Instituto de Ciências da Saúde, Universidade Federal da Bahia, Salvador, Bahia, Brazil.
Department of Pharmaceutical Sciences, University of Cartagena, Cartagena, Colombia.
Transbound Emerg Dis. 2022 Sep;69(5):e2994-e3006. doi: 10.1111/tbed.14650. Epub 2022 Jul 19.
Toxocariasis is an infection caused by the round worms Toxocara canis and Toxocara cati. It occurs worldwide though it is more prevalent in developing countries. For the diagnosis of toxocariasis, the most used method is the indirect enzyme-linked immunosorbent assay (indirect ELISA), based on the detection of specific antibodies using the excreted/secreted products from T. canis larvae (TES) as antigens, but it cross-reacts with several helminth infections. For this reason, there is a need to investigate species-specific immunoreactive proteins, which can be used for the development of a more sensitive and specific diagnosis. This study aims to investigate immunoreactive protein candidates to be used for the development of a more sensitive and specific diagnosis of Toxocara spp. infection in humans. We have used immunoblotting and mass spectrometry to select four Toxocara canis immunoreactive proteins that were recombinantly expressed in bacteria and evaluated as potential new diagnostic antigens (rMUC3, rTES 26, rTES32 and rCTL4). The recognition of these recombinant proteins by total serum IgG and IgG4 was assayed using the purified proteins in an isolated manner or in combination. The IgG ELISAs performed with individual recombinant antigens reached values of sensitivity and specificity that ranged from 91.7% to 97.3% and 94.0% to 97.9%, respectively. Among the analyses, the IgG4 immunoassay was proven to be more effective, revealing a sensitivity that ranged from 88.8% to 98.3% and a specificity of 97.8%-97.9%. The IgG4 ELISA was shown to be more effective and presented no cross-reactivity when using combinations of the rTES 26 and rCTL4 recombinant proteins. The combination of these two molecules achieved 100% sensitivity and specificity. The use of only two recombinant proteins can contribute to improve the current panorama of toxocariasis immunodiagnosis for, with a better optimization and reduced cost.
旋毛虫病是一种由犬旋毛虫和猫旋毛虫引起的感染。它发生在世界各地，尽管在发展中国家更为普遍。对于旋毛虫病的诊断，最常用的方法是间接酶联免疫吸附试验（间接 ELISA），该方法基于使用 T. canis 幼虫的排泄/分泌产物（TES）作为抗原检测特异性抗体，但它与几种寄生虫感染有交叉反应。因此，需要研究种特异性免疫反应蛋白，可用于开发更敏感和特异的诊断方法。本研究旨在调查免疫反应性蛋白候选物，以用于开发更敏感和特异的人类旋毛虫属感染诊断方法。我们使用免疫印迹和质谱法选择了四种犬旋毛虫免疫反应性蛋白，这些蛋白在细菌中重组表达，并作为潜在的新诊断抗原进行评估（rMUC3、rTES 26、rTES32 和 rCTL4）。使用纯化蛋白单独或组合评估了这些重组蛋白对总血清 IgG 和 IgG4 的识别。使用单个重组抗原进行的 IgG ELISA 达到了 91.7%至 97.3%和 94.0%至 97.9%的灵敏度和特异性范围。在分析中，证明 IgG4 免疫测定更有效，显示 88.8%至 98.3%的灵敏度和 97.8%-97.9%的特异性。IgG4 ELISA 显示更有效，并且在使用 rTES 26 和 rCTL4 重组蛋白组合时没有交叉反应。这两种分子的组合实现了 100%的灵敏度和特异性。仅使用两种重组蛋白可以有助于改善当前的旋毛虫病免疫诊断情况，因为优化和成本降低了。
