Large-scale production and antiviral efficacy of multi-target double-stranded RNA for the prevention of white spot syndrome virus (WSSV) in shrimp

BMC Biotechnology

Research Article

Watcharachai Meemetta¹  Thitiporn Thammasorn¹  Vanvimon Saksmerprome²  Saengchan Senapin²  Sarocha Jitrakorn²  Pakkakul Sangsuriya³  Triwit Rattanarojpong⁴ 

[1] Center of Excellence for Shrimp Molecular Biology and Biotechnology, Faculty of Science, Mahidol University, 10400, Bangkok, Thailand;Center of Excellence for Shrimp Molecular Biology and Biotechnology, Faculty of Science, Mahidol University, 10400, Bangkok, Thailand;National Center of Genetic Engineering and Biotechnology, (BIOTEC), Thailand Science Park, 12120, Pathum Thani, Thailand;Department of Biochemistry, Center of Excellence for Molecular Biology and Genomics of Shrimp, Faculty of Science, Chulalongkorn University, Bangkok, Thailand;National Center of Genetic Engineering and Biotechnology, (BIOTEC), Thailand Science Park, 12120, Pathum Thani, Thailand;Department of Microbiology, Faculty of Science, King Mongkut’s University of Technology Thonburi, 10140, Bangkok, Thailand;

关键词: Co-cultivation;    White spot syndrome virus;    dsRNA;    Shrimp;    VP28;    WSSV051;   

DOI  :  10.1186/s12896-015-0226-9

 received in 2014-12-30, accepted in 2015-11-27,  发布年份 2015

来源: Springer

【 摘 要 】

BackgroundRNA interference (RNAi) is a specific and effective approach for inhibiting viral replication by introducing double-stranded (ds)RNA targeting the viral gene. In this study, we employed a combinatorial approach to interfere multiple gene functions of white spot syndrome virus (WSSV), the most lethal shrimp virus, using a single-batch of dsRNA, so-called “multi-WSSV dsRNA.” A co-cultivation of RNase-deficient E. coli was developed to produce dsRNA targeting a major structural protein (VP28) and a hub protein (WSSV051) with high number of interacting protein partners.ResultsFor a co-cultivation of transformed E. coli, use of Terrific broth (TB) medium was shown to improve the growth of the E. coli and multi-WSSV dsRNA yields as compared to the use of Luria Bertani (LB) broth. Co-culture expression was conducted under glycerol feeding fed-batch fermentation. Estimated yield of multi-WSSV dsRNA (μg/mL culture) from the fed-batch process was 30 times higher than that obtained under a lab-scale culture with LB broth. Oral delivery of the resulting multi-WSSV dsRNA reduced % cumulative mortality and delayed average time to death compared to the non-treated group after WSSV challenge.ConclusionThe present study suggests a co-cultivation technique for production of antiviral dsRNA with multiple viral targets. The optimal multi-WSSV dsRNA production was achieved by the use of glycerol feeding fed-batch cultivation with controlled pH and dissolved oxygen. The cultivation technique developed herein should be feasible for industrial-scale RNAi applications in shrimp aquaculture. Interference of multiple viral protein functions by a single-batch dsRNA should also be an ideal approach for RNAi-mediated fighting against viruses, especially the large and complicated WSSV.

【 授权许可】

CC BY   
© Thammasorn et al. 2015