Protection of Shrimp Penaeus monodon from WSSV Infection Using Antisense Constructs

Abstract

White spot syndrome caused by white spot syndrome virus (WSSV) is one of the most threatening diseases of shrimp culture industry. Previous studies have successfully demonstrated the use of DNA- and RNA-based vaccines to protect WSSV infection in shrimp. In the present study, we have explored the protective efficacy of antisense constructs directed against WSSV proteins, VP24, and VP28, thymidylate synthase (TS), and ribonucleotide reductase-2 (RR2) under the control of endogenous shrimp histone-3 (H3) or penaedin (Pn) promoter. Several antisense constructs were generated by inserting VP24 (pH3-VP24, pPn-VP24), VP28 (pH3-VP28, pPn-VP28), TS (pH3-TS, pPn-TS), and RR2 (pH3-RR2) in antisense orientation. These constructs were tested for their protective potential in WSSV infected cell cultures, and their effect on reduction of the viral load was assessed. A robust reduction in WSSV copy number was observed upon transfection of antisense constructs in hemocyte cultures derived from Penaeus monodon and Scylla serrata. When tested in vivo, antisense constructs offered a strong protection in WSSV challenged P. monodon. Constructs expressing antisense VP24 and VP28 provided the best protection (up to 90 % survivability) with a corresponding decrease in the viral load. Our work demonstrates that shrimp treated with antisense constructs present an efficient control strategy for combating WSSV infection in shrimp aquaculture.

Antisense RNA mediated protection from white spot syndrome virus (WSSV) infection in Pacific white shrimp Litopenaeus vannamei

“…Temporal analysis of vp39 gene expression showed it to be expressed after 12 h of viral infection, and its presence was observed in both virions and the viral envelop (Zhu et al, 2006). While 90% protection by the antisense construct in P. monodon was reported by Ahanger et al (2014) with the challenge dose of 1 × 10 3 WSSV, in our study on L. vannamei, 60% protection at day 9 was registered with a challenge dose of 2 × 10 4 copies. There may be a link between the protections offered versus the WSSV challenge dose.…”

“…La Fauce and Owens (2013) used antisense plasmid constructs in live bacterial cells to orally deliver short interfering RNA against the shrimp virus Penaeus merguiensis densovirus, a strain of hepatopancreatic parvovirus, in a cricket model. Later, Ahanger et al (2014) reported the effect of antisense constructs against WSSV infection in P. monodon both in vivo and in vitro. They used antisense RNA constructs against WSSV ORFs (VP24, VP28, thymidylate synthase and ribonucleotide reductase-2) under the control of constitutive host promoters such as histone3 and penaeidin, and found substantial protection from WSSV infection both in haemocyte cell cultures (derived from P. monodon and Scylla serrata) and experimental P. monodon.…”

“…The envelope proteins are the main candidate vaccination molecules because they are the first to interact with the host defence system to stimulate a protective immune response, and are involved in virion morphogenesis and eliciting antiviral mechanisms in the host (Chang et al, 2008;Reske et al, 2007). Ahanger et al (2014) studied the effect of antisense constructs targeting VP28 gene of WSSV in P. monodon and reported 90% survival of the animals. The results are in agreement with previous studies wherein VP28 was observed to be highly immunogenic and hence a potential vaccine candidate against WSSV infection (Kulkarni et al, 2013;Yumiao et al, 2013).…”

“…Escobedo-Bonilla (2011) summarized the role of different shrimp proteins in antiviral immunity brought about by RNAi silencing mechanisms such as toll-like receptor (Labreuche et al, 2009), Rab7-like proteins that participate in virus entry (Wu and Zhang, 2007), caspase-3 protein involved in apoptosis (Rijiravanich et al, 2008) and proPO activity (Charoensapsri et al, 2009). The use of antisense constructs under the control of a constitutive promoter for combating WSSV infection in Penaeus monodon was Aquaculture 435 (2015) 306-309 recently reported by Ahanger et al (2014), who observed that it could elicit good protection in WSSV challenged shrimp. Viral envelope and coat proteins are ideal targets for vaccination studies as they are the first to interact with the host cell and play vital roles in entry and multiplication of the virus inside the host (Chazal and Gerlier, 2003;Chiu and Chang, 2002).…”

RESPONS IMUN UDANG WINDU Penaeus monodon TERHADAP VAKSIN dsRNA VP-24 PADA DOSIS BERBEDA

“…Teknologi vaksinasi menggunakan agen virus dsRNA berkembang sangat pesat terutama dibidang medis, pertanian, dan akuakultur (Jin et al, 2010;Lichner et al, 2003;Rowley & Pope, 2012). Di bidang akuakultur, teknologi ini banyak dikembangkan untuk menanggulangi penyakit yang disebabkan oleh virus pada udang (Mejía-Ruíz et al, 2011;Ahanger et al, 2014). Beberapa penelitian vaksinasi udang menggunakan agen WSSV sebagai sumber vaksin telah menunjukkan kemajuan dan memperoleh hasil yang menggembirakan (Huang et al, 2014;Kumar et al, 2015;Puneeth et al, 2017).…”