship between colony size and cell number was determined for three Microcystis species, including Microcystisflos-aquae, M. aeruginosa and M. wessenbergii. Regression formulas were used to calculate the cell numbers in different- sized colonies. The developed protocol was applied to field sediment samples from Lake Taihu. The results indicated the potential and applicability of flow cytometry as a tool for the rapid analysis of benthic Microcystis. This study provided a new capability for the high frequency monitoring of benthic overwintering and population dynamics of this bloom-forming cyanobacterium.

关 键 词：flow cytometry benthic Microcystis cell enumeration colony size measurement 

分 类 号：Q2-33[生物学—细胞生物学] P512.2[天文地球—地质学]