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Title: Rapid and sensitive detection of Vibrio vulnificus by loop-mediated isothermal amplification combined with lateral flow dipstick targeted to rpoS gene
Authors: Surasilp T.
Longyant S.
Rukpratanporn S.
Sridulyakul P.
Sithigorngul P.
Chaivisuthangkura P.
Keywords: bacterial DNA
fluorescein isothiocyanate
RNA polymerase
RNA polymerase subunit sigma factor s
unclassified drug
bacterial gene
bacterium culture
bacterium detection
bacterium isolate
gene targeting
Listonella anguillarum
loop mediated isothermal amplification
priority journal
RNA hybridization
Vibrio alginolyticus
vibrio campbellii
Vibrio cholerae
Vibrio fluvialis
Vibrio harveyi
Vibrio mimicus
vibrio ordalii
Vibrio parahaemolyticus
Vibrio shilonii
Vibrio vulnificus
Bacterial Proteins
DNA Primers
Molecular Probes
Molecular Sequence Data
Nucleic Acid Amplification Techniques
Sigma Factor
Vibrio vulnificus
Vibrio vulnificus
Issue Date: 2011
Abstract: A novel loop-mediated isothermal amplification (LAMP) combined with amplicon detection by chromatographic lateral flow dipstick (LFD) assay was developed and evaluated for the detection of Vibrio vulnificus. Biotinylated LAMP amplicons were produced by a set of six designed primers that recognized the V. vulnificus RNA polymerase subunit sigma factor S (rpoS) gene followed by hybridization with an FITC-labeled probe and LFD detection. The optimized time and temperature conditions for the LAMP assay were 90 min at 65 °C. The LAMP-LFD method accurately identified 14 isolates of V. vulnificus but did not detect 25 non-vulnificus Vibrio isolates and 37 non-Vibrio isolates. The sensitivity of LAMP-LFD for V. vulnificus detection in pure culture was 1.5 × 103 CFU ml-1 or equivalent to 2.8 CFU per reaction. In the case of spiked oyster samples without enrichment, the detection limit for V. vulnificus was 1.2 × 104 CFU g-1 or equivalent to 11 CFU per reaction. The results show that this method appears to be accurate, precise and valuable tool for identification of V. vulnificus and can be used efficiently for detection of V. vulnificus in contaminated food sample. © 2011 Elsevier Ltd.
ISSN: 8908508
Appears in Collections:SCOPUS 1983-2021

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