Abstract
Rapid detection of pathogenic bacteria is extremely important for public health and safety. Here, we describe for the first time an integrated origami paper-based analytical device (PAD) incorporating cell lysis, molecular recognition, amplification and visual detection of Escherichia coli (E. coli). The device features three components: paper for its ability to extract protein molecules nonspecifically from cells, DNA superstructures for their ability to immobilize RNA-cleaving DNAzymes (RCDs) but undergo target-induced RNA cleavage on paper, and isothermal rolling circle amplification (RCA) for its ability to amplify each cleavage event into repetitive sequence units that can be detected by naked eye. This device can achieve detection of E. coli K12 with a detection limit of as low as 103 CFU·mL−1 in a total turnaround time of 35 min. Furthermore, this device allowed the sensitive detection of E. coli in complex sample matrices such as juice and milk. Given that more specific RCDs can be evolved for diverse bacteria, the integrated PAD holds great potential for rapid, sensitive and highly selective detection of pathogenic bacteria in resource-limited settings.
Highlights
In recent years, infectious diseases caused by food- and water-borne bacterial pathogens have a high incidence throughout the world [1,2,3,4]
We developed a disposable origami paper-based analytical device that fully integrates on-paper cell lysis for protein extraction, target-responsive RNA-cleaving DNAzymes (RCDs) for molecular recognition, rolling circle amplification (RCA) for signal amplification, origami technique for reagent diffusion and flow, and portable readout
The working principle and key functionalities of the oPADs are illustrated in Scheme 1
Summary
Infectious diseases caused by food- and water-borne bacterial pathogens have a high incidence throughout the world [1,2,3,4]. According to the World Health Organization, diarrhea pathogens, such as Escherichia coli, Salmonella, Helicobacter pylori, Campylobacter and Vibrio cholerae [5,6,7,8], account for about one-third infections. Among these pathogenic bacteria, the main cause of diarrhea is Escherichia coli (E. coli) [9]. E. coli is a widely distributed Gram-negative bacterium that mainly lives in the intestines of humans and warm-blooded animals for facultative anaerobic activities It can cause serious symptoms such as sepsis and hemolytic uremic syndrome [10,11], and even result in serious diseases including diarrhea, gastroenteritis, inflammation and malnutrition. Since E. coli can cause pathogenicity at low doses (most likely less than 100 organisms) [12], the development of a rapid, sensitive, low-cost and highly specific method for identifying E. coli is of great significance in the fields of medical diagnostics, food safety, and environmental monitoring
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