Isolation and characterization of a new class of DNA aptamers specific binding to Singapore grouper iridovirus (SGIV) with antiviral activities.
The Singapore grouper iridovirus (SGIV), a member of the genus Ranavirus, is a major viral pathogen that has caused heavy economic losses to the grouper aquaculture industry in China and Southeast Asia. No efficient method of controlling SGIV outbreaks is currently available. Systematic evolution of ligands by exponential enrichment (SELEX) is now widely used for the in vitro selection of artificial ssDNA or RNA ligands, known as aptamers, which bind to targets through their stable three-dimensional structures. In our current study, we generated ssDNA aptamers against the SGIV, and evaluated their ability to block SGIV infection in cultured fish cells and cultured fish in vivo. The anti-SGIV DNA aptamers, LMB-761, LMB-764, LMB-748, LMB-439, LMB-755, and LMB-767, were selected from a pool of oligonucleotides randomly generated using a SELEX iterative method. The analysis of the secondary structure of the aptamers revealed that they all formed similar stem-loop structures. Electrophoretic mobility shift assays showed that the aptamers bound SGIV specifically, as evidenced by a lack cross-reactivity with the soft shell turtle iridovirus. The aptamers produced no cytotoxic effects in cultured grouper spleen cells (GS). Assessment of cytopathic effects (CPE) and viral titer assays showed that LMB-761, LMB-764, LMB-748, LMB-755, and LMB-767 significantly inhibited SGIV infection in GS cells. The in vivo experiments showed that LMB-761 and LMB-764 reduced SGIV-related mortality, and no negative effects were observed in orange-spotted grouper, Epinephelus coioides, indicating that these DNA aptamers may be suitable antiviral candidates for controlling SGIV infections in fish reared in marine aquaculture facilities.
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China; University of Chinese Academy of Sciences, 19 Yuquan Road, Beijing 100049, China.
Pengfei
P
Li
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Yang
Y
Yan
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Shina
S
Wei
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Jingguang
J
Wei
State Key Laboratory Breeding Base for Sustainable Exploitation of Tropical Biotic Resources, College of Marine Science, Hainan University, Haikou 570228, China.
Ren
R
Gao
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Xiaohong
X
Huang
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China.
Youhua
Y
Huang
Analytical and Testing Center, Beijing Normal University, Xinjiekouwai Street, Beijing 100875, China. Electronic address: jgh982@bnu.edu.cn.
Guohua
G
Jiang
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, 164 West Xingang Road, Guangzhou 510301, China. Electronic address: qinqw@scsio.ac.cn.
Qiwei
Q
Qin
D000818:Animals; D000998:Antiviral Agents / Q000302:isolation & purification* / Q000494:pharmacology* / Q000627:therapeutic use; D052157:Aptamers, Nucleotide / Q000302:isolation & purification* / Q000494:pharmacology* / Q000627:therapeutic use; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002681:China; D003588:Cytopathogenic Effect, Viral / Q000187:drug effects; D024202:Electrophoretic Mobility Shift Assay; D005393:Fish Diseases / Q000188:drug therapy; D005399:Fishes; D009690:Nucleic Acid Conformation; D017995:Ranavirus / Q000187:drug effects* / Q000235:genetics; D052156:SELEX Aptamer Technique; D016019:Survival Analysis; D019562:Viral Load
D000998:Antiviral Agents; D052157:Aptamers, Nucleotide
Antiviral agents; DNA aptamers; Grouper; SELEX, SGIV