Universidad Nacional de Salta, Sede Regional Orán, Instituto de Investigaciones de Enfermedades Tropicales (IIET), Salta, Argentina; Universidad Nacional de Salta, Facultad de Ciencias Naturales, Cátedra de Química Biológica, Salta, Argentina; Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), CCT, Salta, Argentina.
Universidad Nacional de Salta, Sede Regional Orán, Instituto de Investigaciones de Enfermedades Tropicales (IIET), Salta, Argentina.
Braz J Infect Dis. 2021 Nov-Dec;25(6):101649. doi: 10.1016/j.bjid.2021.101649. Epub 2021 Nov 10.
Describe the diagnostic characteristics of a conventional multiplex PCR for the diagnosis of S. stercoralis, N. americanus and Ancylostomas spp.
Fecal samples were collected from a cross-sectional study in Orán department, Salta province, Argentina. The stool samples were analyzed using concentration-sedimentation, Harada Mori, McMaster, and Baermann techniques. DNA was extracted from 50 mg fecal sample using the FastPrep® Spin Kit for Soil. Three pairs of primers were used for the amplification of three products of 101, 330, and 577 base pairs (bp) for S. stercoralis, N. americanus and Ancylostoma spp, respectively. The sensitivity and analytical specificity of multiplex PCR were evaluated, as well as the sensitivity and diagnostic specificity, using a composite standard and Bayesian approach.
Multiplex PCR did not present cross-reaction with other intestinal parasites, and the detection limit for multiplex PCR was between 2 and 20 pg of genomic DNA. In addition it presented a diagnostic sensitivity of 97.4% for S. stercoralis and 90.3% for hookworms with a specificity of 100% and 87.6%, respectively. PCR identified a higher proportion (p <0.01) of coinfections (15.3%) than microscopic techniques (3.5%). Also, multiplex PCR showed that there was a positive association between S. stercoralis and hookworms (odds ratio = 2.12). However, this association was due to N. americanus (odds ratio= 3.22), since no association was observed between S. stercoralis and Ancylostoma spp. Neither was an association observed between the two species of hookworms.
描述用于诊断 S. stercoralis、N. americanus 和Ancylostoma spp 的常规多重 PCR 的诊断特征。
从阿根廷萨尔塔省奥伦省的一项横断面研究中收集粪便样本。使用浓缩沉淀、Harada Mori、McMaster 和 Baermann 技术分析粪便样本。使用 FastPrep®Spin Kit for Soil 从 50mg 粪便样本中提取 DNA。使用三对引物分别扩增 101、330 和 577 个碱基对（bp）的三个产物，用于 S. stercoralis、N. americanus 和 Ancylostoma spp。使用复合标准和贝叶斯方法评估了多重 PCR 的灵敏度和分析特异性，以及敏感性和诊断特异性。
多重 PCR 与其他肠道寄生虫无交叉反应，多重 PCR 的检测限为 2 至 20pg 基因组 DNA。此外，它对 S. stercoralis 的诊断灵敏度为 97.4%，对钩虫的灵敏度为 90.3%，特异性均为 100%和 87.6%。PCR 鉴定出的合并感染比例（15.3%）高于显微镜技术（3.5%）（p<0.01）。此外，多重 PCR 显示 S. stercoralis 和钩虫之间存在正关联（优势比=2.12）。然而，这种关联归因于 N. americanus（优势比=3.22），因为在 S. stercoralis 和 Ancylostoma spp.之间未观察到关联。在两种钩虫之间也未观察到关联。
