Immunization With a Live-Attenuated RH:ΔNPT1 Strain of Toxoplasma gondii Induces Strong Protective Immunity Against Toxoplasmosis in Mice

Abstract

Toxoplasmosis, one of the most important health-threatening diseases worldwide, is caused by Toxoplasma gondii , which infects a wide range of warm-blooded animals and humans, leading to enormous health and socioeconomic concerns. T. gondii can establish chronic infection to evade the immune response in hosts. Once a chronic infection has been established, the available treatments cannot efficiently control this stage of T. gondii efficiently. Moreover, the available treatments rely only on a few drugs, such as sulfapyridine and pyrimethamine, that tend to have severe side effects. Given these factors, vaccination has been considered to be the most efficient method to prevent and control this disease. However, there is currently lack of effective vaccine available for use to prevent toxoplasmosis apart form Toxovax ® , the only available vaccine, which is used in sheep to prevent abortion. To address this problem, we knocked out the NPT1 gene of the type I T. gondii strain using the CRISPR-Cas9 system, constructed a live-attenuated vaccine and evaluated its protective efficacy in a mouse model. Immunization of mice with RH:Δ NPT1 induced a high level of Toxoplasma -specific IgG1, IgG2a and total IgG 42 days after immunization. There was a significant increase in the levels of cytokines in the splenocyte suspensions of RH:Δ NPT1 -infected mice, and a mixed Th1/Th2 response was induced in the mice. Remarkably, after heterologous challenges with tachyzoites of the RH, PYS and Pru strains and cysts of the Pru strain by different infection routes, the immunized animals were protected from toxoplasmosis with a 100% survival rate, in both acute and chronic infection. In addition, compared with control mice, the Pru cyst load was clearly reduced in the brains of RH:Δ NPT1 -infected immunization-mice. Our study demonstrated that the RH:Δ NPT1 strain was able to evoke strong anti- Toxoplasma immune responses and provide effective protection against parasite strains with different levels of virulence, suggesting that the RH:Δ NPT1 strain may represent a promising live-attenuated vaccine against toxoplasmosis, which is worthy of further evaluation in food-producing animals and in definitive feline host.

RHΔgra17Δnpt1 Strain of Toxoplasma gondii Elicits Protective Immunity Against Acute, Chronic and Congenital Toxoplasmosis in Mice

“…Consistent with other live-attenuated T. gondii vaccines, mice vaccinated with RH∆gra17∆npt1 exhibited a high level of anti-T. gondii IgG antibodies [13,14,24,25]. These specific IgG antibodies play an important role in protection against T. gondii infection via hindering the attachment of the parasite to the host cells, limiting its ability to establish infection, and activation of the classical complement pathway [26].…”

“…Our data clearly demonstrate the ability of RH∆gra17∆npt1 to significantly reduce 91.4% of the parasite cyst burden compared to unvaccinated mice. In previous studies, it was demonstrated that vaccination using single-gene mutants, RH∆gra17 [13] and RH∆npt1 [24] resulted in 98.8% and 98.6% reduction in the parasite cyst burden, respectively. This finding suggests that although deletion of both gra17 and npt1 genes has considerably attenuated the virulence of the parasite, this was not paralleled by improvement in the double mutant's ability to limit the parasite cyst burden during latent infection.…”

“…We previously demonstrated that live-attenuated T. gondii RH strain deficient in gra17 ( Figure 1A,C) was immunogenic in mice [13]. Interestingly, the NPT1 protein of T. gondii has been reported to play a key role in the growth and virulence of T. gondii, and the deletion of npt1 gene in the RH strain also rendered the parasite immunogenic in mice [24]. We investigated whether a double mutant T. gondii strain deficient in gra17 gene and npt1 gene can be more immunogenic than mutant strain deficient in a single gene only.…”

PLK:Δgra9 Live Attenuated Strain Induces Protective Immunity Against Acute and Chronic Toxoplasmosis

“…and multiple stages of parasite infections (Wang et al, 2017(Wang et al, , 2018Xia et al, 2018b;Yang et al, 2019;Liang et al, 2020). In this study, PLK: gra9 vaccination induced a high level of immune response and resisted the infections of tachyzoites from type I or type II strains and cysts from type II strain.…”

“…Several studies on T. gondii vaccines have been done and reported in animal models. Although compared with nucleic acid vaccines ( Li et al, 2015 ; Zhang et al, 2015 ; Lu et al, 2017 ) and recombinant protein vaccines ( El Bissati et al, 2014 ; Tanaka et al, 2014 ; Wang et al, 2014a , b ), the higher protection against acute or chronic T. gondii infection provided by live attenuated vaccines ( Fox and Bzik, 2015 ; Lagal et al, 2015 ; Abdelbaset et al, 2017 ; Wang et al, 2017 , 2018 ; Xia et al, 2018b ; Yang et al, 2019 ; Liang et al, 2020 ) was highlighted both in short- and long-term vaccination using different strains in the mouse model. However, the safeness of these mutants still needs to be tested in animal models.…”