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A One-Pot Convenient RPA-CRISPR-Based Assay for Salmonella enterica Serovar Indiana Detection
Jiansen Gong1,2, Di Zhang3, Lixia Fu4
1Poultry Institute, Chinese Academy of Agricultural Sciences, Yangzhou 225125, China.
Microorganisms
|March 28, 2024
Summary
A new one-step diagnostic test accurately detects Salmonella Indiana infection using recombinase polymerase amplification (RPA) and CRISPR/Cas12b technology. This rapid and sensitive assay offers a promising tool for identifying Salmonella Indiana in food safety applications.
Area of Science:
- Microbiology
- Molecular Diagnostics
- Food Safety
Background:
- Salmonella enterica serovar Indiana (S. Indiana) is a prevalent cause of foodborne illness globally.
- Accurate and rapid diagnostic methods are crucial for controlling S. Indiana outbreaks.
Purpose of the Study:
- To develop a novel, one-step identification method for S. Indiana using RPA combined with CRISPR/Cas12b.
- To evaluate the sensitivity, specificity, and applicability of the developed assay for S. Indiana detection.
Main Methods:
- A one-pot platform integrating recombinase polymerase amplification (RPA) with CRISPR/Cas12b was established.
- Optimal reaction conditions, including Cas12b and single-guide RNA (sgRNA) concentrations, were determined.
- The assay's performance was validated using various Salmonella strains, other pathogens, and chicken carcass samples.
Main Results:
- The RPA-CRISPR/Cas12b assay achieved a limit of detection of 14.4 copies per reaction.
- 100% accuracy was demonstrated for S. Indiana identification among Salmonella strains and other pathogens.
- The assay showed 100% agreement with traditional culture methods when applied to chicken carcass samples from different processing stages.
Conclusions:
- The developed one-step RPA-CRISPR/Cas12b assay is a sensitive, specific, and accurate tool for S. Indiana detection.
- This method offers a rapid and instrument-free diagnostic solution for S. Indiana infection.
- The assay holds significant potential for improving food safety and public health surveillance.
