Abstract：
The classical dermatophytes diagnosis is based on mycological culture and microscopy observation both human and animal hair, skin, and nail samples. The aim of this work was to develop the new in-house real-time PCR with pan-dematophyte reaction for detection and identification of the main dermatophytes directly from hair samples, providing a simple and rapid diagnosis of dermatophytosis in dogs and cats. An in-house SYBR-Green real-time PCR was designed and used for detecting a DNA fragment encoding chitin synthase 1 (CHS1). A total of 287 samples were processed by culture, microscopic examination with KOH 10%, and real-time PCR (qPCR) analysis. Melting curve analysis of the CHS1 fragment revealed to be reproducible, showing a single distinct peak for each species of dermatophyte, namely Trichophyton mentagrophytes, T. verrucosum, Microsporum canis, and Nannizzia gypsea (formerly M. gypseum). Then, out of the 287 clinically suspected cases of dermatophytosis, 50% were positive for dermatophytes by qPCR, 44% by mycological culture, and 25% by microscopic examination. Microsporum canis was identified in 117 samples tested by culture and 134 samples tested by qPCR, followed by N. gypsea in 5 samples (either tested by culture or qPCR) and T. mentagrophytes detected in 4 and 5 samples when tested by culture or qPCR, respectively. Overall, qPCR allowed the diagnosis of dermatophytosis in clinical samples. The results suggest this newly proposed in-house real-time PCR assay can be used as alternative diagnosis and rapid identification of dermatophytes frequently associated to clinical hair samples of dogs and cats.
The aim of this work was to develop a molecular detection strategy for dermatophytes by SYBR-Green real-time PCR of hair samples from animals. The melting curve analysis of the CHS1 fragment revealed to be reproducible, showing a single distinct peak for distinct dermatophyte species and allowed the diagnosis of dermatophytosis in dogs and cats caused mainly by Trichophyton mentagrophytes, Microsporum sp., and Nannizzia gypsea).
The aim of this work was to develop a molecular detection strategy for dermatophytes by SYBR-Green real-time PCR of hair samples from animals. The melting curve analysis of the CHS1 fragment revealed to be reproducible, showing a single distinct peak for distinct dermatophyte species and allowed the diagnosis of dermatophytosis in dogs and cats caused mainly by Trichophyton mentagrophytes, Microsporum sp., and Nannizzia gypsea).
摘要：
经典的皮肤癣菌诊断基于真菌学培养和显微镜观察人和动物毛发，皮肤,和指甲样本。这项工作的目的是开发新的内部实时PCR与泛脱真菌反应，用于直接从头发样品中检测和鉴定主要的皮肤癣菌，为狗和猫的皮肤癣菌病提供简单快速的诊断。设计了一种内部SYBR-Green实时PCR，用于检测编码几丁质合酶1（CHS1）的DNA片段。共有287个样本经过培养处理,用10%的KOH进行显微镜检查，和实时PCR(qPCR)分析。CHS1片段的熔解曲线分析显示是可重复的，每个皮肤癣菌都有一个独特的峰，即毛癣菌，T.疣,犬小孢子菌,和Nannizziagypsea（以前的M.gypseum）。然后，在287例临床疑似病例中，通过qPCR，50%的皮肤癣菌呈阳性，44％的真菌学文化，和25%的显微镜检查。在117个通过培养检测的样品和134个通过qPCR检测的样品中鉴定出犬小孢子菌，其次是5个样品（通过培养或qPCR测试）和4个和5个样品通过培养或qPCR测试时，分别。总的来说,qPCR允许在临床样品中诊断皮肤癣菌病。结果表明，这种新提出的内部实时PCR检测方法可用作经常与狗和猫的临床毛发样品相关的皮肤癣菌的替代诊断和快速鉴定。
这项工作的目的是通过来自动物的毛发样品的SYBR-Green实时PCR开发皮肤癣菌的分子检测策略。CHS1片段的熔解曲线分析显示是可重复的，显示不同的皮肤癣菌物种的单个明显峰，并允许诊断主要由毛癣菌引起的狗和猫的皮肤癣菌病，微孢子菌。,和Nannizziagypsea）。
这项工作的目的是通过来自动物的毛发样品的SYBR-Green实时PCR开发皮肤癣菌的分子检测策略。CHS1片段的熔解曲线分析显示是可重复的，显示不同的皮肤癣菌物种的单个明显峰，并允许诊断主要由毛癣菌引起的狗和猫的皮肤癣菌病，微孢子菌。,和Nannizziagypsea）。
