Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Cellular and Molecular Research Center, Qom University of Medical Sciences, Qom, Iran.
Acta Parasitol. 2024 Mar;69(1):889-897. doi: 10.1007/s11686-024-00821-2. Epub 2024 Mar 12.
Strongyloides stercoralis is a parasite with special characteristics presenting it as a unique nematode. Iran is an endemic area for S. stercoralis. In this study, nested-qPCR-high resolution melting (HRM) technology was applied on some human isolates of S. stercoralis from this country by focusing on evolutionary genetics analysis.
Twelve human isolates of S. stercoralis were collected from four endemic provinces of Iran. Genomic DNA was extracted from a single filariform larva for every isolate. Using specific primers targeting partial regions in cox1 gene, nested-qPCR-HRM was performed and melting-curve profiles were analyzed alongside the evaluation of genetic proximity and phylogenetic analysis using MEGA7 and DnaSP5 software.
The melting temperature (T) values of the isolates were 77.9 °C-78.3 °C. All isolates from Guilan, Mazandaran, and Khouzestan Provinces shared T values of 78.2 °C to 78.3 °C, while the isolates from Hormozgan Province showed T values of 77.9 °C, 78.0 °C, and 78.1 °C. The phylogenetic tree illustrated that the sequences of the current study included nine haplotypes. Tajima's D index analyses showed that cox1 gene in S. stercoralis isolates was negative (Tajima's D = 0.27).
The isolates were divided into five temperature groups. Although HRM assay compared to PCR sequencing identified more limited genetic changes, it revealed that the mean of T of the isolates from Hormozgan Province was lower than those of other provinces and represented specific haplotypes for this geographical region on the phylogenetic tree.
粪类圆线虫是一种具有特殊特征的寄生虫，表现为一种独特的线虫。伊朗是粪类圆线虫的地方性流行区。在这项研究中，通过聚焦进化遗传学分析，应用巢式 qPCR-高分辨率熔解（HRM）技术对来自该国的一些粪类圆线虫人体分离株进行了研究。
从伊朗四个地方性流行省份收集了 12 个人类粪类圆线虫分离株。从每个分离株的单个丝状幼虫中提取基因组 DNA。使用针对 cox1 基因部分区域的特异性引物，进行巢式 qPCR-HRM，并分析熔解曲线图谱，同时使用 MEGA7 和 DnaSP5 软件评估遗传接近度和系统发育分析。
分离株的熔点（T）值为 77.9°C-78.3°C。来自吉兰、马赞达兰和胡齐斯坦省的所有分离株的 T 值为 78.2°C-78.3°C，而来自霍尔木兹甘省的分离株的 T 值为 77.9°C、78.0°C 和 78.1°C。系统发育树表明，当前研究的序列包括 9 种单倍型。Tajima 的 D 指数分析表明，粪类圆线虫分离株的 cox1 基因呈阴性（Tajima 的 D=0.27）。
分离株分为五个温度组。虽然 HRM 分析与 PCR 测序相比，鉴定出的遗传变化有限，但它表明霍尔木兹甘省分离株的 T 平均值低于其他省份，并且在系统发育树上代表了该地理区域的特定单倍型。
