RNA interference-based therapeutics for shrimp viral diseases

Abstract

RNA interference (RNAi) has emerged as a powerful tool to manipulate gene expression in the laboratory. The presence of a double-stranded RNA (dsRNA) in eukaryotic cells triggers this post-transcriptional gene-silencing mechanism, leading to a sequence-specific degradation of the target mRNA. Among its many potential biomedical applications, silencing of viral genes stands out as a promising therapeutic strategy. Marine shrimp viral diseases, especially white spot disease (WSD), represents one of the most attractive targets for the development of therapeutic RNAi owing to its widespread economic impact. This review summarizes the current knowledge in the therapeutic application of RNAi for combating viral diseases in shrimp. The basic principles of RNAi are described, focusing on features important for its therapeutic manipulation. Subsequently, a stepwise strategy for the development of therapeutic RNAi is presented.KEY WORDS: RNAi therapy · Shrimp diseases · Small interfering RNA · siRNA · Long hairpin RNA · lhRNA Resale or republication not permitted without written consent of the publisherDis Aquat Org 86: [263][264][265][266][267][268][269][270][271][272] 2009 exception of MSGS, are classified as OIE notifiable (OIE 2006). Among these, WSSV has had the greatest impact on shrimp farming and is currently considered the most important shrimp disease in terms of distribution and economic losses. Current therapeutic strategiesManaging viral diseases has been the greatest challenge to the shrimp industry because short-term strategies, such as the use of immunostimulants, bioremediators and probiotics, have their own limitations in terms of their efficacy, practicability, cost, and, above all, reproducibility. Unlike finfishes, the shrimps lack a developed adaptive immune system, which is a prerequisite for the development of any protein vaccines. Subunit vaccines (Witteveldt et al. 2004a(Witteveldt et al. ,b, 2006 have been reported to provide some degree of protection to shrimps from viral infection, but their field level potential is yet to be seen. In this scenario, RNAi, which seems to offer great promise in terms of treating human and animal diseases, is looked upon with great hope by shrimp health managers. ENDOGENOUS RNAi PATHWAYObservations were made in plants (Napoli et al. 1990, van der Krol et al. 1990) and nematodes (Lee et al. 1993),which hinted at the existence of the RNAi pathway years before double-stranded RNA (dsRNA) molecules were identified as the key component of this evolutionarily conserved post-transcriptional silencing pathway in eukaryotic cells (Fire et al. 1998). Since then, we have witnessed significant advances in the understanding of how RNAi functions and identified several effective ways to manipulate it in the laboratory. Although a detailed account on the mechanistic aspects of RNAi is beyond the scope of this review and can be found elsewhere (Bartel 2004, Carmell & Hannon 2004, Cullen 2004, Meister & Tuschl 2004, Murchison & Hannon 2004, Du & Zamore 2005, Kim 20...

Protection of Penaeus monodon from Infection of White spot syndrome virus by DNA Construct Expressing Long Hairpin-RNA Against ICP11 Gene

“…Long hairpin RNA expression construct against WSSV icp11 gene was made following the procedure of Krishnan et al [10,11]. The selected portion (204 bp) of the icp11 gene was amplified from WSSV infected shrimp genomic DNA using a linker primer pair 5 0 -AAAGCTAGCGCTGGTGGGG GATGATACTA-3 0 and 3 0 -AAAAAGCTTGGCACAATCATGGGTTGAAT-5 0 and cloned in sense orientation between NheI and HindIII sites of the pcDNA3.1(?)…”

“…The pICP11-LH prepared in this study and three LH constructs prepared earlier, namely pVP28-LH, pVP19-LH and pGFP-LH [10,11], were used singly and in combination to evaluate their protection efficiency against WSSV. A total of eight treatment groups with six animals each were set up in triplicate.…”

“…Moreover, since Krishnan et al [10,11] could not obtain total protection by targeting vp28, it seemed likely that targeting more than one crucial viral genes simultaneously, would improve protection as it does in case of subunit vaccines. Therefore, lhRNA expressing constructs targeted against icp11, vp28 and vp19 were used in various combinations to see the effect on survival of challenged shrimp.…”

“…This is also important in the light that despite being the most highly expressed protein, it was not reported as a major antigenic protein in numerous studies conducted since the virus was first reported in 1992. A recent study from our lab [10,11] demonstrated the efficacy of DNA constructs capable of expressing lhRNA against WSSV capsid genes (vp19 and vp28) in protecting Penaeus monodon from WSSV infection. The results showed VP28 to be more protective than VP19, which is why several researchers have targeted this protein for vaccine development [8,21,28,31].…”

Sequence-optimized and targeted double-stranded RNA as a therapeutic antiviral treatment against infectious myonecrosis virus in Litopenaeus vannamei

“…The insidious presentation of this disease, which can manifest in a gradual loss of animals over the duration of the growout period, makes IMNV an excellent target for a therapeutic agent because treatments could be initiated immediately following a diagnosis of IMNV in a pond. RNA interference (RNAi) is one possible and promising method to mitigate viral disease in shrimp (Robalino et al 2005, Krishnan et al 2009, Shekhar & Lu 2009, Hirono et al 2011. RNAi has been used to prevent shrimp disease caused by white spot syndrome virus (WSSV) (Robalino et al 2005, Xu et al 2007, yellowhead virus (YHV) (Tirasophon et al 2005, Yodmuang et al 2006, Taura syndrome virus (TSV) (Robalino et al 2004), Penaeus stylirostris densovirus (PstDNV) (formerly called infectious hypodermal and hematopoietic necrosis virus, IHHNV) (Ho et al 2011), and IMNV .…”