Rapid detection of Escherichia coli in beverages using genetically engineered bacteriophage T7

Nicharee Wisuthiphaet,Nitin Nitin,Glenn M Young,Xu Yang

doi:10.1186/s13568-019-0776-7

Nicharee Wisuthiphaet, Nitin Nitin + Show 2 more

Open Access

https://doi.org/10.1186/s13568-019-0776-7

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Journal: AMB Express	Publication Date: Apr 19, 2019
Citations: 46	License type: open-access

Affiliation: University of California, Davis

Abstract

Foodborne illness due to bacterial contamination is a significant issue impacting public health that demands new technology which is practical to implement by food industry. Detection of bacteria in food products and production facilities is a crucial strategy supporting food safety assessments. Bacteriophages were investigated as a tool for bacterial detection due to their ability to infect specific strain of host bacteria in order to improve sensitivity, specificity, and rapidity of bacterial detection. The results of this investigation reveal a novel method for rapid detection. The method employs a genetically engineered bacteriophage, phage T7-ALP, which expresses alkaline phosphatase. Upon infection of Escherichia coli, overexpression of alkaline phosphatase provides an opportunity for rapid sensitive detection of a signal indicative of bacterial presence in model beverage samples as low as 100 bacteria per gram. The method employs a fluorescent precipitated substrate, ELF-97, as a substrate for alkaline phosphatase activity coupled with fluorescence imaging and image analysis allowing single-cell imaging results in high detection sensitivity. The method is easily completed within less than 6 h enabling it to be deployed within most large industrial food processing facilities that have routine 8-h operational shifts.

Highlights

Over the past decades, foodborne disease outbreaks affect millions of people worldwide by causing burdens on public health and significant hindrance in socioeconomic development (WHO 2015)
Several bacteriophagebased bacteria detection approaches have been recently implemented in order to improve bacteria detection sensitivity (Table 1)
These previous studies developed the detection methods based on colorimetric, bioluminescent, and electrochemical detection of enzymatic activity using substrates that yield soluble products; their detection sensitivity relies on the cell concentration and/or immobilization of the reporter enzyme on biomaterial substrates

Summary

Introduction

Foodborne disease outbreaks affect millions of people worldwide by causing burdens on public health and significant hindrance in socioeconomic development (WHO 2015). Immunological-based methods, including variations of enzyme-linked immunosorbent assays (ELISAs), have been developed in order to provide more rapid, economical and simple approach compared to the culture-based and PCR-based methods; on a practical level, ELISAs suffer from an inability to distinguish live from dead bacteria, from low sensitivity due to small sample volume capacity, and from low specificity due to cross-reactivity of polyclonal antibodies (Ivnitski et al 1999; Shen et al 2014) The limitations of these foodborne pathogen detection methods provide incentive for continued research and development into innovative novel approaches for rapid detection that have high sensitivity and specificity to detect bacterial foodborne pathogens, and can be widely adapted for use by a diversity of large and small food manufactures

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