Institute of Parasitology and Centre for Host-Parasite Interactions, Macdonald College, McGill University, Ste Anne de Bellevue, QC, Canada.
Department of Medical Parasitology, Faculty of Medicine, Al-Azhar University, Cairo, Egypt.
Parasit Vectors. 2017 Oct 18;10(1):496. doi: 10.1186/s13071-017-2420-1.
Diagnosis of soil-transmitted helminths (STHs) has traditionally relied on stool microscopy, which has a number of critical deficiencies. Molecular diagnostics are powerful tools to identify closely related species, but the requirement for costly equipment makes their implementation difficult in low-resource or field settings. Rapid, sensitive and cost-effective diagnostic tools are crucial for accurate estimation of STH infection intensity in MDA programmes in which the goal is to reduce morbidity following repeated rounds of chemotherapy.
In this study, colourimetric isothermal assays were developed using SmartAmp2 primer sets and reagents in loop-mediated amplification (LAMP) assays. Species-specific primer sets, designed on a specific target sequence in the β-tubulin gene, were used to identify Necator americanus, Trichuris trichiura and Ascaris lumbricoides. After initial optimization on control plasmids and genomic DNA from adult worms, assays were evaluated on field samples. Assays showed high sensitivity and demonstrated high tolerance to inhibitors in spiked faecal samples. Rapid and sensitive colourimetric assays were successfully developed to identify the STHs in field samples using hydroxy napthol blue (HNB) dye.
Rapid and simple colourimetric diagnostic assays, using the SmartAmp2 method, were developed, with the potential to be applied in the field for detection of STH infections and the estimation of response to treatment. However, further validation on large numbers of field samples is needed.
土壤传播性蠕虫（STH）的诊断传统上依赖粪便显微镜检查，但这种方法存在许多严重的缺陷。分子诊断是识别密切相关物种的有力工具，但由于需要昂贵的设备，在资源匮乏或野外环境中实施这些方法具有一定难度。快速、敏感和具有成本效益的诊断工具对于准确估计 MDA 方案中 STH 感染强度至关重要，因为 MDA 方案的目标是在多次化疗后降低发病率。
在这项研究中，使用 SmartAmp2 引物组和环介导扩增（LAMP）检测试剂开发了比色等温扩增检测法。基于β-微管蛋白基因中的特定目标序列设计了种特异性引物组，用于鉴定美洲钩虫、鞭虫和蛔虫。在对对照质粒和成虫基因组 DNA 进行初步优化后，在现场样本上评估了检测法。检测法显示出较高的灵敏度，并在加标粪便样本中对抑制剂具有较高的耐受性。成功开发了快速灵敏的比色检测法，使用羟基萘酚蓝（HNB）染料来鉴定现场样本中的 STH。
使用 SmartAmp2 方法开发了快速简便的比色诊断检测法，有望在野外用于检测 STH 感染并估计治疗效果。然而，需要进一步在大量现场样本上进行验证。
