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Introduction: Serological assays are alternative laboratory tools for the diagnosis of parasitic infections. The aim of this work was to evaluate the performance of the indirect fluorescent antibody test (IFAT) and Western blotting (WB) for the detection of IgG anti-Giardia antibodies in human sera., Methodology: Sera from individuals infected with Giardia duodenalis, other parasites or non-parasitized were selected for serological assays. Ninety-seven sera were tested by IFAT at 1:20 and 1:40 dilutions and of these, 40 samples were also analyzed by WB., Results: The sensitivity and specificity of the IFAT was 97% and 46.9% at 1:20 sera dilution, and 39.4% and 59.4% at 1:40 sera dilution. The low molecular weight polypeptides fractions of 25 kDa, 27-31 kDa and 45-55 kDa were the most frequently identified by the sera of individuals infected with G. duodenalis, along with low cross-reactivity, presenting an individual sensitivity of 42.8%, 50.0% and 57.1%, and specificity of 83.3%, 83.3% and 91.7%, respectively. The highest overall sensitivity of WB (85.7%) was based on the immunoreactivity of sera with at least one of those proteins. The concordance between the detection of G. duodenalis in feces by microscopy and the WB results was considered substantial (Kappa = 0.61)., Conclusion: Constant exposure to Giardia infection throughout a lifetime can maintain high levels of specific antibodies in serum, even without active infection. Moreover, proteins found in intestinal amoebas may hinder the serological diagnosis of giardiasis in endemic areas due to cross-reactivity, which can be partially solved using Giardia low molecular weight proteins., (© 2023. The Author(s) under exclusive licence to Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Background: Giardia duodenalis is conventionally diagnosed in fecal samples using parasitological methods. However, sensitivity is poor when only a single sample is analyzed, due to intermittent excretion of cysts in feces. Alternatively, the serum antibodies to G. duodenalis can be used for parasite diagnosis and epidemiological studies to determine previous exposure. We compared the rate of G. duodenalis infection between serum anti-Giardia IgG and IgA antibodies and fecal examination in Brazilian children., Methods: Fecal and serum samples were tested from 287 children at a clinical laboratory and from 187 children at daycare centers. Fecal samples were processed using conventional parasitological methods and coproantigen detection for Giardia diagnosis. Serum samples were tested using an in-house ELISA for detection of anti-Giardia IgG and IgA., Results: G. duodenalis was found in 8.2% (N=39) of the 474 children analyzed. The sensitivity and specificity of ELISA were 80.0% and 90.0% for IgG and 80.0% and 83.3% for IgA, respectively. The total positivity rate of anti-Giardia IgG and IgA in the sera was 13.9% (N=66) and 23.6% (N=112). The agreement between the positivity of specific antibodies and the detection of G. duodenalis in feces was moderate for ELISA-IgG, kappa index (95% CI)=0.543 (0.422-0.664), and mild for ELISA-IgA, kappa index (95% CI)=0.283 (0.162-0.404). Among the children infected with other enteroparasites, 11.6% (N=10) and 24.4% (N=21) showed reactivity to anti-Giardia IgG and to IgA, respectively. This cross-reactivity was more frequent in samples from children infected with Endolimax nana and Entamoeba coli., Conclusions: The higher frequency of specific antibody reactivity compared with G. duodenalis diagnosis in feces could reflect continuous exposure of children to G. duodenalis infection, resulting in long-lasting immunological memory and/or cross-reactivity with other intestinal amoebas.

Opportunistic infections, due to disease-related immunosuppression, constitute the major cause of death in American visceral leishmaniasis (AVL). Sera from these patients (AVL sera) non-specifically inhibit the in vitro proliferative response of normal human lymphocytes to lectins or antigens. In the present work, the mediation of this inhibition by IgG, immune complexes and low- or very low-density lipoproteins was studied. AVL serum fractions containing proteins with the molecular weight of IgG, and IgG, purified from AVL sera by anion exchange chromatography, did not suppress the lymphoproliferation. Most of the suppressive activity of AVL sera was associated with a fraction containing molecules with molecular weights above 430 kDa. This would be compatible with it being due to immune complexes and/or lipoproteins, and not to soluble IL-2 receptors as reported previously. However, neither of the two possibilities seem to be the case, as (1) depletion of immune complexes by protein-A followed by protein-G chromatographies did not affect the serum suppressive activity, (2) no correlation between immune complex contents and suppressive activities in individual sera was observed, and (3) plasma lipoproteins (VLDL and LDL), purified from AVL patients and from healthy individuals, had the same degree of immunosuppressive activity.

A semi-quantitative ELISA for complement-fixing, IgG-containing immune complexes (IC) is described. The assay is based on the insolubilization of IC by polyethyleneglycol, their capture by solid-phase anti-C3 antibodies, reaction with peroxidase-labeled anti-IgG antibodies and incubation with a chromogenic peroxidase substrate. It was markedly improved by the use of a single-step procedure which simultaneously washed and precipitated the insolubilized immune complexes. Intra-assay and inter-assay coefficients of variation were lower than 8.6 and 14.7%, respectively. As expected, higher levels of circulating immune complexes, in relation to healthy individuals, were found in patients with American visceral leishmaniasis (AVL), systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), with prevalences comparable to those described in the literature. The ELISA can be quickly assembled from reagents and plasticware widely available commercially, detects immune complexes fulfilling three different criteria and is more sensitive than a previously published method based on the same principles (detection limit for complement-sensitized aggregated IgG of 2 microg ml(-1) as compared with a detection limit above 16 microg ml(-1)).