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

A plasmid construct (pICP11-LH) was designed to constitutively express long-hairpin RNA (1hRNA) against icp11 gene, which is reportedly the most highly expressed gene of White spot syndrome virus (WSSV) and likely to have an important role in viral pathogenesis. The construct was used singly and in combination with other similar constructs designed against vp28 and vp19. A total of 6 treatments, T1 (pICP11-LH; 35 mu g), T2 (pVP28-LH; 35 mu g), T3 (pVP28-LH and pVP19-LH; 17.5 mu g each), T4 (pVP28-LH and pVP19-LH; 25 mu g:10 mu g), T5 (pICP11-LH, pVP28-LH and pVP19-LH; 11.5 mu g each) and T6 (pGFP-LH; 35 mu g) were injected intramuscularly into 20 g Penaeus monodon specimens. The shrimp were challenged with WSSV 24 hpi and protection efficacy was measured in terms of survival and viral load 15 days after challenge. Appropriate negative and positive controls were used. T2 and T3 offered highest protection (75%) followed by T1 (67%) and T4 and T5 groups (58%), while T6 showed 25% protection. In all the target specific treatments, the viral load as estimated by single tube WSSV kit was kept in check (10-100 copies), whereas in the unimmunized challenged controls it progressed to severe infection (>10(5) copies). In spite of over 3 times higher expression of ICP11 compared to VP28, its knockdown by pICP11-LH did not offer any protective advantage over pVP28-LH, either singly or in combination. Moreover, none of the combinations bettered the protection efficacy of pVP28-LH administered alone. To investigate concerns about deleterious effect of plasmid persistence and constitutive expression on shrimp growth, a lab-scale 1 month growth study was conducted with 4 treatments T2, T3, T4 and T6, where no difference in specific growth rate was observed compared to controls.