Agri-environmental Division, Aichi Agricultural Research Center, Nagakute, Aichi, Japan.
FEMS Microbiol Lett. 2014 Jun;355(1):28-35. doi: 10.1111/1574-6968.12453. Epub 2014 May 22.
Root rot of poinsettia, caused by Pythium helicoides at high temperatures in hydroponic cultures, has become a serious problem in many parts of the world. We have developed a species-specific, loop-mediated isothermal amplification (LAMP) assay for the rapid diagnosis of this pathogen. The primers were designed using the ribosomal DNA internal transcribed spacer sequence. Primer specificity was established using 40 Pythium species including P. helicoides, 11 Phytophthora species, and eight other soil-borne pathogens. A sensitivity test was carried out using genomic DNA extracted from P. helicoides, and the detection limit was c. 100 fg which is comparable to that of the polymerase chain reaction (PCR). In addition, we tested the ease of pathogen detection in poinsettia roots. The LAMP results were consistent with those from the conventional plating method and showed more sensitivity than the PCR results. Consequently, the LAMP method developed in this study is effective for the rapid and easy detection of P. helicoides.
一品红根腐病由高温下水培条件下的腐霉螺旋体引起，已成为世界许多地区的严重问题。我们已经开发出一种针对该病原体的快速诊断的种特异性环介导等温扩增（LAMP）检测方法。该引物是使用核糖体 DNA 内部转录间隔区序列设计的。通过使用包括腐霉螺旋体在内的 40 种腐霉、11 种疫霉和其他 8 种土壤传播病原体来确定引物特异性。使用从腐霉螺旋体中提取的基因组 DNA 进行了灵敏度测试，检测限约为 100 fg，与聚合酶链反应（PCR）相当。此外，我们还测试了在一品红根中检测病原体的简便性。LAMP 结果与常规平板法一致，并且比 PCR 结果更敏感。因此，本研究中开发的 LAMP 方法可有效快速简便地检测腐霉螺旋体。
