Dabirzadeh Mansour, Hashemi Mohammad, Maroufi Yahya
Department of Parasitology and Mycology, Zabol University of Medical Sciences, Zabol, IR Iran.
Jundishapur J Microbiol. 2016 May 31;9(6):e33498. doi: 10.5812/jjm.33498. eCollection 2016 Jun.
Zoonotic cutaneous leishmaniasis (ZCL) is polymorphic disease that may show various clinical manifestations.
This study investigates the determination of genetic variation within the species of Leishmania major isolates from new cases in Chabahar, a port city in Southeast Iran (situated at the Iran-Pakistan border). Migration in this region indicates that leishmaniasis is spreading gradually, and a new micro-habitat focus appears each year.
A variety of nucleic acid detection methods that target both DNA and RNA have been developed. The restriction fragment length polymorphism analysis of amplified internal transcribed spacer 1 with polymerase chain reaction (ITS1-RFLP PCR) assay is a multipurpose tool for the diagnosis of Leishmania from clinical samples and for enabling the determination of the infecting Leishmania species. The goal of this study was the identification of species based on ITS1-RFLP in the ribosomal operon of L. major from clinically different forms of ZCL amplified by PCR, followed by the digestion of the PCR product with restriction enzymes. The profiles were observed and visualized in agarose gel under UV light. We used direct smears to identify the parasites. While taking the smear, samples were collected for culture or direct PCR. We used the PCR-RFLP assay of the ITS1 genes for direct identification of Leishmania species in 24 out of 33 suspected patients. PCR-ITS1 amplification was done on the 24 samples confirmed by culture via growth and parasitological methods.
Of the 24 isolates, 21 had 350 bp bands (87.5%) and three had 450 bp bands (12.5%). After using the restriction enzyme, banding patterns including fragments of 210 and 140 bp for L. major were detected in 19 cases.
The L. major species causing ZCL in Chabahar have limited genetic variation. There seems to be little manifestation of diversity between these lesions as a new focus of disease, and new micro-habitats for the disease are appearing in parts of this region.
人兽共患皮肤利什曼病（ZCL）是一种多形性疾病，可能表现出多种临床表现。
本研究调查了来自伊朗东南部港口城市恰巴哈尔（位于伊朗 - 巴基斯坦边境）新病例的硕大利什曼原虫分离株物种内的遗传变异。该地区的人口迁移表明利什曼病正在逐渐蔓延，并且每年都会出现新的微生境疫源地。
已开发出多种针对DNA和RNA的核酸检测方法。聚合酶链反应扩增内部转录间隔区1的限制性片段长度多态性分析（ITS1 - RFLP PCR）检测是一种多用途工具，可用于从临床样本中诊断利什曼原虫，并确定感染的利什曼原虫物种。本研究的目标是基于通过PCR扩增的临床不同形式的ZCL的硕大利什曼原虫核糖体操纵子中的ITS1 - RFLP来鉴定物种，然后用限制性酶消化PCR产物。在紫外光下观察琼脂糖凝胶中的条带并进行可视化。我们使用直接涂片来鉴定寄生虫。涂片时，收集样本用于培养或直接PCR。我们对33例疑似患者中的24例使用ITS1基因的PCR - RFLP检测直接鉴定利什曼原虫物种。对通过培养生长和寄生虫学方法确认的24个样本进行PCR - ITS1扩增。
在24个分离株中，21个有350 bp条带（87.5%），3个有450 bp条带（12.5%）。使用限制性酶后，在19例中检测到包括硕大利什曼原虫210和140 bp片段的条带模式。
在恰巴哈尔引起ZCL的硕大利什曼原虫物种遗传变异有限。作为新的疾病疫源地，这些病变之间似乎几乎没有多样性表现，并且该地区部分地区正在出现该疾病的新微生境。
