Digital metabolic activity assay enables fast assessment of 2D materials bactericidal efficiency.

Background: The identification and quantification of viable Escherichia coli (E. coli) are important in multiple fields including the development of antimicrobial materials, water quality, food safety and infections diagnosis. However, the standard culture-based methods of viable E. coli detection suffer from long detection times (24 h) and complex operation, leaving the unmet requirement for fast assessing the efficiency of antimicrobial materials, early alerting the contamination of water and food, and immediately treatment of infections.

Results: We present a digital β-d-glucuronidase (GUS) assay in a self-priming polydimethylsiloxane (PDMS) microfluidic chip for rapid E. coli identification and quantification. The GUS expression in viable bacteria was investigated to develop a fast GUS assay at the single-cell level. Single E. coli were stochastically discretized in picoliter chambers and identified by specific GUS activity. The digital GUS assay enabled identifying E. coli within 3 h and quantifying within 4 h for different E. coli subtypes. The specificity of our method was confirmed by using blended bacteria including E. coli, Bacillus, Shigella and Vibrio. We utilized digital GUS assay to enumerate viable E. coli after incubated with antibacterial materials for assessing the antibacterial efficiency. Moreover, the degassed chip can realize automatic sample distribution without external instruments.

Significance: The results demonstrated the functionality and practicability of digital GUS assay for single E. coli identification and quantification. With air-tight packaging, the developed chip has the potential for on-site E. coli analysis and could be deployed for diagnosis of E. coli infections, antimicrobial susceptibility testing, and warning the fecal pollution of water. Digital GUS assay provides a paradigm, examining the activity of metabolic enzyme, for detecting the viable bacteria other than E. coli.