莫瑾 王哲 周慧平 朱宏建 朱水芳 廖晓兰

摘要 ：对水稻中多种病原细菌的检测，使用常规方法往往耗时耗力，而多重PCR可以更加高效地进行多种细菌的检测。根据水稻细菌性谷枯病菌gyrB基因，水稻细菌性叶鞘褐腐病菌PfsI/R quorum sensing 位点以及水稻细菌性条斑病菌和水稻白叶枯病菌含铁细胞接受因子基因设计引物，建立4种水稻病菌的多重PCR检测方法，对方法进行特异性和灵敏度测试，并对采自不同地区的水稻样本進行检测。结果显示，多重PCR方法能同步地快速检测出水稻细菌性谷枯病菌、水稻细菌性叶鞘褐腐病菌、水稻细菌性条斑病菌或水稻白叶枯病菌，检测灵敏度达到103 cfu/mL的菌液浓度，利用该方法对我国不同地区的58份水稻种子进行检测，其中17个样本检测出水稻细菌性条斑病菌或水稻白叶枯病菌，未检测到水稻细菌性谷枯病菌和水稻细菌性叶鞘褐腐病菌。

关键词 ：多重PCR; 水稻病原细菌; 检测

中图分类号：

S 435.111

文献标识码： A

DOI： 10.16688/j.zwbh.2020205

A multiplex PCR method for rapid detection of four rice pathogenic bacteria

MO Jin1，2， WANG Zhe2， ZHOU Huiping2， ZHU Hongjian1， ZHU Shuifang1， LIAO Xiaolan1*

（1. College of Plant Protection，Hunan Agricultural University， Changsha 410128， China;

2. Technology Center of Changsha Customs， Changsha 410004， China）

Abstract

Multiplex PCR is an efficient and simple method to detect a variety of pathogenic bacteria in rice at the same time. A multiplex PCR detection method was established for four rice pathogenic bacteria， and the primers were designed based on the gyrB gene of Burkholderia glumae，the PfsI/R quorum sensing locus of Pseudomonas fuscovaginae and the iron-containingcell-receiving factor genes of Xanthomonas oryzae pv. oryzae and X. oryzae pv. oryzicola. The specificity and sensitivity of the multiplex PCR method were validated， and rice samples from different regions were tested using this method. The results showed that the multiplex PCR method can fast and accurately detect four rice pathogenic bacteria. The detectability of this method reached 103 cfu/mL in the bacterial solution. Among 58 rice samples from different regions， neither B.glumae nor P.fuscovaginae was detected，while X.oryzae pv. oryzae or X.oryzae pv. oryzicola were positive in 17 samples.

Key words

multiplex PCR; rice pathogenic bacteria; detection

水稻细菌性谷枯病（病原菌：Burkholderia glumae）[12]、水稻细菌性叶鞘褐腐病（病原菌：Pseudomonas fuscovaginae）[34]、水稻白叶枯病（病原菌：Xanthomonas oryzae pv. oryzae）和水稻细菌性条斑病（病原菌：X.oryzae pv. oryzicola）[56]等水稻细菌性病害，对水稻危害严重，导致严重减产。其中水稻细菌性谷枯病、水稻细菌性条斑病和水稻白叶枯病属于我国进境检疫性病害，水稻细菌性叶鞘褐腐病是马来西亚等稻种进口国的进境检疫性病害。

目前有多种关于这4种病原菌检测技术的报道。Takeuchi等[1]建立了B.glumae的检测方法;罗金燕等[7]研究了B.glumae的生理生化特点……