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Real-time fluorescence loop-mediated isothermal amplification assay for rapid and sensitive detection of Streptococcus gallolyticus subsp. gallolyticus associated with colorectal cancer

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Abstract

The increasing threat of Streptococcus gallolyticus subsp. gallolyticus (SGG) infections has gained considerable attention for its strong association with colorectal cancer (CRC). Herein, we proposed real-time fluorescence loop-mediated isothermal amplification (LAMP) as a novel, simple, rapid, and highly sensitive assay for identifying SGG for the first time. This assay was capable of detecting SGG with initial DNA concentrations ranging from 102 to 108 copies per microliter, under isothermal conditions within 30 min via real-time fluorescence monitoring. Our method was tested for specific identification of SGG strains without cross-reaction with other Streptococcus gallolyticus subspecies and Escherichia coli. The developed LAMP shows a superior performance with shorter time and higher sensitivity compared with conventional polymerase chain reaction (PCR). Significantly, this proposed approach was successfully applied for detecting SGG in clinical urine samples, which is non-invasive diagnosis, showing excellent accuracy and reliability to discriminate healthy controls and CRC patients. For comparison, these samples were also tested against PCR assay. These results yielded an analytical sensitivity of 100% and a specificity of 100% for SGG testing using LAMP. The findings suggest LAMP can be employed for detecting SGG infections which is useful for diagnosis and screening of CRC.

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Acknowledgments

The authors gratefully acknowledge the Department of Clinical Laboratory, Yangpu Hospital (Shanghai, China), for the help in collecting clinical urine samples.

Funding

This work was funded by the National Natural Science Foundation of China (21375027, 21335002, 21427806), Shanghai Pujiang Program (17PJD001, 18PJD047), and Natural Science Foundation of Shanghai (12ZR1401700, 17JC1400100).

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Authors and Affiliations

  1. Department of Chemistry, Fudan University, Shanghai, 200438, China

    Qiuyuan Lin, Xin Ye, Bin Yang, Xueen Fang, Hui Chen & Jilie Kong

  2. Department of Clinical Laboratory, Yangpu Hospital, Tongji University School of Medicine, Shanghai, 200090, China

    Wenhao Weng

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  1. Qiuyuan Lin
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  2. Xin Ye
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  3. Bin Yang
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  5. Hui Chen
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  7. Jilie Kong
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Corresponding authors

Correspondence to Hui Chen, Wenhao Weng or Jilie Kong.

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All participants in this study provided written informed consent prior to testing. The study design was approved by the institutional research commission of Fudan University (Shanghai, China). Ethical approval was obtained from the Ethics Committee of the Department of Clinical Laboratory, Yangpu Hospital (Shanghai, China).

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Lin, Q., Ye, X., Yang, B. et al. Real-time fluorescence loop-mediated isothermal amplification assay for rapid and sensitive detection of Streptococcus gallolyticus subsp. gallolyticus associated with colorectal cancer. Anal Bioanal Chem 411, 6877–6887 (2019). https://doi.org/10.1007/s00216-019-02059-8

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