College of Marine Sciences, South China Agricultural University, Guangdong Laboratory for Lingnan Modern Agriculture, Guangzhou 510642, China.
Laboratory for Marine Biology and Biotechnology, Qingdao National Laboratory for Marine Science and Technology, Qingdao 266000, China.
Viruses. 2022 Apr 30;14(5):945. doi: 10.3390/v14050945.
Largemouth bass virus (LMBV) is a major viral pathogen in largemouth bass culture, usually causing high mortality and heavy economic losses. Accurate and early detection of LMBV is crucial for diagnosis and control of the diseases caused by LMBV. Previously, we selected the specific aptamers, LA38 and LA13, targeting LMBV by systematic evolution of ligands by exponential enrichment (SELEX). In this study, we further generated truncated LA38 and LA13 (named as LA38s and LA13s) with high specificity and affinities and developed an aptamer-based sandwich enzyme-linked apta-sorbent assay (ELASA) for LMBV diagnosis. The sandwich ELASA showed high specificity and sensitivity for the LMBV detection, without cross reaction with other viruses. The detection limit of the ELASA was as low as 1.25 × 10 LMBV-infected cells, and the incubation time of the lysate and biotin labeled aptamer was as short as 10 min. The ELASA could still detect LMBV infection in spleen lysates at dilutions of 1/25, with good consistency of qRT-PCR. For the fish samples collected from the field, the sensitivity of ELASA was 13.3% less than PCR, but the ELASA was much more convenient and less time consuming. The procedure of ELASA mainly requires washing and incubation, with completion in approximately 4 h. The sandwich ELASA offers a useful tool to rapidly detect LMBV rapidly, contributing to control and prevention of LMBV infection.
大口黑鲈病毒（LMBV）是大口黑鲈养殖中的一种主要病毒病原体，通常导致高死亡率和严重的经济损失。准确和早期检测 LMBV 对于诊断和控制由 LMBV 引起的疾病至关重要。以前，我们通过指数富集的配体系统进化（SELEX）选择了针对 LMBV 的特异性适体 LA38 和 LA13。在这项研究中，我们进一步生成了具有高特异性和亲和力的截短的 LA38 和 LA13（命名为 LA38s 和 LA13s），并开发了基于适体的夹心酶联吸附测定法（ELASA）用于 LMBV 诊断。夹心 ELASA 对 LMBV 的检测具有高度特异性和灵敏度，与其他病毒没有交叉反应。ELASA 的检测限低至 1.25×10 LMBV 感染细胞，裂解物和生物素标记适体的孵育时间短至 10 分钟。ELASA 仍可在脾脏裂解物稀释度为 1/25 时检测到 LMBV 感染，与 qRT-PCR 的一致性良好。对于从现场采集的鱼类样本，ELASA 的灵敏度比 PCR 低 13.3%，但 ELASA 更方便，耗时更少。ELASA 的程序主要需要洗涤和孵育，大约 4 小时即可完成。夹心 ELASA 为快速检测 LMBV 提供了有用的工具，有助于控制和预防 LMBV 感染。
