Role of Toxoplasma gondii calcium-binding protein (TgpCaBP) and its implications for the development of attenuated live vaccines

Abstract

ABSTRACT Toxoplasma gondii is an obligate intracellular protozoan parasite and the causative agent of toxoplasmosis in humans and animals. To date, an ideal vaccine against toxoplasmosis has not yet been developed. Our previous study demonstrated that deletion of the TgpCaBP gene, which encodes a calcium-binding EF-hand protein, impaired calcium flux, invasion, and egress, thereby reducing T. gondii virulence. However, the mechanisms underlying this loss of virulence and the potential of RHΔTgpCaBP as a live attenuated vaccine remain unclear. Here, we showed that TgpCaBP deletion significantly downregulated the expression of proteins associated with invasion, egress, and pyrimidine metabolism. In vivo evaluations revealed that co-administration of RHΔTgpCaBP and sulfadiazine showed limited pathological adverse effects in mice—all mice survived immunization with 100 tachyzoites without obvious organ injury, and conferred protection against lethal wild-type RH strain challenge (≤1,000 tachyzoites). Flowcytometry assays demonstrated that RHΔTgpCaBP strain immunization increased IFN-γ production in circulating CD4+ and CD8+ T cells and the frequencies of splenic CD4+ and CD8+ effector memory T cells, memory B cells, and activated B cells. In conclusion, TgpCaBP plays an important role in regulating the virulence of T. gondii by modulating the expression of proteins related to pyrimidine metabolism, invasion, and egress. Co-immunization with 100 RHΔTgpCaBP tachyzoites and sulfadiazine shows favorable safety profiles and protective efficacy in mice, triggers observable immune memory, and may serve as a promising candidate for an attenuated live vaccine against toxoplasmosis. IMPORTANCE Toxoplasmosis, caused by T. gondii, poses substantial public health and economic burdens worldwide. Although medicinal plants are traditionally the most promising source for the treatment of toxoplasmosis, no effective therapeutic regimen has been developed to date. Thus, the development of safe, effective, and long-lasting vaccines has become key for the prevention and control of toxoplasmosis. This study demonstrated that the deletion of TgpCaBP, a gene encoding an EF-hand calcium-binding protein, profoundly attenuated the virulence of the virulent RH strain by disrupting key pathogenic processes. More importantly, the RHΔTgpCaBP strain, administered in combination with sulfadiazine, emerged as a promising live attenuated vaccine candidate. It exhibited favorable safety profiles in mice and elicited an immunomodulatory trend. Although further optimization is warranted, this study establishes a foundational framework for developing live attenuated vaccines against toxoplasmosis. Toxoplasmosis, caused by T. gondii, poses substantial public health and economic burdens worldwide. Although medicinal plants are traditionally the most promising source for the treatment of toxoplasmosis, no effective therapeutic regimen has been developed to date. Thus, the development of safe, effective, and long-lasting vaccines has become key for the prevention and control of toxoplasmosis. This study demonstrated that the deletion of TgpCaBP, a gene encoding an EF-hand calcium-binding protein, profoundly attenuated the virulence of the virulent RH strain by disrupting key pathogenic processes. More importantly, the RHΔTgpCaBP strain, administered in combination with sulfadiazine, emerged as a promising live attenuated vaccine candidate. It exhibited favorable safety profiles in mice and elicited an immunomodulatory trend. Although further optimization is warranted, this study establishes a foundational framework for developing live attenuated vaccines against toxoplasmosis.