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Abstract：
BACKGROUND: The sensitivity of parasitological and molecular methods is unsatisfactory for the diagnosis of strongyloidiasis, and serological techniques are remaining as the most effective diagnostic approach. The present study aimed to design and produce a chimeric recombinant antigen from Strongyloides stercoralis immunoreactive antigen (SsIR) and Ss1a antigens, using immune-informatics approaches, and evaluated its diagnostic performance in an ELISA system for the diagnosis of human strongyloidiasis.
RESULTS: The coding sequences for SsIR and Ss1a were selected from GenBank and were gene-optimized. Using bioinformatics analysis, the regions with the highest antigenicity that did not overlap with other parasite antigens were selected. The chimeric recombinant antigen SsIR- Ss1a, was constructed. The solubility and physicochemical properties of the designed construct were analyzed and its tertiary structures were built and evaluated. The construct was expressed into the pET-23a (+) expression vector and the optimized DNA sequences of SsIR-Ss1a (873 bp) were cloned into competent E. coli DH5α cells. Diagnostic performances of the produced recombinant antigen, along with a commercial kit were evaluated in an indirect ELISA system, using a panel of sera from strongyloidiasis patients and controls. The physicochemical and bioinformatics evaluations revealed that the designed chimeric construct is soluble, has a molecular with of 35 KDa, and is antigenic. Western blotting confirmed the immunoreactivity of the produced chimeric recombinant antigen with the sera of strongyloidiasis patients. The sensitivity and specificity of the indirect ELISA system, using the produced SsIR-Ss1a chimeric antigen, were found to be 93.94% (95% CI, 0.803 to 0.989) and 97.22% (95% CI, 0.921 to 0.992) respectively.
CONCLUSIONS: The preliminary findings of this study suggest that the produced SsIR-Ss1a chimeric antigen shows promise in the diagnosis of human strongyloidiasis. However, these results are based on a limited panel of samples, and further research with a larger sample size is necessary to confirm its accuracy. The construct has potential as an antigen in the ELISA system for the serological diagnosis of this neglected parasitic infection, but additional validation is required.
RESULTS: The coding sequences for SsIR and Ss1a were selected from GenBank and were gene-optimized. Using bioinformatics analysis, the regions with the highest antigenicity that did not overlap with other parasite antigens were selected. The chimeric recombinant antigen SsIR- Ss1a, was constructed. The solubility and physicochemical properties of the designed construct were analyzed and its tertiary structures were built and evaluated. The construct was expressed into the pET-23a (+) expression vector and the optimized DNA sequences of SsIR-Ss1a (873 bp) were cloned into competent E. coli DH5α cells. Diagnostic performances of the produced recombinant antigen, along with a commercial kit were evaluated in an indirect ELISA system, using a panel of sera from strongyloidiasis patients and controls. The physicochemical and bioinformatics evaluations revealed that the designed chimeric construct is soluble, has a molecular with of 35 KDa, and is antigenic. Western blotting confirmed the immunoreactivity of the produced chimeric recombinant antigen with the sera of strongyloidiasis patients. The sensitivity and specificity of the indirect ELISA system, using the produced SsIR-Ss1a chimeric antigen, were found to be 93.94% (95% CI, 0.803 to 0.989) and 97.22% (95% CI, 0.921 to 0.992) respectively.
CONCLUSIONS: The preliminary findings of this study suggest that the produced SsIR-Ss1a chimeric antigen shows promise in the diagnosis of human strongyloidiasis. However, these results are based on a limited panel of samples, and further research with a larger sample size is necessary to confirm its accuracy. The construct has potential as an antigen in the ELISA system for the serological diagnosis of this neglected parasitic infection, but additional validation is required.
摘要：
背景：寄生虫学和分子学方法的敏感性不能令人满意，血清学技术仍然是最有效的诊断方法。本研究旨在设计和生产一个嵌合的重组抗原从圆圆圆线虫免疫反应性抗原（SsIR）和Ss1a抗原，使用免疫信息学方法，并评价了其在诊断人圆线虫病的ELISA系统中的诊断性能。
结果:从GenBank中选择SsIR和Ss1a的编码序列并进行基因优化。利用生物信息学分析，选择与其他寄生虫抗原不重叠的免疫原性最高的区域.嵌合重组抗原SsIR-Ss1a,是建造的。分析了设计的构建体的溶解度和理化性质，并建立和评估了其三级结构。将构建体表达到pET-23a(+)表达载体中,并将SsIR-Ss1a(873bp)的优化DNA序列克隆到感受态大肠杆菌DH5α细胞中。产生的重组抗原的诊断性能，与商业试剂盒一起在间接ELISA系统中进行评估，使用一组来自圆线虫病患者和对照组的血清。物理化学和生物信息学评估表明，设计的嵌合构建体是可溶性的，具有35KDa的分子，而且是抗原性的.Western印迹证实了所产生的嵌合重组抗原与圆线虫病患者血清的免疫反应性。间接ELISA系统的敏感性和特异性，使用产生的SsIR-Ss1a嵌合抗原，分别为93.94%（95%CI，0.803至0.989）和97.22%（95%CI，0.921至0.992）。
结论：这项研究的初步结果表明，所产生的SsIR-Ss1a嵌合抗原在人类圆线虫病的诊断中显示出希望。然而,这些结果是基于有限的一组样本，并且需要用更大的样本量进行进一步的研究来确认其准确性。该构建体在ELISA系统中具有作为抗原的潜力，用于这种被忽视的寄生虫感染的血清学诊断，但需要额外的验证。
结果:从GenBank中选择SsIR和Ss1a的编码序列并进行基因优化。利用生物信息学分析，选择与其他寄生虫抗原不重叠的免疫原性最高的区域.嵌合重组抗原SsIR-Ss1a,是建造的。分析了设计的构建体的溶解度和理化性质，并建立和评估了其三级结构。将构建体表达到pET-23a(+)表达载体中,并将SsIR-Ss1a(873bp)的优化DNA序列克隆到感受态大肠杆菌DH5α细胞中。产生的重组抗原的诊断性能，与商业试剂盒一起在间接ELISA系统中进行评估，使用一组来自圆线虫病患者和对照组的血清。物理化学和生物信息学评估表明，设计的嵌合构建体是可溶性的，具有35KDa的分子，而且是抗原性的.Western印迹证实了所产生的嵌合重组抗原与圆线虫病患者血清的免疫反应性。间接ELISA系统的敏感性和特异性，使用产生的SsIR-Ss1a嵌合抗原，分别为93.94%（95%CI，0.803至0.989）和97.22%（95%CI，0.921至0.992）。
结论：这项研究的初步结果表明，所产生的SsIR-Ss1a嵌合抗原在人类圆线虫病的诊断中显示出希望。然而,这些结果是基于有限的一组样本，并且需要用更大的样本量进行进一步的研究来确认其准确性。该构建体在ELISA系统中具有作为抗原的潜力，用于这种被忽视的寄生虫感染的血清学诊断，但需要额外的验证。
