Location: Harry K. Dupree Stuttgart National Aquaculture Research CntrTitle: Molecular detection and quantification of the fish pathogen Saprolegnia using Loop Mediated Isothermal Amplification (LAMP)
|GHOSH, SATYAKI - Bowling Green State University|
|Straus, David - Dave|
|GOOD, CHRISTOPHER - Freshwater Institute|
|PHUNTUMART, VIPAPORN - Bowling Green State University|
Submitted to: International Symposium on Aquatic Animal Health
Publication Type: Abstract Only
Publication Acceptance Date: 7/16/2018
Publication Date: 9/2/2018
Citation: Ghosh, S., Straus, D.L., Good, C., Phuntumart, V. 2018. Molecular detection and quantification of the fish pathogen Saprolegnia using Loop Mediated Isothermal Amplification (LAMP) [abstract]. International Symposium on Aquatic Animal Health. p. 422.
Technical Abstract: Saprolegniasis is one of the most serious diseases of fish. This is caused by an oomycete pathogen, Saprolegnia spp., and has resulted in significant losses to fish in both natural ecosystems and commercial setups. The goal of this study is to establish rapid and sensitive molecular tools for the on-field detection and quantification of this pathogen from water samples. Here, we have developed a Loop Mediated Isothermal Amplification (LAMP) assay to detect zoospores, the predominant infective propagule of the pathogen, by targeting the Internal Transcribed Spacer (ITS) region of the ribosomal DNA. Visual detection of the amplified products was possible using SYBR Green I dye. Using purified Saprolegnia spp. DNA, our developed LAMP protocol enables femtogram-level detection sensitivity. We also developed a qPCR protocol based on the same ITS marker that could detect Saprolegnia at the picogram level. Furthermore, this qPCR method could also detect one zoospore directly from water samples. Combining qPCR and LAMP would facilitate the detection and quantification of pathogen loads. The currently established protocol is being tested for on-field detection of the pathogen from water samples collected from Recirculating Aquaculture Systems (RAS).