Abstract
Toxoplasmosis, caused by infection of the protozoan parasite Toxoplasma gondii, is associated with mild disease in healthy individuals, whereas individuals with depressed immunity may develop encephalitis, neurologic disorders, and other organ diseases. Women who develop acute toxoplasmosis during pregnancy are at risk of transmitting the infection to the fetus, which may lead to fetal damage. A diagnosis is usually confirmed by measuring IgG, or IgM where it is important to determine the onset of infection. A negative IgM result essentially excludes acute infection, whereas a positive IgM test is largely uninterpretable because IgM can persist for up to 18 months after infection. To identify antigens for improved diagnosis of acute infection, we probed protein microarrays displaying the polypeptide products of 1357 Toxoplasma exons with well-characterized sera from Turkey. The sera were classified according to conventional assays into (1) seronegative individuals with no history of T. gondii infection; (2) acute infections defined by clinical symptoms, high IgM titers, and low avidity IgG; (3) chronic/convalescent cases with high avidity IgG but persisting IgM; (iv) true chronic infections, defined by high avidity IgG and no IgM. We have identified 38 IgG target antigens and 108 IgM target antigens that can discriminate infected patients from healthy controls, one or more of which could form the basis of a 'tier-1' test to determine current or previous exposure. Of these, three IgG antigens and five IgM antigens have the potential to discriminate chronic/IgM persisting or true chronics from recent acutely infected patients (a 'tier-2' test). Our analysis of the antigens revealed several enriched features relative to the whole proteome, which include transmembrane domains, signal peptides, or predicted localization at the outer membrane. This is the first protein microarray survey of the antibody response to T. gondii, and will help in the development of improved serodiagnostics and vaccines.
Highlights
IntroductionWomen that become infected for the first time during the first trimester of pregnancy may transmit infection to the fetus (congenital toxoplasmosis), causing spontaneous abortion or stillbirth, or overt symptoms in the newborn
From the ‡Division of Infectious Diseases, Department of Medicine, University of California, Irvine, CA 92697; §Ege University Medical School, Department of Parasitology, Bornova/Izmir, 35100, Turkey; ¶Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA, 90095
For PCR primer design, the genomic sequence of type II strain ME49 of T. gondii was obtained from the Toxoplasma Genomics Resource and a sequential bioinformatic filtering strategy was applied to prioritize genes targeted for cloning based on features we have found in previous studies to be enriched within seroreactive antigen sets
Summary
Women that become infected for the first time during the first trimester of pregnancy may transmit infection to the fetus (congenital toxoplasmosis), causing spontaneous abortion or stillbirth, or overt symptoms in the newborn. Routine diagnosis of toxoplasmosis is based on the detection of T. gondii-specific antibodies. The original test, developed in 1948, is the Sabin-Feldman Dye Test (DT) test for specific IgG [11]. This is a sensitive and specific bioassay in Molecular & Cellular Proteomics 10.7. A negative result in someone with clinical symptoms of toxoplasmosis requires the test is repeated after 2–3 weeks, after which an immunocompetent infected individual should seroconvert. Diagnosis based on other biomarkers or PCR based molecular diagnostic techniques are being sought for these patients [26, 27]
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