Bull C T, Rubio I
USDA-ARS Agricultural Research Station, Salinas, CA.
Undergraduate Research Opportunities Center, California State University, Monterey Bay, Seaside.
Plant Dis. 2011 Aug;95(8):1027. doi: 10.1094/PDIS-11-10-0804.
In 1978 and 1979, Pseudomonas syringae pv. maculicola strains DAR 33362, DAR 33363, and DAR 33406 were isolated from diseased Brassica hirta, B. nigra, and B. napus var. napus, respectively, in Wagga Wagga and Armatree, NSW, Australia (2). Peters et al. (2) demonstrated that these strains were similar to P. syringae pv. maculicola ICMP 4326 (CFBP 1637), which was recently transferred to Pseudomonas cannabina pv. alisalensis (1). We evaluated these Australian strains to determine if they might also be P. cannabina pv. alisalensis. Amplification of DNA using the BOXA1R primer and PCR resulted in identical DNA fragment banding patterns for Australian strains DAR 33362 and DAR 33363 and P. cannabina pv. alisalensis ICMP 4326 and CFBP 6875. The third Australian strain, DAR 33406, was 90% similar to P. cannabina pv. alisalensis; in contrast, it was only 77% similar to P. syringae pv. maculicola. All strains of P. cannabina pv. alisalensis, including the pathotype strain (CFBP 6866) and all three Australian strains, were lysed by bacteriophage PBS1, which is specific for P. cannabina pv. alisalensis strains (1). To complete Koch's postulates, pathogenicity was evaluated on B. hirta, B. nigra, and B. napus var. napus. In two independent experiments, two plants of each species were inoculated with each Australian strain or a phosphate buffer control treatment. In separate experiments, pathogenicity was evaluated on the differential hosts radish (Raphanus sativus cv. Comet) and broccoli raab (Brassica rapa cv. Sorrento), and plants inoculated with the pathotypes of P. cannabina pv. alisalensis and P. syringae pv. maculicola served as additional control treatments. Inoculum was prepared by growing the bacteria on nutrient agar for 48 h (27°C), suspending the bacteria in 0.01 M phosphate buffer (pH 7.0), and adjusting each suspension to 0.6 OD at 600 nm (approximately 10 CFU/ml). Treatments were applied by spraying until runoff. DAR 33362, DAR 33363, and DAR 33406 caused typical bacterial blight symptoms on B. hirta, B. nigra, and B. napus var. napus. Infected leaves became yellow, followed by the development of small (<2 mm in diameter), angular, water-soaked, and eventually, shot-holed spots. Bacteria isolated from symptomatic tissue following surface disinfestation of tissue with sodium hypochlorite (0.525%) had identical characteristics (rep-PCR DNA fragment banding patterns and phage sensitivity) to the strains used to inoculate the plants. Additionally, DAR 33362, DAR 33363, and DAR 33406, as well as P. cannabina pv. alisalensis, caused symptoms on radish and broccoli raab while P. syringae pv. maculicola and the buffer control did not. These data support the transfer of the Australian crucifer strains, originally identified as P. syringae pv. maculicola, to P. cannabina pv. alisalensis. To our knowledge, this is the first report of a bacterial disease of crucifers caused by P. cannabina pv. alisalensis in Australia. Differentiation of these pathogens will inform crop rotation strategies for disease management. References: (1) C. T. Bull et al. Syst. Appl. Microbiol. 33:105, 2010. (2) B. J. Peters et al. Plant Pathol. 53:3, 2004.
1978年和1979年，分别从澳大利亚新南威尔士州瓦加瓦加和阿马特里患病的白芥、黑芥和甘蓝型油菜甘蓝变种中分离出丁香假单胞菌黄斑变种菌株DAR 33362、DAR 33363和DAR 33406（2）。彼得斯等人（2）证明，这些菌株与丁香假单胞菌黄斑变种ICMP 4326（CFBP 1637）相似，该菌株最近被转移到大麻假单胞菌阿利萨变种（1）。我们对这些澳大利亚菌株进行了评估，以确定它们是否也是大麻假单胞菌阿利萨变种。使用BOXA1R引物和PCR扩增DNA，结果显示澳大利亚菌株DAR 33362和DAR 33363与大麻假单胞菌阿利萨变种ICMP 4326和CFBP 6875具有相同的DNA片段条带模式。第三个澳大利亚菌株DAR 33406与大麻假单胞菌阿利萨变种的相似度为90%；相比之下，它与丁香假单胞菌黄斑变种的相似度仅为77%。所有大麻假单胞菌阿利萨变种菌株，包括致病型菌株（CFBP 6866）和所有三个澳大利亚菌株，都被特异性针对大麻假单胞菌阿利萨变种菌株的噬菌体PBS1裂解（1）。为了完成科赫法则，我们评估了这些菌株对白芥、黑芥和甘蓝型油菜甘蓝变种的致病性。在两个独立的实验中，每种植物的两株分别接种每种澳大利亚菌株或磷酸盐缓冲液对照处理。在单独的实验中，我们评估了这些菌株对鉴别寄主萝卜（萝卜品种彗星）和抱子芥（芜菁品种索伦托）的致病性，并用接种了大麻假单胞菌阿利萨变种和丁香假单胞菌黄斑变种致病型的植物作为额外的对照处理。通过在营养琼脂上培养细菌48小时（27°C）来制备接种物，将细菌悬浮在0.01M磷酸盐缓冲液（pH 7.0）中，并将每种悬浮液在600nm处调整至0.6 OD（约10 CFU/ml）。通过喷雾处理直至径流。DAR 33362、DAR 33363和DAR 33406对白芥、黑芥和甘蓝型油菜甘蓝变种引起典型的细菌性叶斑病症状。受感染的叶片变黄，随后出现小的（直径<2mm）、角状、水渍状斑点，最终形成穿孔斑。用次氯酸钠（0.525%）对有症状组织进行表面消毒后，从症状组织中分离出的细菌与用于接种植物的菌株具有相同的特征（rep-PCR DNA片段条带模式和噬菌体敏感性）。此外，DAR 33362、DAR 33363和DAR 33406以及大麻假单胞菌阿利萨变种对萝卜和抱子芥引起症状，而丁香假单胞菌黄斑变种和缓冲液对照则未引起症状。这些数据支持将最初鉴定为丁香假单胞菌黄斑变种的澳大利亚十字花科菌株转移到大麻假单胞菌阿利萨变种。据我们所知，这是澳大利亚首次报道由大麻假单胞菌阿利萨变种引起的十字花科细菌性病害。这些病原体的鉴别将为病害管理的作物轮作策略提供依据。参考文献：（1）C.T. Bull等人，《系统与应用微生物学》33:105，2010年。（2）B.J. Peters等人，《植物病理学》53:3，2004年。
