Laboratorio de Inmunología y Virología, Universidad Nacional de Quilmes, Bernal, B1876BXD Buenos Aires, Argentina.
Biomed Res Int. 2013;2013:298598. doi: 10.1155/2013/298598. Epub 2013 Jul 24.
Group A rotaviruses are the major etiologic agents of acute gastroenteritis worldwide in children and young animals. Among its structural proteins, VP6 is the most immunogenic and is highly conserved within this group. Lactococcus lactis is a food-grade, Gram-positive, and nonpathogenic lactic acid bacteria that has already been explored as a mucosal delivery system of heterologous antigens. In this work, the nisin-controlled expression system was used to display the VP6 protein at the cell surface of L. lactis. Conditions for optimal gene expression were established by testing different nisin concentrations, cell density at induction, and incubation times after induction. Cytoplasmic and cell wall protein extracts were analyzed by Western blot and surface expression was confirmed by flow cytometry. Both analysis provided evidence that VP6 was efficiently expressed and displayed on the cell surface of L. lactis. Furthermore, the humoral response of mice immunized with recombinant L. lactis was evaluated and the displayed recombinant VP6 protein proved to be immunogenic. In conclusion, this is the first report of displaying VP6 protein on the surface of L. lactis to induce a specific immune response against rotavirus. These results provide the basis for further evaluation of this VP6-displaying L. lactis as a mucosal delivery vector in a mouse model of rotavirus infection.
A 组轮状病毒是全球儿童和幼小动物急性胃肠炎的主要病原体。在其结构蛋白中，VP6 是最具免疫原性的，并且在该组内高度保守。乳球菌是一种食品级、革兰氏阳性、非致病性的乳酸菌，已经被探索作为异源抗原的黏膜传递系统。在这项工作中，使用乳链菌肽控制表达系统将 VP6 蛋白展示在乳球菌的细胞表面。通过测试不同的乳链菌肽浓度、诱导时的细胞密度和诱导后的孵育时间，确定了最佳基因表达的条件。通过 Western blot 分析细胞质和细胞壁蛋白提取物，并通过流式细胞术确认表面表达。这两种分析都提供了证据表明 VP6 被有效地表达并展示在乳球菌的细胞表面上。此外，还评估了用重组乳球菌免疫的小鼠的体液免疫反应，证明了展示的重组 VP6 蛋白具有免疫原性。总之，这是首次报道将 VP6 蛋白展示在乳球菌表面上，以诱导针对轮状病毒的特异性免疫反应。这些结果为进一步评估这种展示 VP6 的乳球菌作为轮状病毒感染小鼠模型中的黏膜传递载体提供了基础。
