title: Rapid and sensitive detection of shrimp infectious myonecrosis virus using a reverse transcription loop-mediated isothermal amplification and visual colorogenic nanogold hybridization probe assay.
abstract: This study reports a novel strategy for the detection of reverse transcription loop-mediated isothermal amplification (RT-LAMP) products derived from infectious myonecrosis virus (IMNV), causes a serious myonecrosis in Penaeus (Litopenaeus) vannamei, by using a ssDNA-labeled with gold nanoparticle (AuNP) probe. This technique relies on a self-aggregation method, when the AuNP aggregation is induced by an increasing of salt concentrations with visual detection. The presence of IMNV-LAMP target prevented an AuNP aggregation and a solution remained as pink color of AuNP, while non-complementary targets cannot prevent AuNP aggregation, resulting in a visible color change to purple color after addition of salt. By using the combination of LAMP and AuNP probe system, the total assay interval required approximately 50 min (exclude RNA preparation). Detection limit was 10 copies of IMNV RNA in vitro transcript that comparable to that of LAMP followed by LFD and nested RT-PCR, but it was 100-times more sensitive than RT-PCR methods. This assay can be adapted easily for rapid detection of other shrimp infectious diseases agents at low-cost with robust reagents and using a simple colorimetric detection method.
authors.affiliation: Center of Excellence for Shrimp Molecular Biology and Biotechnology (CENTEX Shrimp), Faculty of Science, Mahidol University, Rama VI Road, Bangkok 10400, Thailand; National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency, Pathumthani 12120, Thailand.
authors.forename: Narong
authors.initials: N
authors.lastname: Arunrut
authors.forename: Jantana
authors.initials: J
authors.lastname: Kampeera
authors.forename: Rungkarn
authors.initials: R
authors.lastname: Suebsing
authors.forename: Wansika
authors.initials: W
authors.lastname: Kiatpathomchai
mesh_terms: D000818:Animals; D003124:Colorimetry / Q000379:methods*; D005516:Food Microbiology / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D033561:Penaeidae / Q000821:virology*; D012328:RNA Viruses / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013997:Time Factors
keywords: Colorimetric; Gold nanoparticle; Infectious myonecrosis virus; Penaied shrimp; Reverse transcription loop-mediated isothermal amplification