Naked-eyes detection of Largemouth bass ranavirus in clinical fish samples using gold nanoparticles as colorimetric sensor

Abstract

Largemouth bass virus (LMBV) infection causes a lethal disease (ulcerative syndrome) in largemouth bass (Micropterus salmoides), which is an economically important freshwater fish species cultured in south-east China. For rapid detection of this virus, a simple and sensitive method was developed using probe-modified gold nanoparticles (AuNPs) as a colorimetric biosensor. A species-specific probe was attached to the surface of AuNPs, and thereby increasing the resistance of AuNPs to NaCl-induced aggregation. The NaCl concentration and ratio of probe to AuNPs in reaction mixture were evaluated, and the assay was optimized to detect only LMBV DNA sequence. Under optimum conditions, a linear relationship was observed between the ratio of A670/A520 and LMBV DNA concentrations over the range from 2.33 to 23.3 ng/μL and as low as 6.99 ng/μL of unamplied virus DNA could be detected only utilizing naked eyes. The entire assay was finished within 30 min, and the detection results were comparable to PCR examination in terms of reliability. Therefore, the visual assay is a potentially useful tool for aquaculture farmers to screen LMBV, and an ideal candidate for prevention and control of ulcerative syndrome in fish.