Enhancement of shrimp immunity against white spot syndrome virus by Macrobrachium rosenbergii nodavirus-like particle encapsulated VP28 double-stranded RNA Pitchanee Jariyapong a,b , Wattana Weerachatyanukul c , Sataporn Direkbusarakom b,d , Ikuo Hirono e , Suwit Wuthisuthimethavee b,d , Charoonroj Chotwiwatthanakun f, ⁎ a School of Medicine, Walailak University, Thasala District, Nakhonsrithammarat 80161, Thailand b Research Center of Excellence on Shrimp, Walailak University, Thasala District, Nakhonsrithammarat 80161, Thailand c Department of Anatomy, Faculty of Science, Mahidol University, Rama 6 Road, Phyathai, Bangkok 10400, Thailand d School of Agricultural Technology, Walailak University, Thasala District, Nakhonsrithammarat 80161, Thailand e Laboratory of Genome Science, Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato, Tokyo 108-8477, Japan f Nakhonsawan Campus, Mahidol University, Nakhonsawan 60130, Thailand abstract article info Article history: Received 18 March 2015 Received in revised form 8 May 2015 Accepted 11 May 2015 Available online 13 May 2015 Keywords: Macrobrachium rosenbergii nodavirus Virus-like particles Encapsulation WSSV VP28 dsRNA RNA-induced silencing complex We investigated the efficiency and downstream effects of Macrobrachium rosenbergii nodavirus-like particle (MrNv-VLP) encapsulated VP28 double-stranded RNA against white spot syndrome virus (WSSV) in shrimp. Our results showed that the VP28 gene of WSSV was significantly silenced at 72 h post-viral challenge in group pre-treated with the VLP encapsulated VP28 dsRNA. At 24 h post-treatment, the amount of VP28 dsRNA was significantly higher (two-fold) in encapsulated VP28 dsRNA-VLP pre-treated shrimp compared with non-encapsulated VP28 dsRNA pre-treated shrimp. Using quantitative real time polymerase chain reaction (PCR), we found higher and longer expression of RNA-induced silencing complex (RISC) and immune-related genes in shrimp pre-treated with encapsulated VP28 dsRNA compared to the control groups. Increasing expression of apoptotic-related genes was also associated with the encapsulated VP28 dsRNA-VLP pretreatment group at 24 h post-WSSV challenge. These results indicate that MrNv-VLP was able to efficiently deliver VP28 dsRNA into shrimp tissues, which in turn, triggered a better anti-viral response. This may represent a novel strategy for aquaculture disease management. Statement of relevance: This study developed methods for WSSV protection. © 2015 Elsevier B.V. All rights reserved. 1. Introduction Shrimp aquaculture has been severely affected by widespread infection of white spot syndrome virus (WSSV), which results in cumulative mortality up to 100% within 3 to 10 days. This impact of WSSV infection has prompted the development of many strategies to protect animals from and control the disease. One of the most effective strategies against WSSV infection is the application of RNA interference (RNAi) based therapy, which is a universal gene-silencing mechanism. RNAi antiviral response is triggered by double-stranded RNA (dsRNA) specific to the viral major capsid protein genes of WSSV, including VP19, VP24, and VP28, and is able to partially block viral progression and increase the survival rate of infected shrimp when compared to shrimp that received random dsRNA (Mejía-Ruíz et al., 2011; Sarathi et al., 2008). These findings indicate the natural existence of intact RNAi machinery within shrimp tissues. To date, two key protein components of RNAi machinery – dicer and argonaute – have been identified in Penaeus shrimp and experimentally proven to play an important role in RNA-induced silencing complex (RISC) (Chen et al., 2011; Labreuche et al., 2010; Yao et al., 2010). Similar to other types of nucleic acids, dsRNA is also negatively charged and faces a major barrier from the negatively charged cell membrane when entering the cytoplasm. To improve gene silencing, compaction of dsRNA by a cationic delivery system is a potential strategy for RNA-based vaccination (Manjila et al., 2013). Ideally, the delivery vehicle should be enhanced by intracellular uptake and be biodegradable and biocompatible without compromising the gene silencing efficacy of the dsRNA. Several non-viral delivery vectors where dsRNA could either be conjugated to or encapsulated into the interior of the delivery vehicle have been demonstrated in plants and even humans (Liu et al., 2014; Numata et al., 2014; Sakurai et al., 2003; Wang et al., 2010). Despite many reports that dsRNA is ready to use without any adjuvants or carriers, our previous results demonstrate the benefit of using an MrNV virus-like particle (MrNV-VLP)-based delivery system to carry Aquaculture 446 (2015) 325–332 ⁎ Corresponding author at: Mahidol University, Nakhonsawan Campus, Phayuhakiri, Nakhonsawan 60130, Thailand. E-mail address: Charoonroj.cho@mahidol.ac.th (C. Chotwiwatthanakun). http://dx.doi.org/10.1016/j.aquaculture.2015.05.016 0044-8486/© 2015 Elsevier B.V. All rights reserved. Contents lists available at ScienceDirect Aquaculture journal homepage: www.elsevier.com/locate/aqua-online