A colorimetric Loop-mediated isothermal amplification (LAMP) assay based on HRP-mimicking molecular beacon for the rapid detection of Vibrio parahaemolyticus.

Journal: Biosensors & bioelectronics
PMID: 31999578

Abstract

In the world wide, food poisoning accidents related to Vibrio spp. are on the rise, even numbers of food poisoning by other foodborne pathogens are decreasing. Therefore, the requirement of the rapid, sensitive and convenient detection method for V. parahaemolyticus has been grown. The objective of this study is to develop a colorimetric loop-mediated isothermal amplification (LAMP) assay using a molecular beacon (HRPzyme connected with complementary oligonucleotides at the 5' and 3' ends) for the rapid, sensitive, and convenient detection of V. parahaemolyticus. The colorimetric LAMP assay optimized at 58.8°C showed a detection limit of 1 × 10 CFU/mL and was confirmed to be specific to V. parahaemolyticus. The colorimetric LAMP assay can be finished within 1 h including DNA extraction step. The method was successfully applied to flatfish samples artificially inoculated with known amount of V. parahaemolyticus, and its cut-off value for the flatfish samples was 1 × 10 CFU/g. In addition, the colorimetric LAMP assay developed in the study was found to be able to correct false-positive results, which are known to be a disadvantage of conventional LAMP assays. Therefore, these results indicated that the colorimetric LAMP method is a useful tool for the rapid, sensitive and convenient detection of V. parahaemolyticus in fishes and can also be used as a point-of-care molecular diagnostic technique since it does not require any expensive equipment such as a thermocycler and detectors.

Authors

  • Jeong-Eun Lee
    Division of Applied Life Science, Graduate School, Gyeongsang National University, Jinju, Gyeongnam, 52828, South Korea.
  • Hyoyoung Mun
    Apteasy MJ inc., BI Center, 123 Cheomdangwagi-ro, Buk-gu, Gwangju, 61005, South Korea.
  • Se-Ri Kim
    Agro-Food Safety & Crop Protection Department, National Institution of Agricultural Science, Rural Development Administration, Wanju, 55365, South Korea.
  • Min-Gon Kim
    Department of Chemistry, School of Physics and Chemistry, Gwangju Institute of Science and Technology (GIST), Gwangju, 61005, South Korea.
  • Ji-Yoon Chang
    Advanced Process Technology and Fermentation Research Group, World Institute of Kimchi, Gwangju, 61755, South Korea.
  • Won-Bo Shim
    Institute of Agriculture and Life Science, Gyeongsang National University, Jinju, Gyeongnam, 52828, South Korea; Department of Agricultural Chemistry and Food Science & Technology, Gyeongsang National University, Jinju, Gyeongnam, 52828, South Korea. Electronic address: wbshim@gnu.ac.kr.

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