Genetic Disruption of Toxoplasma gondii peroxiredoxin (TgPrx) 1 and 3 Reveals the Essential Role of TgPrx3 in Protecting Mice from Fatal Consequences of Toxoplasmosis

Ragab M Fereig,Yoshifumi Nishikawa

doi:10.3390/ijms23063076

Abstract

Toxoplasma gondii is a worldwide protozoan parasite that endangers human health and causes enormous economic losses to the animal production sector. A safe and effective vaccine or treatment is needed to reduce these hazards. In this study, we revealed the cyto-nuclear and mitochondrial localization of TgPrx1 and TgPrx3 proteins, respectively. We knocked out the T. gondii peroxiredoxin (TgPrxKO) 1 and 3 genes using a parental type II Prugniaud strain lacking KU80 and HXGPRT genes (PruΔku80Δhxgprt) via CRISPR-Cas9 technology. The successful KO was confirmed using PCR, IFAT, and Western blotting in two clones of both target genes, named TgPrx1KO and TgPrx3KO. Regarding in vitro assays, no significant variations between any of the knocked-out clones in TgPrx1KO or TgPrx3KO parasite strains, or even PruΔku80Δhxgprt, were obtained in rates of infection, proliferation, or egress. Nevertheless, mice that were infected with tachyzoites of the TgPrx3KO strain showed a marked decrease in survival rate compared with TgPrx1KO- and PruΔku80Δhxgprt-infected mice. This effect was confirmed using different mouse strains (ICR and C57BL/6J mice), sexes (male and female), and immunological backgrounds (ICR and SCID mice). In addition, TgPrx1KO and TgPrx3KO induced high levels of interferon gamma (IFN-γ) in infected mice at 8 days post infection, and increased IL-6 and IL-12p40 production from murine macrophages cultivated in vitro. The results of the present study suggested that TgPrx3 can induce anti-T. gondii immune responses that protect the mice from fatal consequences of toxoplasmosis. The results of our current and previous studies represent TgPrx3 as an excellent candidate for sub-unit vaccines, suggesting it may contribute to the control of toxoplasmosis for susceptible humans and animals.

Highlights

National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Department of Animal Medicine, Faculty of Veterinary Medicine, South Valley University, Qena City 83523, Qena, Egypt
The results of the present study suggested that TgPrx3 can induce anti-T. gondii immune responses that protect the mice from fatal consequences of toxoplasmosis
To determine the cellular localization of TgPrx1 and TgPrx3, T. gondii-infected Vero cells were stained via specific anti-sera against TgPrx1 and TgPrx3 prepared in BALB/c mice

Summary

Introduction

Toxoplasma gondii is a worldwide protozoan parasite that endangers human health and causes enormous economic losses to the animal production sector. No significant variations between any of the knocked-out clones in TgPrx1KO or TgPrx3KO parasite strains, or even Pru∆ku80∆hxgprt, were obtained in rates of infection, proliferation, or egress. The results of the present study suggested that TgPrx can induce anti-T. gondii immune responses that protect the mice from fatal consequences of toxoplasmosis. The results of our current and previous studies represent TgPrx as an excellent candidate for sub-unit vaccines, suggesting it may contribute to the control of toxoplasmosis for susceptible humans and animals. As a member of the phylum Apicomplexa, Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite that can infect almost all warm-blooded animals such as humans and cats. There are no successful Toxoplasma vaccines that can be applied clinically [2]

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Conclusion