Technical University of Denmark, Denmark.
Qiagen, Aarhus, Denmark.
Fish Shellfish Immunol. 2019 Feb;85:99-105. doi: 10.1016/j.fsi.2018.06.056. Epub 2018 Jun 30.
The highly effective DNA vaccines against diseases caused by fish rhabdoviruses in farmed fish consist of a DNA plasmid vector encoding the viral glycoprotein under the control of a constitutive cytomegalovirus promoter (CMV). Among others, attempts to improve efficacy and safety of these DNA vaccines have focused on regulatory elements of plasmid vectors, which play a major role in controlling expression levels of vaccine antigens. Depending on the context, use of a fish-derived promoter with minimal activity in mammalian cells could be preferable. Another aspect related to the CMV promoter is that constitutive expression of the vaccine antigen may lead to rapid elimination of antigen expressing cells in the fish and thereby potentially reduce the long-term effects of the vaccine. In this study, we compared DNA vaccines with the interferon-inducible Mx promoter from rainbow trout and the CMV promoter, respectively. Plasmid constructs encoding the enhanced green fluorescent protein (EGFP) were used for the in vitro analysis, whereas DNA vaccines encoding the glycoprotein (G) of the viral haemorrhagic septicaemia virus (VHSV) were applied for the in vivo examination. The in vitro analysis showed that while the DNA vaccine with the CMV promoter constitutively drove the expression of EGFP in both fish and human cell lines, the DNA vaccine with the Mx promoter inducibly enhanced the expression of EGFP in the fish cell line. To address the impact on protection, a time-course model was followed as suggested by Kurath et al. (2006), where vaccinated fish were challenged with VHSV at 2, 8 and 78 weeks post-vaccination (wpv). The DNA vaccine with the CMV promoter protected at all times, while vaccination with the DNA vaccine containing the Mx promoter only protected the fish at 8 wpv. However, following induction with Poly (I:C) one week before the challenge, high protection was also evident at 2 wpv. In conclusion, the results revealed a more fish host dependent activity of the trout Mx promoter compared to the traditionally used cross species-active CMV promoter, but improvements will be needed for its application in DNA vaccines to ensure long term protection.
针对养殖鱼类中鱼类呼肠孤病毒引起的疾病的高效 DNA 疫苗，由在组成型巨细胞病毒启动子（CMV）控制下编码病毒糖蛋白的 DNA 质粒载体组成。除其他外，提高这些 DNA 疫苗的功效和安全性的尝试集中在质粒载体的调节元件上，这些元件在控制疫苗抗原的表达水平方面起着主要作用。根据具体情况，使用在哺乳动物细胞中活性最低的鱼类来源启动子可能更可取。与 CMV 启动子相关的另一个方面是，疫苗抗原的组成型表达可能导致鱼类中表达抗原的细胞迅速消除，从而可能降低疫苗的长期效果。在这项研究中，我们比较了干扰素诱导的虹鳟鱼 Mx 启动子和 CMV 启动子的 DNA 疫苗。分别使用编码增强型绿色荧光蛋白（EGFP）的质粒构建体进行体外分析，而编码病毒性出血性败血症病毒（VHSV）糖蛋白（G）的 DNA 疫苗则用于体内检查。体外分析表明，尽管具有 CMV 启动子的 DNA 疫苗在鱼类和人类细胞系中持续驱动 EGFP 的表达，但具有 Mx 启动子的 DNA 疫苗可诱导鱼类细胞系中 EGFP 的表达增强。为了解决保护的影响，遵循了 Kurath 等人（2006 年）提出的时间过程模型，其中在接种疫苗后 2、8 和 78 周（wpv）用 VHSV 对接种疫苗的鱼进行攻毒。具有 CMV 启动子的 DNA 疫苗在所有时间都提供保护，而含有 Mx 启动子的 DNA 疫苗仅在 8 wpv 时保护鱼类。然而，在攻毒前一周用 Poly（I：C）诱导后，在 2 wpv 时也明显具有高保护作用。总之，结果表明，与传统的跨物种活性 CMV 启动子相比，虹鳟鱼 Mx 启动子在鱼类宿主中的活性更高，但为了确保长期保护，还需要对其在 DNA 疫苗中的应用进行改进。
