Selection and characterization of ssDNA aptamers specifically recognizing pathogenic Vibrio alginolyticus

作者：Qing Yu, Mingzhu Liu, Hongfei Su, Hehe Xiao, Siting Wu, Xianling Qin, Siqiao Li, Huizhi Mi, Zijun Lu, Deqiang Shi, Pengfei Li · 发表于：Journal of Fish Diseases · 年份：2019 · DOI：10.1111/jfd.12985 · 被引用次数：40 · 研究领域：Advanced biosensing and bioanalysis techniques、Aquaculture disease management and microbiota、RNA Interference and Gene Delivery

Vibrio alginolyticus (V. alginolyticus) is a major opportunistic pathogen to both marine animals and humans, which has also caused heavy economic losses to mariculture. The aim of this study was to develop highly specific aptamers for V. alginolyticus. Single-stranded DNA (ssDNA) aptamers with high binding affinity to viable V. alginolyticus were generated by Systematic Evolution of Ligands by Exponential Enrichment (SELEX) and identified by flow cytometric analysis in this study. The selected aptamers showed high specificity for V. alginolyticus and low apparent binding for other bacteria. The aptamers formed distinct stem-loop structures, which could form the basis of aptamers' specific binding to the target V. alginolyticus. Aptamer VA2 and VA8 showed particularly high binding affinity constant (Kd) of 14.31 ± 4.26 and 90.00 ± 13.51 nM, respectively. The aptamers produced no cytotoxic effects in vitro and in vivo. ssDNA aptamers were successfully selected against the viable bacteria pathogen V. alginolyticus by SELEX. The aptamers selected in this study could be not only applied as specific chemical molecular probes for studying V. alginolyticus pathogenesis to Trachinotus ovatus, but also developing rapid convenient diagnosis assay for V. alginolyticus infection, even when applied to the complex sample matrix, such as food and environment samples.