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An Adapted Optical Density-Based Microplate Assay for Characterizing Actinobacteriophage Infection
Published on: June 30, 2023
Filter-based assay for Escherichia coli in aqueous samples using bacteriophage-based amplification.
Ratmir Derda1, Matthew R Lockett, Sindy K Y Tang
1Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138, United States.
Analytical Chemistry
|July 16, 2013
Summary
This study presents a novel bacteriophage (phage) based method for rapid bacterial detection in water. The technique amplifies bacterial signals for sensitive and accessible microbial testing.
Area of Science:
- Microbiology
- Biotechnology
- Environmental Science
Background:
- Bacterial detection in aqueous samples is crucial for public health.
- Existing methods for bacterial detection can be time-consuming and require laboratory equipment.
- Bacteriophages (phages) offer potential for sensitive and specific biological detection systems.
Purpose of the Study:
- To develop a rapid and accessible method for detecting bacteria in aqueous samples using bacteriophages.
- To demonstrate a phage-based amplification strategy for enhanced bacterial detection sensitivity.
- To evaluate the performance of the developed method in various sample types.
Main Methods:
- Bacteria were captured on a syringe filter.
- Target bacteria were infected with a specific bacteriophage (M13KE phage).
- The phage delivered a gene encoding a peptide motif, leading to the expression of the omega-domain of beta-galactosidase (β-gal) in Escherichia coli K12.
- Enzymatic activity of β-gal was detected colorimetrically for signal amplification.
Main Results:
- The phage-based assay detected as few as five colony-forming units (CFUs) of Escherichia coli K12 per liter of water using an overnight culture-based method.
- A solution-based assay with visual readout detected 50 CFUs of E. coli K12 in 1 L of water in under 4 hours.
- The solution-based assay demonstrated effectiveness in various matrices including orange juice and skim milk.
- The method showed potential for point-of-access testing without specialized laboratory equipment.
Conclusions:
- A bacteriophage-based amplification method enables sensitive and rapid detection of bacteria in aqueous samples.
- The developed solution-based assay is accessible, rapid, and suitable for point-of-access applications.
- This phage-based detection strategy can potentially be adapted for a wide range of bacterial targets by utilizing phages encoding different reporter genes.
