Reactivity of Synthetic SAG1 (p30) Peptide Sequences with RH, S273 and Beverley Strain-Induced Anti-<i>Toxoplasma gondii</i> Antibodies

Abstract

Objectives: We compared the reactivity of IgG1 and IgG2a antibodies in mouse sera after infection with virulent RH and low-virulent S273 and Beverley strains of Toxoplasma gondii against RH SAG1 recombinant p30 (rp30) and synthetic SAG1 peptides. Methods: Infected mouse serum samples were collected 9 days after infection, and the level of total IgG, IgG1 and IgG2a against the RH SAG1 rp30 protein and twenty peptides of the RH SAG1 protein were assessed. The glycosylphosphatidylinositol (GPI) modification site, the hydrophilic-hydrophobic structure, the transmembrane region and the secondary structure of the SAG1 sequence of virulent and low-virulent strains were analyzed using software. Results: The virulent strain-infected mice produced a higher level of IgG1 but a lower IgG2a against the rp30 antigen, while the low-virulent strain-infected mice produced a higher level of IgG2a than the virulent strain. The difference in the secondary structure of SAG1 protein between the virulent and low-virulent strain was largely confined to amino acid positions 291–336, showing mutations and GPI anchor site. Conclusion:The difference in the reactivity of IgG against the rp30 antigen and synthetic peptides between virulent and low-virulent strains points to the importance of the primary and secondary structure assumed by antigens in the activation of Th cells and, subsequently, in the induction of IgG and its subclasses.

High diagnostic efficiency of IgM-ELISA with the use of multiple antigen peptides (MAP1) from T. gondii ESA (SAG-1, GRA-1 and GRA-7), in acute toxoplasmosis

“…In the initial evaluation of the different peptide combinations by the assay, we observed that when SAG-1 was present, as in the MAP1, the sensitivity and specificity in the detection of IgG, IgM and IgA antibodies in different toxoplasmosis phases was higher compared with those without SAG-1, as MAP4 (Table 1). Our results are consistent with the results of other investigators who have found that SAG-1 improves the sensitivity and specificity of serological assays 19,25 . In the study of other peptide combinations such as, MAP2, 3 and 4, the assay presented low diagnostic performance, and the acute/recent, non-acute/recent and chronic phase could not be distinguished as in the assay using MAP1.…”

“…However, current laboratory practice using highly sensitive automated equipment has led to problems with the detection of parasite specific IgG and IgM antibodies 12,33 . Several authors have shown that IgM to T. gondii can be detected for a long period after the primary infection and false positivities with IgG to T. gondii have been reported which are non-significant and erroneously interpreted as positive results indicating acute infection and protection 19 . Research and development have continued to advance and new parameters have been developed for the diagnosis of toxoplasmosis in its different phases, such as the avidity test of IgG antibodies and detection of the parasite by a molecular method such as polymerase chain reaction (PCR).…”

“…The Fmoc protecting group was removed by using a mild base treatment with 2% piperidine 1,8-diazabicyclo [5.4.0]undec-7-ene (DBU) in dimethyl sulfoxide-70% domethylformamide with in situ activation. The peptide sequence chosen for SAG-1 is based in KATO et al 19 ; for GRA-1 in BEGHETTO et al 3 ; and for GRA-7 in SOUSA et al 31 .…”

Peptide Microarray Analysis ofIn Silico-Predicted Epitopes for Serological Diagnosis of Toxoplasma gondii Infection in Humans

“…Several experimental approaches have been used to identify protein regions and epitopes suitable for the serological diagnosis of T. gondii, including phage display of cDNA libraries, epitope mapping, and reactivity with monoclonal antibodies (5,11,26,29,41,49). For this study, a bioinformatic approach was used for in silico prediction of 20 B-cell linear epitopes on the T. gondii antigenic proteins GRA1, GRA2, GRA4, and MIC3.…”

“…For this study, a bioinformatic approach was used for in silico prediction of 20 B-cell linear epitopes on the T. gondii antigenic proteins GRA1, GRA2, GRA4, and MIC3. These 20 novel peptides and 18 peptides reported in studies previously published by others (6,11,25,26) were printed into a peptide microarray and validated by analyzing a large number of well-characterized human serum samples.…”