Ahani Azari Ania, Amanollahi Reza, Jafari Jozani Razi, Trott Darren J, Hemmatzadeh Farhid
Department of Microbiology, Gorgan branch, Islamic Azad University, Gorgan, Iran.
Department of Clinical Sciences, School of Veterinary Medicine, Shiraz University, Shiraz, Iran.
Trop Anim Health Prod. 2020 May;52(3):1043-1047. doi: 10.1007/s11250-019-02098-4. Epub 2019 Nov 1.
Mycoplasma species cause wide ranges of infectious diseases in human and animals. The aim of the present study was to evaluate a real-time polymerase chain reaction (RT-PCR) followed by a high-resolution melting curve assay (HRM) for rapid differentiation of Mycoplasma species isolated from clinical cases of bovine and porcine respiratory disease. Lung samples from suspected cases to respiratory infections from cows and pigs were cultured on specific media, and the extracted DNA were tested by conventional polymerase chain reaction (PCR) assays for Mycoplasma. A set of universal primers specific for the 16S ribosomal RNA gene was designed and used for RT-PCR and HRM. The HRM analysis was able to differentiate between five different species of Mycoplasmas, namely, M. hyopneumoniae, M. bovis, M. hyorhinis, M. hyosynoviae and other uncultured Mycoplasma. All results were confirmed based on 16S rRNA gene sequencing. This rapid and reliable assay was as a simple alternative to PCR and sequencing, differentiating bovine and porcine mycoplasmas in species level.
支原体可引起人和动物的多种传染病。本研究的目的是评估实时聚合酶链反应（RT-PCR）结合高分辨率熔解曲线分析（HRM），用于快速鉴别从牛和猪呼吸道疾病临床病例中分离出的支原体种类。将疑似牛和猪呼吸道感染病例的肺组织样本在特定培养基上培养，提取的DNA通过常规聚合酶链反应（PCR）检测支原体。设计了一组针对16S核糖体RNA基因的通用引物，并用于RT-PCR和HRM。HRM分析能够区分五种不同的支原体，即猪肺炎支原体、牛支原体、猪鼻支原体、猪滑膜支原体和其他未培养的支原体。所有结果均通过16S rRNA基因测序得到证实。这种快速可靠的检测方法是PCR和测序的一种简单替代方法，可在种水平上区分牛和猪支原体。
