Division of Small Animal Surgery, Department of Clinical Veterinary Sciences, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
Institute of Parasitology, DIP, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
Front Cell Infect Microbiol. 2021 Aug 25;11:675219. doi: 10.3389/fcimb.2021.675219. eCollection 2021.
(LM) has been proposed as vaccine vector in various cancers and infectious diseases since LM induces a strong immune response. In this study, we developed a novel and safe LM-based vaccine vector platform, by engineering a triple attenuated mutant (Lm3Dx) (ΔactA, ΔinlA, ΔinlB) of the wild-type LM strain JF5203 (CC 1, phylogenetic lineage I). We demonstrated the strong attenuation of Lm3Dx while maintaining its capacity to selectively infect antigen-presenting cells (APCs) . Furthermore, as proof of concept, we introduced the immunodominant (Nc) surface antigen NcSAG1 into Lm3Dx. The NcSAG1 protein was expressed by Lm3Dx_SAG1 during cellular infection. To demonstrate safety of Lm3Dx_SAG1 , we vaccinated BALB/C mice by intramuscular injection. Following vaccination, mice did not suffer any adverse effects and only sporadically shed bacteria at very low levels in the feces (<100 CFU/g). Additionally, bacterial load in internal organs was very low to absent at day 1.5 and 4 following the 1 vaccination and at 2 and 4 weeks after the second boost, independently of the physiological status of the mice. Additionally, vaccination of mice prior and during pregnancy did not interfere with pregnancy outcome. However, Lm3Dx_SAG1 was shed into the milk when inoculated during lactation, although it did not cause any clinical adverse effects in either dams or pups. Also, we have indications that the vector persists more days in the injected muscle of lactating mice. Therefore, impact of physiological status on vector dynamics in the host and mechanisms of milk shedding requires further investigation. In conclusion, we provide strong evidence that Lm3Dx is a safe vaccine vector in non-lactating animals. Additionally, we provide first indications that mice vaccinated with Lm3Dx_SAG1 develop a strong and Th1-biased immune response against the Lm3Dx-expressed neospora antigen. These results encourage to further investigate the efficiency of Lm3Dx_SAG1 to prevent and treat clinical neosporosis.
（LM）已被提议作为各种癌症和传染病的疫苗载体，因为 LM 能诱导强烈的免疫反应。在这项研究中，我们通过工程改造野生型 LM 菌株 JF5203（CC1，系统发育谱系 I）的三重减毒突变体（Lm3Dx）（ΔactA、ΔinlA、ΔinlB），开发了一种新型且安全的 LM 基于疫苗载体平台。我们证明了 Lm3Dx 的强烈衰减，同时保持其选择性感染抗原呈递细胞（APC）的能力。此外，作为概念验证，我们将免疫优势抗原（Nc）表面抗原 NcSAG1 引入 Lm3Dx。NcSAG1 蛋白在细胞感染期间由 Lm3Dx_SAG1 表达。为了证明 Lm3Dx_SAG1 的安全性，我们通过肌肉内注射对 BALB/C 小鼠进行了疫苗接种。接种后，小鼠没有出现任何不良反应，仅在粪便中以非常低的水平（<100 CFU/g）偶尔排出细菌。此外，在接种后第 1.5 天和第 4 天以及第 2 次加强免疫后第 2 周和第 4 周，在内部器官中的细菌载量非常低或不存在，与小鼠的生理状态无关。此外，在怀孕前和怀孕期间对小鼠进行疫苗接种不会干扰怀孕结果。然而，当在哺乳期接种时，Lm3Dx_SAG1 会排入乳汁中，但无论是在母鼠还是幼鼠中都没有引起任何临床不良影响。此外，我们有迹象表明，载体在哺乳期小鼠的注射肌肉中持续存在更长的时间。因此，生理状态对宿主中载体动力学的影响和乳汁脱落的机制需要进一步研究。总之，我们提供了强有力的证据表明 Lm3Dx 是一种在非哺乳期动物中安全的疫苗载体。此外，我们提供了初步迹象表明，用 Lm3Dx_SAG1 接种的小鼠对 Lm3Dx 表达的新孢子抗原产生了强烈的 Th1 偏向性免疫反应。这些结果鼓励进一步研究 Lm3Dx_SAG1 预防和治疗临床新孢子虫病的效率。
