Noble Rachel T, Weisberg Stephen B
University of North Carolina at Chapel Hill, Institute of Marine Sciences, 3431 Arendell St, Morehead City, NC 28557, USA.
J Water Health. 2005 Dec;3(4):381-92. doi: 10.2166/wh.2005.051.
Monitoring of recreational beaches for fecal indicator bacteria is currently performed using culture-based technology that can require more than a day for laboratory analysis, during which time swimmers are at risk. Here we review new methods that have the potential to reduce the measurement period to less than an hour. These methods generally involve two steps. The first is target capture, in which the microbial group of interest (or some molecular/chemical/or biochemical signature of the group) is removed, tagged or amplified to differentiate it from the remaining material in the sample. We discuss three classes of capture methods: 1) Surface and whole-cell recognition methods, including immunoassay techniques and molecule-specific probes; 2) Nucleic acid methods, including polymerase chain reaction (PCR), quantitative PCR (Q-PCR), nucleic acid sequence based amplification (NASBA) and microarrays; and 3) Enzyme/substrate methods utilizing chromogenic or fluorogenic substrates. The second step is detection, in which optical, electrochemical or piezoelectric technologies are used to quantify the captured, tagged or amplified material. The biggest technological hurdle for all of these methods is sensitivity, as EPA's recommended bathing water standard is less than one cell per ml and most detection technologies measure sample volumes less than 1 ml. This challenge is being overcome through addition of preconcentration or enrichment steps, which have the potential to boost sensitivity without the need to develop new detector technology. The second hurdle is demonstrating a relationship to health risk, since most new methods are based on measuring cell structure without assessing viability and may not relate to current water quality standards that were developed in epidemiology studies using culture-based methods. Enzyme/substrate methods may be the first rapid methods adopted because they are based on the same capture technology as currently-approved EPA methods and their relationship to health risk can be established by demonstrating equivalency to existing procedures. Demonstration of equivalency may also be possible for some surface and whole-cell recognition methods that capture bacteria in a potentially viable state. Nucleic acid technologies are the most versatile, but measure nonviable structure and will require inclusion in epidemiological studies to link their measurement with health risk.
目前，对休闲海滩的粪便指示菌监测是采用基于培养的技术进行的，这种技术可能需要一天多的时间进行实验室分析，在此期间游泳者会面临风险。在此，我们回顾了一些有可能将测量时间缩短至不到一小时的新方法。这些方法通常包括两个步骤。第一步是目标捕获，即去除、标记或扩增感兴趣的微生物组（或该组的一些分子/化学/或生化特征），以便将其与样品中的其余物质区分开来。我们讨论了三类捕获方法：1）表面和全细胞识别方法，包括免疫测定技术和分子特异性探针；2）核酸方法，包括聚合酶链反应（PCR）、定量PCR（Q-PCR）、基于核酸序列的扩增（NASBA）和微阵列；3）利用显色或荧光底物的酶/底物方法。第二步是检测，即使用光学、电化学或压电技术对捕获、标记或扩增的物质进行定量。所有这些方法最大的技术障碍是灵敏度，因为美国环境保护局（EPA）推荐的游泳用水标准是每毫升少于一个细胞，而大多数检测技术测量的样品体积小于1毫升。通过添加预浓缩或富集步骤正在克服这一挑战，这些步骤有可能提高灵敏度而无需开发新的检测技术。第二个障碍是证明与健康风险的关系，因为大多数新方法是基于测量细胞结构而不评估生存能力，可能与使用基于培养的方法在流行病学研究中制定的当前水质标准无关。酶/底物方法可能是首先采用的快速方法，因为它们基于与目前已批准的EPA方法相同的捕获技术，并且可以通过证明与现有程序等效来确立它们与健康风险的关系。对于一些以潜在存活状态捕获细菌的表面和全细胞识别方法，也有可能证明等效性。核酸技术用途最广泛，但测量的是非存活结构，需要纳入流行病学研究以将其测量结果与健康风险联系起来。
