Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok, 10400, Thailand.
Center of Excellence for Shrimp Molecular Biology and Biotechnology (Centex Shrimp), Faculty of Science, Mahidol University, Bangkok, 10400, Thailand.
BMC Microbiol. 2021 Mar 23;21(1):88. doi: 10.1186/s12866-021-02148-8.
Viruses cause significant economic losses to shrimp aquaculture worldwide. In severe cases, they can lead to 100% mortality within a matter of days, hence the aquaculture industry requires antiviral strategies to minimize economic impacts. Currently, a double-stranded RNA (dsRNA)-based platform has been proven effective at a laboratory scale. The bottleneck for its industrialization is the lack of low-cost, efficient and practical delivery approaches. In an effort to bridge the gap between laboratory and farm applications, virus-like particles (VLP) have been used as nanocarriers of dsRNA. However, the implementation of this approach still suffers from high costs and a lengthy procedure, co-expression of subunits of VLP or capsid proteins (CPs) and dsRNA can be the solution for the problem. CP and dsRNA are traditionally expressed in two different E. coli hosts: protease-deficient and RNase III-deficient strains. To condense the manufacturing of dsRNA-containing VLP, this study constructed a novel E. coli strain that is able to co-express viral capsid proteins and dsRNA in the same E. coli cell.
A novel bacterial strain DualX-B15(DE3) was engineered to be both protease- and RNase III-deficiency via P1 phage transduction. The results revealed that it could simultaneously express recombinant proteins and dsRNA.
Co-expression of viral capsid proteins and dsRNA in the same cell has been shown to be feasible. Not only could this platform serve as a basis for future cost-effective and streamlined production of shrimp antiviral therapeutics, it may be applicable for other applications that requires co-expression of recombinant proteins and dsRNA.
病毒给全球虾类养殖业造成了巨大的经济损失。在严重的情况下，它们可能在几天内导致 100%的死亡率，因此养殖业需要抗病毒策略来最大程度地减少经济影响。目前，双链 RNA（dsRNA）为基础的平台已被证明在实验室规模上有效。其工业化的瓶颈是缺乏低成本、高效和实用的输送方法。为了弥合实验室和农场应用之间的差距，病毒样颗粒（VLP）已被用作 dsRNA 的纳米载体。然而，这种方法的实施仍然存在成本高和程序冗长的问题，VLP 的亚基或衣壳蛋白（CP）与 dsRNA 的共表达可以解决这个问题。CP 和 dsRNA 传统上在两种不同的大肠杆菌宿主中表达：缺乏蛋白酶和缺乏 RNase III 的菌株。为了简化含 dsRNA 的 VLP 的制造，本研究构建了一种新型大肠杆菌菌株，该菌株能够在同一大肠杆菌细胞中共同表达病毒衣壳蛋白和 dsRNA。
通过 P1 噬菌体转导，构建了一个新型细菌菌株 DualX-B15(DE3)，使其同时缺乏蛋白酶和 RNase III。结果表明，它可以同时表达重组蛋白和 dsRNA。
在同一细胞中共同表达病毒衣壳蛋白和 dsRNA是可行的。该平台不仅可以作为未来虾类抗病毒治疗剂的经济高效和简化生产的基础，还可能适用于需要共表达重组蛋白和 dsRNA 的其他应用。
