Rapid On-Site Monitoring of Bacteria in Freshwater Environments Using a Portable Microfluidic Counting System
Rapid On-Site Monitoring of Bacteria in Freshwater Environments Using a Portable Microfluidic Counting System
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DOI：
10.1248/bpb.b19-00549
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发表时间：
2020-01-01
影响因子：
2
通讯作者：
Fujii, Yudai
中科院分区：
文献类型：
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作者：
Yamaguchi, Nobuyasu;Fujii, Yudai
淡水环境和天然水上公园是重要的娱乐场所;然而，在河边或湖边享受娱乐的人有时会感染病原微生物。微生物监测是日常水质评价的基础。荧光染色技术被认为是最有用的快速微生物学方法之一;然而，荧光显微镜样品的制备通常是劳动密集型的，并且通常必须将样品带到实验室进行测量，这通常会改变样品中细菌的可培养性。这些因素创造了一个快速和简单的方法，可以在现场进行淡水中的细菌定量的需求。在这项研究中，我们应用我们的微流控装置，这是最初设计的芯片上的荧光染色和半自动计数的目标微生物细胞与荧光抗体染色，计数淡水中的细菌细胞。这与自制的便携式系统相结合，用于细菌细胞的快速现场监测。采用6-羧基荧光素二乙酸酯和SYBR绿色II分别对池塘水样进行芯片染色，利用便携式微流控计数系统成功地测定了池塘水样中的酯酶活性细菌和细菌总数。计数在1小时内完成（淡水预过滤30分钟，片上染色和计数30分钟）。结果表明，该方法可快速、准确地对淡水中的细菌进行计数，可用于淡水微生物质量控制中的“现场初筛”。
Freshwater environments and natural water parks are important as recreation areas; however, people enjoying recreation at the river- or lake-side are sometimes infected with pathogenic microbes. Microbiological monitoring is fundamental for the routine evaluation of water quality. Fluorescent staining techniques are regarded as among the most useful rapid microbiological methods; however, preparation of samples for fluorescence microscopy is often labor-intensive, and one usually has to take the samples to a laboratory for measurement, which often alters the culturability of bacteria in the samples. These factors have created demand for a rapid and simple method of bacterial quantification in freshwater that can be performed on-site. In this study, we applied our microfluidic device, which was originally designed for on-chip fluorescent staining and semi-automated counting of target microbial cells with fluorescent antibody-staining, to enumerate bacterial cells in freshwater. This was combined with a self-made portable system for rapid on-site monitoring of the bacterial cells. Numbers of both esterase-active bacteria and total bacteria in pond water samples could be successfully determined by on-chip staining with 6-carboxyfluorescein diacetate and SYBR Green II, respectively, using the portable microfluidic counting system. The counting was completed within 1h (30 min for pre-filtration of freshwater and 30 min for on-chip staining and counting). These results indicate that rapid and accurate counting of bacterial cells in freshwater can be performed and this technique could be applied for "on-site first screening" purposes in microbial quality control of freshwater.