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Loop-mediated isothermal amplification combined with colorimetric nanogold for detection of the microsporidian Enterocytozoon hepatopenaei in penaeid shrimp.
MedLine Citation:
PMID:  23387348     Owner:  NLM     Status:  Publisher    
Abstract/OtherAbstract:
AIMS: Enterocytozoon hepatopenaei is an emerging microsporidian parasite that has been linked to recent losses caused by white feces syndrome (WFS) in cultivated giant or black tiger shrimp Penaeus (Penaeus) monodon and whiteleg shrimp Penaeus (Litopenaeus) vannamei in Asia. To more accurately assess its impact on shrimp production and to determine reservoir carriers for control measures, our objective was to establish a loop-mediated isothermal amplification (LAMP) assay combined with colorimetric nanogold (AuNP) for rapid, sensitive and inexpensive detection of this parasite. METHODS AND RESULTS: A set of six specific primers was designed to successfully detect the SSU rRNA gene of E. hepatopenaei by a LAMP reaction of 45 min at 65°C combined with visual detection of the amplification product via hybridization at 65°C for 5 min with a ssDNA-labled nanogold probe, followed by salt induced AuNP aggregation (total assay time approximately 50 minutes). This method gave similar results to LAMP followed by electrophoresis or spectrophotometric detection, and it was more sensitive (0.02 fg total DNA) than a conventional nested PCR reaction (0.2 fg total DNA). The new method gave negative results with shrimp DNA templates extracted from diseased shrimp containing other pathogens, indicating that the LAMP-AuNP assay was specific for E. hepatopenaei. CONCLUSIONS: Without sacrificing sensitivity or specificity, the new LAMP-AuNP assay significantly reduced the time, ease and cost for molecular detection of E. hepatopenaei in shrimp. © 2013 The Authors Journal of Applied Microbiology © 2013 The Society for Applied Microbiology.
Authors:
Rungkarn Suebsing; Photchanathorn Prombun; Jiraporn Srisala; Wansika Kiatpathomchai
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Publication Detail:
Type:  JOURNAL ARTICLE     Date:  2013-2-7
Journal Detail:
Title:  Journal of applied microbiology     Volume:  -     ISSN:  1365-2672     ISO Abbreviation:  J. Appl. Microbiol.     Publication Date:  2013 Feb 
Date Detail:
Created Date:  2013-2-7     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9706280     Medline TA:  J Appl Microbiol     Country:  -    
Other Details:
Languages:  ENG     Pagination:  -     Citation Subset:  -    
Copyright Information:
© 2013 The Authors Journal of Applied Microbiology © 2013 The Society for Applied Microbiology.
Affiliation:
Center of Excellence for Shrimp Molecular Biology and Biotechnology (CENTEX Shrimp), Faculty of Science, Mahidol University, Rama VI rd., Bangkok, 10400, Thailand; National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency, Pathumthani, 12120, Thailand.
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