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Rapid and simple detection of Pediococcus using ARMS-CRISPR/Cas12a method.

Xiaoyan Li¹, Haiqiang Lu¹, Shaozeng Li²

¹College of Food Science and Technology, Hebei Agricultural University, Baoding, 071001, People's Republic of China.

|April 19, 2026

Summary

A new detection system using amplification-resistant mutation system-based polymerase chain reaction (ARMS-CRISPR/Cas12a) accurately identifies Pediococcus spp. This rapid method offers high sensitivity and specificity for detecting these bacteria in various applications.

Keywords:

ARMS-PCR CRISPR/Cas12a Pediococcus spp.SNP

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Area of Science:

Microbiology
Molecular Biology
Biotechnology

Background:

Pediococcus spp. are common lactic acid bacteria found in diverse environments, including the intestinal microbiota of humans and animals.
Accurate and sensitive detection methods for Pediococcus spp. are crucial but currently face significant challenges.
Distinguishing Pediococcus spp. from other bacterial genera requires highly specific diagnostic tools.

Purpose of the Study:

To develop a novel, highly sensitive, and specific detection system for Pediococcus spp.
To identify single nucleotide polymorphism (SNP) sites within the 16S rRNA gene for precise genus identification.
To establish a rapid and effective diagnostic tool for Pediococcus spp. relevant to food production.

Main Methods:

Development of a fluorescent detection system utilizing amplification-resistant mutation system-based polymerase chain reaction (ARMS-PCR) combined with CRISPR/Cas12a.
Bioinformatic analysis of Pediococcus spp. 16S rRNA gene sequences to identify specific SNP sites.
Design of specific primers targeting Pediococcus pentosaceus STS-6 for optimized detection.

Main Results:

The ARMS-CRISPR/Cas12a system demonstrated high specificity and sensitivity in detecting Pediococcus spp. under optimized conditions (Cas12a:crRNA ratio of 1:1 at 37°C).
The limit of detection for genomic DNA was 8.15 × 10⁻⁵ ng/μL, surpassing the sensitivity of traditional gel electrophoresis.
The entire detection process was completed in approximately 1.5 hours, indicating a rapid diagnostic capability.

Conclusions:

The established ARMS-CRISPR/Cas12a detection system provides a rapid, sensitive, and specific method for identifying Pediococcus spp.
This system effectively detects the 16S rRNA gene of clinically relevant Pediococcus spp. probiotics.
The developed method meets the stringent requirements for bacterial detection in food production settings.