A Live Attenuated AP2X-1-Deficient Toxoplasma Strain Confers Protective Immunity Against Acute and Chronic...

Toxoplasma gondii is an obligate intracellular apicomplexan parasite that poses serious risks to immunocompromised individuals and the global livestock industry. Our previous study showed that AP2X-1 regulates stage conversion and virulence of T. gondii; however, whether AP2X-1-deficient strains can serve as live-attenuated vaccine candidates is unknown. Here, we evaluated the in vivo virulence, protective efficacy, and induced immunity of the ap2X-1 knockout strain PruΔap2X-1. In Kunming mice, infection with even the highest tested dose (5 × 10⁶ tachyzoites) of PruΔap2X-1 resulted in 100% survival without detectable brain cysts. Vaccination with 10⁶ PruΔap2X-1 tachyzoites conferred complete protection (100% survival) against acute challenge with homologous type II Pru or heterologous ToxoDB#9 PYS strains, whereas control mice exhibited only 0% to 16.7% survival. Moreover, immunized mice orally challenged with Pru cysts showed 100% survival, and no brain cysts were detected under the conditions tested, compared with only 30% survival in control mice challenged with 10 cysts and no survival with 40 cysts. The vaccination induced a Th1-biased response as evidenced by significantly elevated T. gondii-specific total IgG, IgG2a, and the Th1 cytokines IFN-γ and IL-2, along with a significantly reduced IgG1/IgG2a ratio. Collectively, the attenuated PruΔap2X-1 strain generates strong protective immunity against acute and chronic infection with low-virulence strains (the type II Pru and ToxoDB#9 PYS), as well as partial protection against the hypervirulent type I RH strain, positioning it as a potential candidate for a live-attenuated vaccine, with a favorable safety profile observed under the experimental conditions.