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利用内转录间隔区1 PCR-RFLP技术对伊朗不同地区人类皮肤利什曼病和犬内脏利什曼病病原体进行分子鉴定

Molecular Identification of Agents of Human Cutaneous Leishmaniasis and Canine Visceral Leishmaniasis in Different Areas of Iran Using Internal Transcribed Spacer 1 PCR-RFLP.

作者信息

Teimouri Aref, Mohebali Mehdi, Kazemirad Elham, Hajjaran Homa

机构信息

Department of Medical Parasitology and Mycology, Tehran University of Medical Sciences, Tehran, Iran.

Students Scientific Research Center, Tehran University of Medical Sciences, Tehran, Iran.

出版信息

J Arthropod Borne Dis. 2018 Jun 13;12(2):162-171. eCollection 2018 Jun.

DOI:
PMID:30123810
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6091798/
Abstract

BACKGROUND: Leishmaniasis is a major medical health problem and distributes in nearly half of 31 provinces of Iran. We aimed to identify cutaneous and visceral spp. isolated from infected humans and domestic dogs in various regions of Iran, 2010-2013. METHODS: DNA was extracted from 108 lesion exudate samples of suspected patients to cutaneous leishmaniasis and nine liver and spleen aspirates of infected dogs cultured in RPMI-1640 and amplified using partial sequence of ITS1 gene. The PCR amplicons were digested using III endonuclease enzyme and used in restriction fragment length polymorphism (RFLP) assay. Then, 48 amplicons representing various hosts were sequenced and compared to sequences from GenBank databases using BLAST. RESULTS: PCR-RFLP analysis showed that 60 and 48 CL patients were infected by and , respectively. From nine canine visceral leishmaniasis (CVL) isolates, eight isolates were identified as and one as . The greatest similarity of 95.7% in ITS1 region was seen between and . Furthermore, the lowest similarity with 65.7% was seen between and . Intra-species comparison of ITS1 region in and isolates were showed 100%, 98.2% and 72.4 % similarities, respectively. CONCLUSION: PCR-RFLP based on ITS1 region is an appropriate method to distinguish three spp. of , and . In intra-species comparison of ITS1 region, genotypic variations showed that isolates were more heterogeneous than and isolates.

摘要

背景：利什曼病是一个主要的医疗卫生问题，分布于伊朗31个省份中近一半的地区。我们旨在鉴定2010 - 2013年期间从伊朗不同地区受感染的人类和家犬中分离出的皮肤利什曼原虫和内脏利什曼原虫种类。 方法：从108例疑似皮肤利什曼病患者的皮损渗出液样本以及9例感染犬的肝脏和脾脏穿刺液中提取DNA，在RPMI - 1640培养基中培养，使用ITS1基因的部分序列进行扩增。PCR扩增产物用III型内切酶消化，并用于限制性片段长度多态性（RFLP）分析。然后，对代表不同宿主的48个扩增产物进行测序，并使用BLAST与GenBank数据库中的序列进行比较。 结果：PCR - RFLP分析表明，60例和48例皮肤利什曼病患者分别感染了 和 。在9例犬内脏利什曼病（CVL）分离株中，8株被鉴定为 ，1株被鉴定为 。在 和 之间，ITS1区域的相似度最高，为95.7%。此外，在 和 之间，相似度最低，为65.7%。 和 分离株ITS1区域的种内比较分别显示出100%、98.2%和72.4%的相似度。 结论：基于ITS1区域的PCR - RFLP是区分 、 和 三种利什曼原虫的合适方法。在ITS1区域的种内比较中，基因型变异表明 分离株比 和 分离株更具异质性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/06e70d81d3e6/JAD-12-162-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/8380cb5ea763/JAD-12-162-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/ca0c9f4f6b83/JAD-12-162-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/db645b56947f/JAD-12-162-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/8c769e8f7fe9/JAD-12-162-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/06e70d81d3e6/JAD-12-162-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/8380cb5ea763/JAD-12-162-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/ca0c9f4f6b83/JAD-12-162-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/db645b56947f/JAD-12-162-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/8c769e8f7fe9/JAD-12-162-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91cf/6091798/06e70d81d3e6/JAD-12-162-g005.jpg

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本文引用的文献

[1]
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Identification and phylogenetic relationship of Iranian strains of various Leishmania species isolated from cutaneous and visceral cases of leishmaniasis based on N-acetylglucosamine-1-phosphate transferase gene.

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Molecular identification and polymorphism determination of cutaneous and visceral leishmaniasis agents isolated from human and animal hosts in Iran.

Biomed Res Int. 2013-10-28

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