A Loop-Mediated Isothermal Amplification Integrated G-Quadruplex Molecular Beacon (LAMP-GMB) Method for the Detection of Staphylococcus aureus in Food
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Staphylococcus aureus is a common pathogen in foodstuffs, and an effective detection assay is therefore essential for ensuring food safety. In this study, we established a label-free loop-mediated isothermal amplification integrated G-quadruplex molecular beacon (LAMP-GMB) chemiluminescence method for S. aureus detection based on its nuc gene that encodes a thermostable nuclease. Target DNA was amplified using the LAMP technique; the result LAMP products were specially recognized by GMB detection probe, subsequently induced the release of G-rich sequence, and formed a complex spatial structure G-quadruplex. The structure can bind with hemin and catalyze the H2O2-mediated oxidation of luminol reagent to produce the chemiluminescence signal for chemiluminescent detection. Under the optimized conditions, the developed method showed a good linear relationship (R2 = 0.9882) in the range of 101-105 fg μL−1 and was able to detect as low as 10 fg μL−1 genomic DNA from S. aureus. Using spiked milk samples, the method exhibited a linear relationship (R2 = 0.9893) ranging from 5 × 101 to 5 × 107 CFU mL−1 with the limit of detection of 50 CFU mL−1. Importantly, the detection assay did not require the preparation and use of labeled primers for the LAMP reaction, increasing the reliability and efficiency of the assay. Therefore, the developed LAMP-GMB method has the potential to be widely applied to S. aureus monitoring work in food field.
KeywordsStaphylococcus aureus LAMP G-quadruplex Luminol Chemiluminescence
This study was funded by the Fund for Qing Lan Project of Jiangsu, the Fundamental Research Funds for the Central Universities (KYYJ201708), and Special Funds of Agro-Product Quality Safety Risk Assessment of Ministry of Agriculture of the People’s Republic of China (GJFP201701505).
Compliance with Ethical Standards
Conflict of Interest
Jingguo Xu declares that he has no conflict of interest. Yimin Hu declares that she has no conflict of interest. Jia Guo declares that she has no conflict of interest. Yumeng Yang declares that she has no conflict of interest. Jiarong Qiu declares that she has no conflict of interest. Xuanxuan Li declares that he has no conflict of interest. Zhihong Xin declares that he has no conflict of interest
This article does not contain any studies with human participants or animals performed by any of the authors.
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