Zakeri Hamideh, Shokohi Tahereh, Badali Hamid, Mayahi Saba, Didehdar Mojtaba
Student Research Committee, Faculty of Medicine, Mazandaran University of Medical Sciences, Sari, IR Iran.
Invasive Fungi Research Center (IFRC), Mazandaran University of Medical Sciences, Sari, IR Iran ; Department of Medical Mycology and Parasitology, Mazandaran University of Medical Sciences, Sari, IR Iran.
Jundishapur J Microbiol. 2015 Jul 27;8(7):e19107. doi: 10.5812/jjm.19107v2. eCollection 2015 Jul.
The high degree of phenotypic similarity among Trichophyton species makes their identification difficult.
The current study aims to establish the use of rolling circle amplification (RCA) based on internal transcribed spacer ribosomal DNA (ITS rDNA) as a powerful, simple, and rapid procedure for distinguishing closely related organisms, and specifically to identify Trichophyton species, which cause human and animal disorders.
A total of sixty-one isolates belonging to three species of Trichophyton were identified to the species level based on microscopic and macroscopic examinations and their ITS rDNA regions were sequenced. Three specific circular oligonucleotide probes targeting the ITS1 and ITS2 regions were designed to differentiate Trichophyton rubrum, T. mentagrophytes, and T. tonsurans.
Of the 61 putative Trichophyton clinical isolates, 52 were identified to the species level. The most common species was T. mentagrophytes var. interdigitale (31 isolates), followed by T. rubrum (11 isolates), T. tonsurans (9 isolates), and T. violaceum (1 isolates); moreover, 9 isolates were identified as non-Trichophyton species. The RCA method correctly identified four Trichophyton species and was 100% specific for each species. Neither cross-reaction between the examined species of Trichophyton nor false positive or false negative results were observed.
Species identification of Trichophyton is crucially important for epidemiological and phylogenetic purposes and for genotype delineation. RCA based on ITS polymorphisms can be used to generate identification barcodes and as an alternative to DNA sequencing; it is a very fast, specific, and economical tool for species identification.
毛癣菌属物种之间高度的表型相似性使得它们的鉴定变得困难。
本研究旨在建立基于内转录间隔区核糖体DNA（ITS rDNA）的滚环扩增（RCA）方法，作为一种强大、简单且快速的程序，用于区分密切相关的生物体，特别是鉴定引起人和动物疾病的毛癣菌属物种。
基于显微镜和宏观检查，将总共61株属于三种毛癣菌的分离株鉴定到种水平，并对其ITS rDNA区域进行测序。设计了三种靶向ITS1和ITS2区域的特异性环状寡核苷酸探针，以区分红色毛癣菌、须癣毛癣菌和断发毛癣菌。
在61株假定的毛癣菌临床分离株中，52株被鉴定到种水平。最常见的物种是指间型须癣毛癣菌（31株分离株），其次是红色毛癣菌（11株分离株）、断发毛癣菌（9株分离株）和紫色毛癣菌（1株分离株）；此外，9株分离株被鉴定为非毛癣菌属物种。RCA方法正确鉴定了四种毛癣菌物种，并且对每个物种具有100%的特异性。在所检测的毛癣菌物种之间未观察到交叉反应，也未出现假阳性或假阴性结果。
毛癣菌的物种鉴定对于流行病学和系统发育目的以及基因型划分至关重要。基于ITS多态性的RCA可用于生成鉴定条形码并作为DNA测序的替代方法；它是一种非常快速、特异且经济的物种鉴定工具。
