Urine proteomics for profiling of mouse toxoplasmosis using liquid chromatography tandem mass spectrometry analysis.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that causes toxoplasmosis. Urine is an easily obtained clinical sample that has been widely applied for diagnostic purposes. However, changes in the urinary proteome during T. gondii infection have never been investigated. METHODS: Twenty four-hour urine samples were obtained from BALB/c mice with acute infection [11 days post infection (DPI)], mice with chronic infection (35 DPI) and healthy controls, and were analyzed using a label-free liquid chromatography tandem mass spectrometry analysis. RESULTS: We identified a total of 13,414 peptides on 1802 proteins, of which 169 and 47 proteins were significantly differentially expressed at acute and chronic infection phases, respectively. Clustering analysis revealed obvious differences in proteome profiles among all groups. Gene ontology analysis showed that a large number of differentially expressed proteins (DEPs) detected in acute infection were associated with biological binding activity and single-organism processes. KEGG pathway enrichment analysis showed that the majority of these DEPs were involved in disease-related and metabolic pathways. CONCLUSIONS: Our findings revealed global reprogramming of the urine proteome following T. gondii infection, and data obtained in this study will enhance our understanding of the host responses to T. gondii infection and lead to the identification of new diagnostic biomarkers.

Detection of Toxoplasma gondii from sera and urine of experimentally infected mice by PCR.

Toxoplasms gondii was detected in sera and urine of acutely infected mice. Animals were inoculated intraperitoneally with tachyzoites of T. gondii, RH strain. Ten animals were killed every day from day 1 up to day 7 post infection. Urine and sera of animals were collected and stored at -20 degree C until use. PCR performed by B1 gene amplification. Toxoplasma was detected in sera from 3 days post infection and in urine from 5 days post infection. No parasite was detected in control group.

[Detection of circulating antigen in urine from mice infected with Toxoplasma tachyzoites by SPA-ELISA].

OBJECTIVE: To explore a simple and convenient immunoassay for early diagnosis of Taxoplasma infection. METHODS: Urine samples collected from three groups of mice infected with different doses of tachyzoites were detected for Taxoplasma circulating antigen (TCA) by dot-ELISA using HRP-SPA as a second antibody (SPA-ELISA). RESULTS: Taxoplasma circulating antigens were detected in all three groups of infected mice in contrast with the normal control group. Taxoplasma circulating antigen was detected on d6 and d3 after infection in mice of light- and moderate-infection groups, respectively. CONCLUSION: SPA-ELISA is a simple and convenient method for early immunodiagnosis of recent Taxoplasma infection.

Detection of Toxoplasma gondii tachyzoites and bradyzoites in blood, urine, and brains of infected mice.

Different techniques for identifying Toxoplasma gondii were compared. PCR was used to amplify part of the major surface antigen P30 gene of T. gondii. Amplified-DNA detection with the DNA enzyme immunoassay (PCR-DEIA) was more sensitive than ethidium bromide staining after agarose gel electrophoresis and as sensitive as nested PCR. PCR-DEIA, using common enzyme-linked immunosorbent assay (ELISA) methods, avoids agarose gel electrophoresis for the identification of amplified products. T. gondii can also be detected with equal sensitivity in infected fibroblasts, but only after at least 8 days of cell culture. PCR-DEIA is thus recommended because of its sensitivity and convenience for detecting early parasitemia in the surveillance of toxoplasmosis among pregnant women and immunocompromised hosts. The courses of infection in mice infected with two strains of T. gondii were compared. Tachyzoites of the virulent strain T. gondii RH, killing the host in 4 days, were identified in urine specimens and blood samples of mice 24 to 94 h after inoculation but not in brains, but no antibodies were detected. After intraperitoneal inoculation with cysts of the low-level virulence Beverley strain of T. gondii, parasites were identified in blood samples 4 days later and up to 17 days (but not in urine specimens) and in the brain from day 6 through day 525. By ELISA, high antibody titers were found from day 11 to day 525, with parasitemia preceding the appearance of antibodies. The usefulness of PCR-DEIA tests in conjunction with the search for circulating antibodies for the early diagnosis of toxoplasmosis in humans is discussed.

[Detection of circulating antigen in urine from mice infected with Toxoplasma tachyzoites].

Urine samples collected from three groups of mice infected experimentally with different numbers doses of Toxoplasma trophozoites were detected for the presence of Toxoplasma circulating antigen by using fast ELISA. The results showed that presence of circulating antigen in all three infected groups of mice in comparison with the normal control group. Toxoplasma circulating antigen was detected on days 5, 4 and 3 after infection in light-, moderate- and heavy-infection groups, respectively. The concentration of circulating antigen was on a parallel with the duration of infection. Western blot analysis of the Toxoplasma circulating antigen in urine revealed the existence of seven specific bands with molecular weights of 75, 67, 55, 43, 30, 28 and 22 kDa.