Digital E. coli Counter: A Microfluidics and Computer Vision-Based DNAzyme Method for the Isolation and Specific Detection of E. coli from Water Samples

Biological water contamination detection-based assays are essential to test water quality; however, these assays are prone to false-positive results and inaccuracies, are time-consuming, and use complicated procedures to test large water samples. Herein, we show a simple detection and counting method for E. coli in the water samples involving a combination of DNAzyme sensor, microfluidics, and computer vision strategies. We first isolated E. coli into individual droplets containing a DNAzyme mixture using droplet microfluidics. Upon bacterial cell lysis by heating, the DNAzyme mixture reacted with a particular substrate present in the crude intracellular material (CIM) of E. coli. This event triggers the dissociation of the fluorophore-quencher pair present in the DNAzyme mixture leading to a fluorescence signal, indicating the presence of E. coli in the droplets. We developed an algorithm using computer vision to analyze the fluorescent droplets containing E. coli in the presence of non-fluorescent droplets. The algorithm can detect and count fluorescent droplets representing the number of E. coli present in the sample. Finally, we show that the developed method is highly specific to detect and count E. coli in the presence of other bacteria present in the water sample.

Automated summary

In this study, a simple and efficient method for detecting and counting E. coli in water samples was developed, which involved the use of a DNAzyme sensor, microfluidics, and computer vision strategies. The method successfully isolated E. coli into individual droplets, and the reaction of DNAzyme mixture with E. coli's intracellular material resulted in fluorescence signal, indicating the presence of E. coli. The computer vision algorithm accurately detected and counted the fluorescent droplets, providing a reliable measurement of E. coli in the sample.