基于高分辨率熔解曲线的常见五种皮肤癣菌分子鉴定

皮肤癣菌是浅表真菌感染的重要致病菌,常见致病菌种包括红色毛癣菌、趾间毛癣菌、絮状表皮癣菌、犬小孢子菌和内弯小孢子菌。本研究基于实时荧光PCR的高分辨率熔解曲线分析技术,以几丁质合成酶1为目标基因设计扩增引物,对经测序鉴定的上述5种皮肤癣菌进行熔解曲线分析,并在方法建立后,对已鉴定的临床菌株进行验证。实验结果表明,使用该方法上述5种皮肤癣菌均能扩增,并表现出不同的熔解曲线和熔解温度值,可有效区分;对其余对照菌及人基因均无法扩增,特异性为100%;临床菌株的熔解曲线分析结果与DNA测序结果一致。本研究提供了一种快速、准确鉴别5种常见皮肤癣菌的方法,可为临床应用和分子流行病学研究提供参考。

Molecular identification of five common dermatophytes based on high-resolution melting curves

Dermatophytes are important pathogens of superficial fungal infections, including five common pathogens: Trichophyton rubrum, Trichophyton interdigitale, Epidermophyton floccosum, Microsporum canis and Microsporum incurvatum. Real-time PCR-based high-resolution melting (HRM) assay was used to differentiate the five common dermatophytes, which was identified by DNA sequencing, and chitin synthase 1 was selected as the target gene to design amplification primers. Experimental results showed that five common dermatophytes could be effectively amplified and distinguished by different melting curves and melting temperature values. Negative controls and human DNA could not be amplified with 100% specificity. The analytical result of clinical strains was consistent with that of DNA sequencing. This study provides a rapid and accurate method for identification of five common dermatophytes, and valuable references for clinical applications and molecular epidemiological research.