Simple and reliable enumeration of Escherichia coli concentrations in wastewater samples by measuring β-d-glucuronidase (GUS) activities via a microplate reader
Simple and reliable enumeration of Escherichia coli concentrations in wastewater samples by measuring β-d-glucuronidase (GUS) activities via a microplate reader
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通过酶标仪测量 β-d-葡萄糖醛酸酶 (GUS) 活性，简单可靠地计数废水样品中的大肠杆菌浓度
DOI：
10.1016/j.scitotenv.2020.136928
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发表时间：
2020
影响因子：
9.8
通讯作者：
Okabe Satoshi
中科院分区：
文献类型：
--
作者：
Satoh Hisashi;Kikuchi Kai;Katayose Yutaka;Tsuda Shu;Hirano Reiko;Hirakata Yuga;Kitajima Masaaki;Ishii Satoshi;Oshiki Mamoru;Hatamoto Masashi;Takahashi Masahiro;Okabe Satoshi
污水处理厂大肠杆菌浓度的监测对保证工艺性能和保护公众健康具有重要意义。然而，传统的E.大肠杆菌计数方法复杂且耗时耗力。本文报道了一种新的基于β-D-葡萄糖醛酸酶（GUS）活性测定的简单可靠的方法。废水（WW）样品中的共浓度。一等份（20 μL）含E.将180 μL WW样品加入96孔微孔板的一个孔中。将微孔板置于37 °C的微孔板读数器中。为此，荧光强度的荧光酶底物为E。在3小时内每10分钟测量一次大肠杆菌以确定GUS活性。代表GUS活性的荧光强度的线性增加与E.废水样品中的浓度。然而，相关方程是特定的污水处理厂，这可能是由于在E的差异。污水处理厂中的大肠菌群结构。我们观察到废水基质不是测量GUS活性的限制，并且可以使用废水处理厂特定的相关方程作为校准曲线来估计E。从该地点收集的样本中的浓度。将所得结果与培养相关Colilert法的结果进行了比较，证明了该方法简单、实用，可用于大肠杆菌的计数。废水样品中的浓度可靠。
Monitoring ofEscherichia coliconcentrations at wastewater treatment plants (WWTPs) is important to ensure process performance and protect public health. However, conventionalE. colienumeration methods are complicated and time- and labor-consuming. Here, we report a novel simple and reliable method based on β-d-glucuronidase (GUS) activity assay to enumerateE. coliconcentrations in wastewater (WW) samples. An aliquot (20 μL) of the medium with fluorogenic enzyme substrate forE. coliand 180 μL of a WW sample were added to one well of a 96-well microplate. The microplate was placed in a microplate reader at 37 °C. To this end, the fluorescence intensity of a fluorogenic enzyme substrate forE. coliwas measured every 10 min over 3 h to determine GUS activity. The linear increase in the fluorescence intensity representing the GUS activities showed a positive correlation withE. coliconcentrations in wastewater samples. However, the correlation equations were specific to WWTPs, which could be due to the difference in theE. colipopulation structures among WWTPs. We observed that the wastewater matrix is not a limitation to measure the GUS activity, and a WWTP-specific correlation equation can be used as a calibration curve to estimate theE. coliconcentrations in the samples collected from that site. A comparison of the results with those of culture-dependent Colilert method proved that the current method is simple and useful for the enumeration ofE. coliconcentrations in wastewater samples reliably.