Yang Jinru, Nie Linchong, Song Yining, Yang Zipeng, Yang Liulu, Ren Hongjie, Li Wenhao, Mahmmod Yasser, Zhang Xiu-Xiang, Yuan Ziguo, Yuan Hao, Zhang Yan
College of Science, Shenyang University, Shenyang 110044, China.
Key Laboratory of Zoonosis Prevention and Control of Guangdong Province, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China.
Vaccines (Basel). 2025 Aug 11;13(8):851. doi: 10.3390/vaccines13080851.
() is a significant opportunistic zoonotic protozoan, presenting a substantial risk to human health and livestock. Consequently, the development of an effective vaccine against toxoplasmosis is imperative. This study focuses on the GRA12 protein as a target for developing a recombinant protein vaccine, with its efficacy evaluated through immunization trials in cats. We expressed recombinant GRA12 protein in and immunized cats with the purified antigen. The cats were categorized into four groups: G1 (PBS control), G2 (ISA 201 adjuvant alone), G3 (rGRA12 vaccine), and G4 (rGRA12 combined with ISA 201 adjuvant). All cats underwent subcutaneous immunizations on days 0, 14, and 28. Subsequently, serum levels of IgG (including IgG1 and IgG2a subclasses) and cytokines (IFN-γ, IL-2, TNF-α, IL-4, IL-10) were measured by enzyme-linked immunosorbent assay (ELISA). Two weeks after the third immunization (42 DPI), each cat was intraperitoneally infected with 1 × 10 RH tachyzoites. Oocyst shedding, survival duration, and burden were monitored to assess vaccine-induced immunity. The results indicate that immunization with recombinant rGRA12 protein significantly elevated IgG, IgG1, and IgG2a antibody levels in cats. G4 displayed elevated IgG levels post-immunization compared to G1 and G2, with an IgG1/IgG2a ratio > 1, indicating a mixed Th1/Th2 immune response. G4 also showed significantly increased IFN-γ, IL-2, TNF-α, and IL-4 levels compared to G1 ( < 0.05), while IL-10 remained unchanged. After infection, total oocyst counts were 4.61 × 10 (G1), 4.49 × 10 (G2), 3.58 × 10 (G3), and 2.59 × 10 (G4), with G3/G4 showing 20.1-27.9% reduction relative to G1 ( < 0.05). Survival analysis revealed that groups G3 and G4 exhibited significantly longer median survival times (38 and 60 days, respectively; G4 with no mortality) compared to G1 and G2 (19 and 26 days, respectively). Additionally, parasite burdens in the brain, heart, lungs, liver, and spleen were significantly reduced in G3/G4 compared to G1/G2 ( < 0.01). In summary, the recombinant GRA12 vaccine significantly enhanced host survival and reduced parasite burden, demonstrating its potential as an effective toxoplasmosis vaccine candidate. These findings provide valuable data for future toxoplasmosis vaccine development.
（）是一种重要的机会性人畜共患原生动物，对人类健康和家畜构成重大风险。因此，开发一种有效的抗弓形虫病疫苗势在必行。本研究聚焦于GRA12蛋白作为开发重组蛋白疫苗的靶点，并通过在猫身上进行免疫试验来评估其疗效。我们在 中表达重组GRA12蛋白，并用纯化的抗原来免疫猫。将猫分为四组：G1（PBS对照组）、G2（仅ISA 201佐剂组）、G3（rGRA12疫苗组）和G4（rGRA12与ISA 201佐剂联合组）。所有猫在第0、14和28天进行皮下免疫。随后，通过酶联免疫吸附测定（ELISA）测量血清中IgG（包括IgG1和IgG2a亚类）和细胞因子（IFN-γ、IL-2、TNF-α、IL-4、IL-10）的水平。在第三次免疫后两周（感染后天数42天），每只猫腹腔内感染1×10个RH速殖子。监测卵囊排出、存活时间和 负荷以评估疫苗诱导的免疫力。结果表明，用重组rGRA12蛋白免疫可显著提高猫体内IgG、IgG1和IgG2a抗体水平。与G1和G2相比，G4免疫后IgG水平升高，IgG1/IgG2a比值>1，表明存在混合的Th1/Th2免疫反应。与G1相比，G4的IFN-γ、IL-2、TNF-α和IL-4水平也显著升高（<0.05），而IL-10保持不变。感染后，总卵囊计数分别为：G1组4.61×10、G2组4.49×10、G3组3.58×10、G4组2.59×10，G3/G4组相对于G1组显示减少20.1 - 27.9%（<0.05）。生存分析显示，与G1和G2组（分别为19天和26天）相比，G3和G4组的中位生存时间显著更长（分别为38天和60天；G4组无死亡）。此外，与G1/G2组相比，G3/G4组脑、心、肺脏、肝脏和脾脏中的寄生虫负荷显著降低（<0.01）。总之，重组GRA12疫苗显著提高了宿主的存活率并降低了寄生虫负荷，证明其作为一种有效的弓形虫病疫苗候选物的潜力。这些发现为未来弓形虫病疫苗的开发提供了有价值的数据。
