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Simultaneous detection of multiple bacterial and viral aquatic pathogens using a fluorogenic loop-mediated isothermal

Qian-Jin Zhou1,2, Jian-Fei Lu2, Xiu-Rong Su1,2

  • 1State Key Laboratory for Managing Biotic and Chemical Threats to the Quality and Safety of Agro-products, Ningbo University, Ningbo, China.

Journal of Fish Diseases
|December 19, 2020
PubMed
Summary

A new dual-sample microfluidic chip with loop-mediated isothermal amplification (LAMP) enables rapid, simultaneous detection of 10 aquatic pathogens. This user-friendly assay offers quick, reliable on-site screening for aquaculture disease management.

Keywords:
aquatic pathogendual-sample microfluidic chiploop-mediated isothermal amplificationmultiplex detectionreal-time detection

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Area of Science:

  • Aquatic microbiology
  • Biotechnology
  • Diagnostic assay development

Background:

  • Rapid and user-friendly diagnostic tests are crucial for early disease detection in aquaculture.
  • On-site screening of pathogenic microorganisms is essential for preventing disease outbreaks.

Purpose of the Study:

  • To develop a dual-sample microfluidic chip integrated with real-time fluorogenic loop-mediated isothermal amplification (LAMP) assay.
  • To simultaneously detect 10 common pathogenic microorganisms in aquaculture.

Main Methods:

  • Development of a dual-sample on-chip LAMP assay for simultaneous analysis of two samples.
  • Detection of 10 pathogenic microorganisms including bacteria (Aeromonas hydrophila, Edwardsiella tarda, Vibrio spp.) and viruses (IHHNV, ISKNV, WSSV).
  • Real-time fluorogenic detection with automated protocol and short run times (<30 min).

Main Results:

  • Achieved limits of detection (LOD) from 10^0 to 10^-1 pg/μl for bacterial DNA and 10^-4 to 10^-5 pg/μl for viral DNA.
  • Demonstrated robust reproducibility with a coefficient of variation <10% for time-to-positive.
  • Reported clinical sensitivity of 93.52% and specificity of 85.53% compared to conventional methods.

Conclusions:

  • The on-chip LAMP assay is an effective tool for dual-sample, multiple pathogen analysis.
  • This assay is suitable for on-site detection and routine monitoring of multiple pathogens in aquaculture.
  • The developed technology facilitates rapid and user-friendly disease screening in aquaculture settings.