Kursa Olimpia, Woźniakowski Grzegorz, Tomczyk Grzegorz, Sawicka Anna, Minta Zenon
Department of Poultry Diseases, National Veterinary Research Institute, Partyzantów 57 Avenue, 24-100, Puławy, Poland.
Arch Microbiol. 2015 Mar;197(2):319-25. doi: 10.1007/s00203-014-1063-2. Epub 2014 Nov 21.
Mycoplasma synoviae (MS) remains a serious concern in production of poultry and affects world production of chickens and turkeys. Loop-mediated isothermal amplification (LAMP) of DNA has been recently used for the identification of different economically important avian pathogens. The aim of this study was to develop LAMP for simple and inexpensive detection of MS strains in poultry using specifically designed primers targeting hemagglutin A (vlh) gene. The assay was conducted in a water bath for 1 h at 63 °C. The results were visualized after addition of SYBR Green(®) fluorescent dye. LAMP was specific exclusively for MS without cross-reactivity with other Mycoplasma species. The sensitivity of LAMP was determined as 10(-1) CFU/ml and was 1,000 times higher than MS-specific polymerase chain reaction. LAMP assay was conducted on 18 MS field strains to ensure its reliability and usefulness. This is the first report on LAMP development and application for the rapid detection of MS isolated from chickens. This simple method may be applied by diagnostic laboratories without access to expensive equipment.
鸡滑液囊支原体(MS)仍是家禽生产中的一个严重问题,影响着全球鸡肉和火鸡的产量。DNA的环介导等温扩增(LAMP)技术最近已被用于鉴定不同的具有重要经济意义的禽类病原体。本研究的目的是利用针对血凝素A(vlh)基因设计的特异性引物,开发一种用于简单、廉价检测家禽中MS菌株的LAMP方法。该检测在63℃的水浴中进行1小时。加入SYBR Green(®)荧光染料后观察结果。LAMP仅对MS具有特异性,与其他支原体物种无交叉反应。LAMP的灵敏度测定为10^(-1) CFU/ml,比MS特异性聚合酶链反应高1000倍。对18株MS田间菌株进行了LAMP检测,以确保其可靠性和实用性。这是关于开发LAMP并将其应用于快速检测从鸡中分离出的MS的首次报告。这种简单的方法可供没有昂贵设备的诊断实验室使用。