A biosensor platform for rapid detection of E. coli in drinking water

Abstract

The need for rapid, specific and sensitive assays that provide a detection of bacterial indicators are important for monitoring water quality. Rapid detection using biosensor is a novel approach for microbiological testing applications. Besides, validation of rapid methods is an obstacle in adoption of such new bio-sensing technologies. In this study, the strategy developed is based on using the compound 4-methylumbelliferyl glucuronide (MUG), which is hydrolyzed rapidly by the action of E. coli β-D-glucuronidase (GUD) enzyme to yield a fluorogenic product that can be quantified and directly related to the number of E. coli cells present in water samples.The detection time required for the biosensor response ranged from 30 to 120 minutes, depending on the number of bacteria. The specificity of the MUG based biosensor platform assay for the detection of E. coli was examined by pure cultures of non-target bacterial genera and also non-target substrates. GUD activity was found to be specific for E. coli and no such enzymatic activity was detected in other species. Moreover, the sensitivity of rapid enzymatic assays was investigated and repeatedly determined to be less than 10 E. coli cells per reaction vial concentrated from 100 mL of water samples.The applicability of the method was tested by performing fluorescence assays under pure and mixed bacterial flora in environmental samples. In addition, the procedural QA/QC for routine monitoring of drinking water samples have been validated by comparing the performance of the biosensor platform for the detection of E. coli and culture-based standard techniques such as Membrane Filtration (MF). The results of this study ii indicated that the fluorescence signals generated in samples using specific substrate molecules can be utilized to develop a bio-sensing platform for the detection of E. coli in drinking water. The procedural QA/QC of the biosensor will provide both industry and regulatory authorities a useful tool for near real-time monitoring of E. coli in drinking water samples. Furthermore, this system can be applied independently or in conjunction with other methods as a part of an array of biochemical assays in order to reliably detect E. coli in water.

A Strategy to Establish a Quality Assurance/Quality Control Plan for the Application of Biosensors for the Detection of E. coli in Water

“…When E. coli metabolizes Collilert-18’s nutrient-indicator, MUG, the sample also fluoresces. It is reported that the method is able to detect a single viable coliform or E. coli per sample and eliminates false positive detection of non-target organisms [5,6,7,8,9,10,11,12]. For each test, contents of one pack of Collilert-18 were added to a 100 mL sample in a sterile, transparent, non-fluorescing vessel which was then capped and shaken.…”

“…In our previous study, we identified specific bacterial enzymatic-biochemical signatures that can be used in a custom designed opto-electronic biosensor platform for the detection of E. coli and other bacterial cells in water samples [5,6]. The new generations of biosensors rely on bacterial enzymatic responses to specific fluorogenic substrates.…”

“…This study only focuses on the parameters effecting the reaction cocktail and QA/QC procedure of the assay. The effect of water contaminant interferences such as heavy metals on assay sensitivity with lower concentrations of bacteria have been documented in our previous publications [5,6] which focused on the application of this innovative method in environmental water, including water from lakes that receive regular application of copper sulfate for algal control. The result showed that no significant interference from non-GUD sources in the water samples existed.…”

Rapid Water Quality Monitoring for Microbial Contamination

“…42 With this handheld fluorescence detector, the E. coli concentrations of 7 CFU/mL are detected within 30-60 min after testing. Similarly, Hesari, et al, 43 developed custom designed fluorescence biosensor to detect E. coli. This method also uses the enzymatic activity of the MUG substrate with the β-D-glucuronidase enzyme.…”

“…Several sensors for water quality monitoring have been designed and developed for rapid detection of bacteria (total coliform and E. coli) in water samples. They are a paper-based microfluidic method, handheld fluorometer, [42][43][44] Mobile Water Kit, [45][46][47] hydrogel-based plunger-tube assembly, [48][49][50] and glucometer based detection. 51 We provide the details of each method briefly.…”