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Background <p>Enterovirus G (EV-G) is widely prevalent in pigs worldwide, and co-infections of EV-G and other diarrhea-causing pathogens have been reported in many countries, threatening the pig farming industry. There are some methods available for EV-G detection; however, the RT-LAMP method for detecting EV-G has not yet been developed. The aim of thisstudy was to establish a highly sensitive and visual RT-LAMP assay for the detection of EV-G.</p> Results <p>A colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed and evaluated for the visual detection of porcine enterovirus <i>G</i> (EV-G). The assay can be completed in 45&#xa0;min at 64&#xa0;°C, and the results can be observed by the unaided eye using the colorimetric LAMP master mix. The limit of detection (LOD) of the RT-LAMP assay was 46.8 copies of EV-G RNA, which is comparable to that of the previously described real-time RT-PCR (qRT-PCR) method and is 100-fold more sensitive than that of conventional RT-PCR. The assay specifically amplified the RNA of EV-Gs, and there was no cross-amplification with other porcine pathogens. In the clinical evaluation, the Kappa coefficient of the established RT-LAMP assay and the previously reported qRT-PCR was was 0.862 (<i>P</i> &lt; 0.01), and the nucleic acid of EV-G could be tested from the fecal samples of porcine at different infection stages.</p> Conclusions <p>In this study, we successfully established a visual, sensitive, specific, rapid, and convenient RT-LAMP method for the detection of EV-G, which was successfully applied in the detection of EV-Gs in clinical samples.</p>

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Colorimetric reverse transcription loop-mediated isothermal amplification assay for visual, sensitive, and specific detection of enterovirus G

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年8月29日

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  • Zhan-hong Li,
  • Zhen-xing Zhang,
  • Pei Zhu,
  • Zhuo-ran Li,
  • Peng Liu,
  • Qi Yang,
  • Qiu-yan Yang,
  • Zhen Yang,
  • Jian-ling Song

摘要

Background

Enterovirus G (EV-G) is widely prevalent in pigs worldwide, and co-infections of EV-G and other diarrhea-causing pathogens have been reported in many countries, threatening the pig farming industry. There are some methods available for EV-G detection; however, the RT-LAMP method for detecting EV-G has not yet been developed. The aim of thisstudy was to establish a highly sensitive and visual RT-LAMP assay for the detection of EV-G.

Results

A colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed and evaluated for the visual detection of porcine enterovirus G (EV-G). The assay can be completed in 45 min at 64 °C, and the results can be observed by the unaided eye using the colorimetric LAMP master mix. The limit of detection (LOD) of the RT-LAMP assay was 46.8 copies of EV-G RNA, which is comparable to that of the previously described real-time RT-PCR (qRT-PCR) method and is 100-fold more sensitive than that of conventional RT-PCR. The assay specifically amplified the RNA of EV-Gs, and there was no cross-amplification with other porcine pathogens. In the clinical evaluation, the Kappa coefficient of the established RT-LAMP assay and the previously reported qRT-PCR was was 0.862 (P < 0.01), and the nucleic acid of EV-G could be tested from the fecal samples of porcine at different infection stages.

Conclusions

In this study, we successfully established a visual, sensitive, specific, rapid, and convenient RT-LAMP method for the detection of EV-G, which was successfully applied in the detection of EV-Gs in clinical samples.