Chimeric Antigens ofToxoplasma gondii: Toward Standardization of Toxoplasmosis Serodiagnosis Using Recombinant Products

Abstract

We have evaluated the diagnostic utility of six antigenic regions of the Toxoplasma gondii MIC2, MIC3, M2AP, GRA3, GRA7, and SAG1 gene products, assembled in recombinant chimeric antigens by genetic engineering, in order to replace the soluble, whole-cell tachyzoite extract in serological assays. Serum samples from 100 adults with acquired T. gondii infection and from 30 infants born to mothers with primary toxoplasmosis contracted during pregnancy, of whom 20 were congenitally infected, were included. Immunoglobulin G (IgG) and IgM antibodies against epitopes carried by chimeric antigens were measured by performing parallel enzyme immunoassays (recombinant enzyme-linked immunosorbent assays [Rec-ELISAs]), and the results obtained by standard commercial assays with the whole-cell Toxoplasma antigen and assays with the chimeric antigens were compared. Our results demonstrate that IgG and IgM Rec-ELISAs with individual chimeric antigens have performance characteristics comparable to those of the corresponding commercial assays. Furthermore, we show that IgM-capture assays based on chimeric antigens improve the ability to diagnose congenital toxoplasmosis postnatally compared with the ability to diagnose congenital toxoplasmosis by the use of standard assays. The use of recombinant chimeric antigens is effective in distinguishing T. gondii-infected individuals from T. gondii-uninfected individuals and shows that immunoassays based on recombinant products could provide the basis for standardized commercial tests for the serodiagnosis of toxoplasmosis.Human infection by Toxoplasma gondii is generally asymptomatic and induces a self-limiting disease. In contrast, primary T. gondii infection acquired during gestation can be transmitted to the fetus through the placenta and may cause miscarriage, permanent neurological damage, premature birth, and visual impairment (15,29,32). Toxoplasmosis during gestation represents a formidable task for the clinician due to its subclinical course in the majority of pregnant women and the unpredictable long-term outcome of congenital infection (11,16,32). To implement suitable therapies in good time and to avoid neonatal malformations or reduced eyesight in newborns, it is essential to establish when the primary infection has been acquired in the mother and to determine if vertical transmission to the fetus has occurred.The diagnosis of T. gondii infection can be established by detecting parasite-specific DNA sequences in body fluids and tissues or Toxoplasma-specific immunoglobulins in sera from infected individuals (2,20,30). Serological methods are generally preferred, since the sensitivities and the specificities of other methods can be affected by the appropriateness of sample handling and shipping and storage conditions. Furthermore, parasitemia may be of short duration, further limiting the value of detection of DNA in amniotic fluid and peripheral blood.The diagnosis of toxoplasmosis by serological methods routinely uses immunoenzymatic assays to detect the presence of the var...

Serology using rROP2 antigen in the diagnostic of toxoplasmosis in pregnant women

“…Several other recombinant antigens give similar results, but they were not included in the commercial assays. As the results were similar but weaker than for whole extract ELISA, attempts have been made to introduce a mixture of several recombinant antigens or chimeric proteins composed of several epitopes of diverse T. gondii proteins (BUFFOLANO et al, 2005;BEGHETTO et al, 2006;PIETKIEWICZ et al, 2007). However, all attempts have given similar results without improving upon the results of the whole extract ELISA.…”

Seroprevalence of Toxoplasma gondii in southern districts of Tamil Nadu using IgG-ELISA

“…To date, many serologic kits commercially available for diagnosis of T. gondii infection are based primarily on the use of whole extracts of tachyzoites (Beghetto et al 2006). However, these extracts contained large amount of proteins and other macromolecules; each of them can influence the results of the test.…”

“…Clinically, however, the diagnosis of toxoplasmosis is difficult because the clinical signs and symptoms are varied and mimics those of a variety of other diseases (Hurt and Tammaro 2007). To date, many diagnostic methods including pathogenic, immunologic, and molecular techniques have been used for detection of T. gondii infection; of them, serologic methods such as dye test, indirect hemagglutination test, latex agglutination test, indirect fluorescent antibody test, and enzyme-linked immunosorbent assay (ELISA) are most commonly used (Beghetto et al 2006;Taylor et al 1990).…”

Human Antibody Response to Thioredoxin Peroxidase-1 and Tandem Repeat Proteins as Immunodiagnostic Antigen Candidates for Schistosoma japonicum Infection

“…Fusion proteins have been shown effective in the serodiagnosis of other parasites. 35,36 On the basis of the ELISA results, Sj7TR is a promising candidate antigen for diagnostic purposes, although GeneDB defined it only as an expressed protein. Immunolocalization and expression profiling therefore of this schistosome antigen should be done to understand its role for the parasite.…”