Identification of candidate antigens from adult stages of Toxocara canis for the serodiagnosis of human toxocariasis

Abstract

In the present work, we identified adult Toxocara canis antigens through sodium dodecyl sulfate-polyacrylamide gel electrophoresis for potential use in human toxocariasis immunodiagnosis. The sensitivity and specificity of several semi-purified antigens, as well as their cross-reactivity with other parasitic infections, were assessed by IgM and IgG-enzime linked immunosorbent assay. Whilst we found that the crude extract of the parasite presented limited sensitivity, specificity and high cross-reactivity against other parasites, we identified 42, 58, 68 and 97-kDa semi-purified antigens as the most promising candidates for immunodiagnosis. Moreover, the 58 and 68-kDa antigens presented the lowest IgM cross-reactivity. When tested as a combination, a mixture of the 58 and 68-kDa antigens presented 100% sensitivity and specificity, as well as minor cross-reactivity. Although the combination of the 42, 58, 68 and 97-kDa antigens presented 100% sensitivity at a dilution of 1:40, the low specificity and high cross-reactivity observed suggested a limited use for diagnostic purposes. Our data suggested that the 58 and 68-kDa antigens might be most suitable for the immunodiagnosis of human toxocariasis.

Toxocariasis: a silent threat with a progressive public health impact

“… Unavailable for discrimination of past and recent infection Recombinant antigens Less cross-reactivity with antibodies from other helminth infections in endemic regions where poly-parasitism is common, in contrast to TES-Ag [ 14 ]. rTES-30, rTES-26 or rTES-120 [ 67 , 136 ] Recommended as the best option for diagnosis of human toxocariasis [ 67 , 137 – 139 ]. Nucleic acid amplification [ 9 ] RFLP Requires a large quantity of genomic DNA, which is not readily available for parasites of small sizes, particularly larvae and eggs ITS-1 ITS-2 High sensitivity and specificity; useful for species identification and quantification of parasite burden Technically demanding, requires skilled laboratory technicians RAPD Low reproducibility and specificity; cannot distinguish between eggs of T. canis and T. cati .…”

Reactivity of recombinant Toxocara canis TES‐30/120 in experimentally infected mice

“…Most studies that involve the use the recombinant antigens in toxocariasis diagnosis use both rTES‐30 and rTES‐120 as predicted models with high antigenic potential. Albeit human samples have shown a potential response for rTES‐30 and rTES‐120, as published by Mohamad et al (2009), Zahabiun et al (2015), Fong et al (2003) and Peixoto et al (2011), in mouse, the animal model for this disease, rTES‐120 did not prove itself as an antigenic protein (Figure ) . Sperotto et al (2017) published a proteomic analysis of in vitro T. canis larvae and identified both proteins, rTES‐30 and rTES‐120, indicating that larvae cultivated in vitro express them both .…”

“…In humans, recombinant proteins have been widely studied for toxocariasis diagnostic, with sensibility and specificity around 70%‐100% and 92%‐100%, respectively . By contrast, there is scarce information in animal studies, especially in mice, where the standard diagnostic methods are still utilized to study various infection paradigms, such as humoral immune response …”

Human toxocariasis – A look at a neglected disease through an epidemiological ‘prism’

“…In humans, the diagnosis of Toxocara infection or toxocariasis is usually achieved by serology, occasionally combined with imaging methods to detect granulomata (with encapsulated larvae) in tissues. Methods such as enzyme-linked immunosorbent assays (ELISAs), utilising Toxocara [canis] excretory/secretory (TES) antigens (de Savigny et al, 1979), have been used for seroprevalence studies of Toxocara infection/exposure in humans (Elefant et al, 2016;Jin et al, 2013;Magnaval et al, 1991;Mohamad et al, 2009;Noordin et al, 2005;Peixoto et al, 2011;Roldán et al, 2015;Watthanakulpanich et al, 2008; Table 1). ELISA results can be verified by Western blotting to reduce false-positive results relating to background-or cross-reactivity with other parasites (Moreira et al, 2014).…”

“…The sensitivity and specificity of serodiagnostic tests for human toxocariasis depend on the antigens used (e.g., somatic products from T. canis larvae, native or recombinant TES products, or either glycan antigens or deglycosylated TES), the isotype of antibodies (e.g., IgG, IgG subclass or IgM) being detected and the extent of optimisation of the assays (Peixoto et al, 2011;Roldán et al, 2015). Thus, numerous ELISAs have been established and evaluated including those listed in Table 1.…”