1、基本信息

      本文利用CRISPR/Cas9技术对弓形虫尿苷酸从头与补救合成途径的关键基因ompdc、uprt同时进行敲除，获得了基因复制缺陷型虫株，本研究表明该虫株在小鼠和猫体内均为无毒。将RHΔompdcΔuprt虫株作为减毒活疫苗免疫小鼠诱导产生了强烈的体液和细胞免疫反应，对强毒株、弱毒株的攻击感染保护力为100%。值得注意的是，该疫苗免疫猫后产生了有效的保护力，ME49包囊攻毒后免疫组猫的卵囊排放总量显著降低，减卵囊率达到95.3%，能有效控制弓形虫的传播。该研究为开发预防弓形虫病的减毒活疫苗提供了新方向。

2、英文摘要

A live attenuated RHΔompdcΔuprt mutant of Toxoplasma gondii induces strong protective immunity against toxoplasmosis in mice and cats

Abstract

Toxoplasma gondii is an obligate intracellular apicomplexan parasite and is responsible for zoonotic toxoplasmosis. It is essential to develop an effective anti-T. gondii vaccine for the control of toxoplasmosis, and this study is to explore the immunoprotective effects of a live attenuated vaccine in mice and cats. The RHΔompdcΔuprt were constructed by the CRISPR-Cas9 system. Compared with the wild-type strain, the mutant notably reduced proliferation (P < 0.05). In addition, the mutant exhibited virulence attenuation in both murine (BALB/c and BALB/c-nu) and cat models. Notably, limited pathological changes were found in tissues from RHΔompdcΔuprt-injected mice. Furthermore, compared with nonimmunized group, high levels of IgG (IgG1 and IgG2a) antibodies and cytokines (IFN-γ, IL-4, IL-10, IL-2 and IL-12) in mice were detected by the mutant (P < 0.05). Remarkably, all RHΔompdcΔuprt-vaccinated mice survived a lethal challenge with RHΔku80 and ME49 and WH6 strains. The immunized sera and splenocytes, especially CD8+ T cells, could significantly extend (P < 0.05) the survival time of mice challenged with the RHΔku80 strain compared with naïve mice. In addition, compared with nonimmunized cats, cats immunized with the mutant produced high levels of antibodies and cytokines (P < 0.05), and notably decreased the shedding numbers of oocysts in feces (95.3%). The avirulent RHΔompdcΔuprt strain can provide strong anti-T. gondii immune responses, and is a promising candidate for developing a safe and effective live attenuated vaccine.