Kaldis Angelo, Uddin Muhammed Salah, Guluarte Jose Ortiz, Martin Coby, Alexander Trevor W, Menassa Rima
London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, Canada.
Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, Lethbridge, AB, Canada.
Front Plant Sci. 2023 Sep 15;14:1251046. doi: 10.3389/fpls.2023.1251046. eCollection 2023.
Bovine respiratory disease (BRD) affects feedlot cattle across North America, resulting in economic losses due to animal treatment and reduced performance. In an effort to develop a vaccine candidate targeting a primary bacterial agent contributing to BRD, we produced a tripartite antigen consisting of segments of the virulence factor Leukotoxin A (LktA) and lipoprotein PlpE from , fused to a cholera toxin mucosal adjuvant (CTB). This recombinant subunit vaccine candidate was expressed in the leaves of plants, with accumulation tested in five subcellular compartments. The recombinant protein was found to accumulate highest in the endoplasmic reticulum, but targeting to the chloroplast was employed for scaling up production due the absence of post-translational modification while still producing feasible levels. Leaves were freeze dried, then orally administered to mice to determine its immunogenicity. Sera from mice immunized with leaf tissue expressing the recombinant antigen contained IgG antibodies, specifically recognizing both LktA and PlpE. These mice also had a mucosal immune response to the CTB+LktA+PlpE protein as measured by the presence of LktA- and PlpE-specific IgA antibodies in lung and fecal material. Moreover, the antigen remained stable at room temperature with limited deterioration for up to one year when stored as lyophilized plant material. This study demonstrated that a recombinant antigen expressed in plant tissue elicited both humoral and mucosal immune responses when fed to mice, and warrants evaluation in cattle.
牛呼吸道疾病（BRD）影响着北美饲养场的牛，因动物治疗和性能下降而导致经济损失。为了开发一种针对导致BRD的主要细菌病原体的候选疫苗，我们制备了一种三联抗原，该抗原由来自的毒力因子白细胞毒素A（LktA）和脂蛋白PlpE的片段组成，并与霍乱毒素粘膜佐剂（CTB）融合。这种重组亚单位候选疫苗在植物叶片中表达，并在五个亚细胞区室中测试了其积累情况。发现重组蛋白在内质网中积累最高，但由于缺乏翻译后修饰且仍能产生可行的水平，因此将其靶向叶绿体用于扩大生产。将叶片冷冻干燥，然后口服给小鼠以确定其免疫原性。用表达重组抗原的叶片组织免疫的小鼠血清中含有IgG抗体，能特异性识别LktA和PlpE。通过在肺和粪便中检测到LktA和PlpE特异性IgA抗体，这些小鼠对CTB + LktA + PlpE蛋白也有粘膜免疫反应。此外，当作为冻干植物材料储存时，该抗原在室温下保持稳定，长达一年仅有有限的降解。这项研究表明，在植物组织中表达的重组抗原在喂给小鼠时能引发体液免疫和粘膜免疫反应，值得在牛身上进行评估。
