Abstract
BackgroundSoft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis). More efficient methods of controlling and detecting STIV infections are urgently needed. MethodsIn this study, we generated eight single-stranded DNA (ssDNA) aptamers against STIV using systematic evolution of ligands by exponential enrichment (SELEX).ResultsThe aptamers formed representative stem-loop secondary structures. Electrophoretic mobility shift assays and fluorescent localization showed that the selected aptamers had high binding affinity for STIV. Aptamer QA-36 had the highest calculated binding affinity (Kd) of 53.8 nM. Flow cytometry and fluorescence microscopy of cell-aptamer interactions demonstrated that QA-12 was able to recognize both STIV-infected cells and tissues with a high level of specificity. Moreover, the selected aptamers inhibited STIV infection in vitro and in vivo, with aptamer QA-36 demonstrating the greatest protective effect against STIV and inhibiting STIV infection in a dose-dependent manner.DiscussionWe generated DNA aptamers that bound STIV with a high level of specificity, providing an alternative means for investigating STIV pathogenesis, drug development, and medical therapies for STIV infection.ConclusionsThese DNA aptamers may thus be suitable antiviral candidates for the control of STIV infections.
Highlights
Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis)
We investigated the specificities of aptamer-binding to STIV and to STIV-infected cells in vitro and to STIV-infected tissues in vivo
Selection of single-stranded DNA (ssDNA) aptamers against STIV Specific enrichment of each selected pool was monitored by enzyme-linked immunosorbent assay (ELISA). ssDNA binding increased as the selection cycles progressed, confirming that the selected pools were specific to STIV. ssDNA binding to STIV peaked at the 8th pool (Figure 1a), and eight ssDNA aptamers were isolated from the 8th pool (Table 1) based on their highly-specific STIV binding
Summary
Soft-shelled turtle iridovirus (STIV) causes severe systemic disease in farmed soft-shelled turtles (Trionyx sinensis). The rapid growth of turtle aquaculture has led to outbreaks of viral, bacterial, and parasitic diseases that threaten the industry’s future development Among these outbreaks, soft-shelled turtle iridovirus (STIV) isolated from T. sinensis with ‘red neck disease’ has caused great economic losses [1], indicating an urgent need for effective diagnostic and therapeutic agents to control STIV infections in aquaculture systems. Aptamers have distinct three-dimensional structures and are characterized by complex structural features including stems, loops, hairpins, and pseudoknots [8, 9] Their high specificity, low immunogenicity and lack of toxicity mean that aptamers have been used as specific probes in many areas, such as diagnostics, pathogen detection, and cancer research [10,11,12,13,14]. RNA aptamers with therapeutic potential for viral hemorrhagic septicemia virus and Hirame rhabdovirus in fish have been generated using purified virus particles as targets [20, 21]
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