Proteomics Analysis Of Mouse Serum Following The Immunization With Live UV-attenuated Toxoplasma Gondii RH Strain

Toxoplasma gondii is an intracellular protozoa that can infect almost all warm-blooded animals,including humans.Although there are many anti-Toxoplasma gondii vaccines,the systematically biological evaluation of vaccine efficacy are missing.This study mainly contains two parts.Firstly we prepared and evaluated three vaccination models.Finally we used a quantitative proteomics technology to profile the protein variations in mouse sera following the immunization with live UV-attenuated Toxoplasma gondii RH strain.Vaccine preparations and evaluations:We firstly constructed the TGME49₂76860 gene knockout RH strain using CRISPR-Cas9 technology,and found that the virulence of this gene knockout strain was not significantly reduced by using in vitro plaque experiments and in vivo tests.Next,we successfully constructed a DNA vaccine that simultaneously expressed TGME49₂76860 and TgSPATR.After immunization with this DNA vaccine mice significantly prolonged their survival time.However,these mice died within 15 days post infection.Finally,we successfully prepared an attenuated live vaccine by using ultraviolet irradiation.Plaque experiments,flow cytometry,electron microscopy and animal experiments showed that the optimal exposure time was 14min.Animal experiments showed that the immunized mice survive from challenge with 100 wild-type RH tachyzoites and the survival rate reached 100%.Proteomics analysis and evaluation:According to the animal model of ultraviolet attenuated vaccine,we collected the blood from mice in the control group and the 28-day post-challenge group,respectively.Serum samples were analysed by using TMT?tandem mass tags?proteomics.The results showed that immunized mice showed specific proteomic profiles when compared with controls.By using GO analysis and KEGG pathway analysis,we found that the differentially expressed proteins?DEPs?in immunized mice were significantly enriched in signaling pathways related to immunity and antagonism.Worthy to be mentioned,disease-related signaling pathways were also enriched.In summary,we firstly profiled the proteome in mouse sera during the immunization process with the ultraviolet attenuated vaccine.This study not only lays the foundation for further exploration of the mechanism underlying Toxoplasma gondii vaccination,but also provides valuable data for the control and treatment of toxoplasmosis.