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Multiplex Detection of Bacteria in Complex Clinical and Environmental Samples using Oligonucleotide-coupled Fluorescent Microspheres

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Published on: October 23, 2011

Multispectral pathogens detection in food using multiplex hyperbranched saltatory rolling circle amplification.

Nanfack V D Carole¹, Lina Sheng¹, Jian Ji¹

¹School of Food Science and Technology, International Joint Laboratory on Food Safety, Synergetic Innovation Center of Food Safety and Quality Control, Jiangnan University, Wuxi, Jiangsu, 214122, PR China.

|August 8, 2024

Summary

A new triple-probe multiplex rolling circle amplification method offers rapid and accurate detection of Salmonella Typhimurium and Staphylococcus aureus. This technique provides a simple visual identification for foodborne pathogen testing.

Keywords:

Food safety Isothermal nucleic acid amplification Pathogen detection Rolling circle amplification

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Area of Science:

Food safety
Microbiology
Molecular diagnostics

Background:

Foodborne illnesses caused by Salmonella and Staphylococcus aureus pose significant public health risks.
Current detection methods can be complex and time-consuming.
A need exists for simple, rapid, and accurate bacterial detection techniques.

Purpose of the Study:

To develop a straightforward method for simultaneous detection and identification of Salmonella Typhimurium and Staphylococcus aureus.
To utilize a triple-probe multiplex rolling circle amplification technique for enhanced sensitivity and specificity.

Main Methods:

A triple-probe multiplex rolling circle amplification assay was designed targeting specific genes (invA for S. Typhimurium, glnA for S. aureus).
Fluorophore-labeled probes and a pH-based detection approach were employed for visual identification.
The assay was optimized for speed and sensitivity, tested with synthetic targets and real bacteria, and evaluated in milk samples.

Main Results:

The assay achieved simultaneous detection of S. Typhimurium and S. aureus within 30 minutes for synthetic targets and 2 hours for real bacteria.
High specificity was observed, with limits of detection as low as 10 CFU/mL (S. Typhimurium) and 5 CFU/mL (S. aureus) in milk samples.
The detection method demonstrated stability for over 3 months.

Conclusions:

The developed triple-probe multiplex rolling circle amplification method is specific, sensitive, and easy to interpret.
This technique offers a promising solution for rapid and reliable detection of foodborne pathogens in food samples.
The method contributes to improving food safety and mitigating the impact of foodborne illnesses.