Zhang J X, Lin B R, Shen H F, Pu X M, Chen Z N, Feng J J
Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China.
Shenzhen Academy of Inspection and Quarantine, Shenzhen 518010, China.
Plant Dis. 2012 Dec;96(12):1819. doi: 10.1094/PDIS-07-12-0620-PDN.
Potato (Solanum tuberosum L.) is a major crop in China, with 80.0 million tons being produced in 2010 on 3.3 million ha. Pectobacterium carotovorum subsp. carotovorum Jones 1901; Hauben et al. 1999 causes soft rot worldwide on a wide range of hosts including potato, carrot, and cabbage. During spring 2010, a soft rot with a foul smell was noted in stored potato tubers of different cultivars in the Guangdong Province. Symptoms on tubers appeared as tan, water-soaked areas with watery ooze. The rotted tissues were white to cream colored. Stems of infected plants with typical inky black symptoms could also be found in the fields prior to harvest. Three different potato fields were surveyed, and 13% of the plants had the symptoms. Twenty-seven samples (three symptomatic tubers per sample) were collected. Bacteria were successfully isolated from all diseased tissues on nutrient agar media supplemented with 5% sucrose and incubated at 26 ± 1°C for 36 h. After purification on tripticase soy agar media, four typical strains (7-3-1, 7-3-2, 8-3-1, and 8-3-2) were identified using the following deterministic tests: gram-negative rods, oxidase negative, facultatively anaerobic, able to degrade pectate, sensitive to erythromycin, negative for phosphatase, unable to produce acid from α-methyl-glucoside, and produced acid from trehalose. Biolog analysis (Ver 4.20.05, Hayward, CA) identified the strains as P. carotovorum subsp. carotovorum (SIM 0.808, 0.774, 0.782, and 0.786, respectively). The identity of strains 7-3-1 (GenBank Accession No. JX258132), 7-3-2 (JX258133), and 8-3-1 (JX196705) was confirmed by 16S rRNA gene sequencing (4), since they had 99% sequence identity with other P. carotovorum subsp. carotovorum strains (GenBank Accession Nos. JF926744 and JF926758) using BLASTn. Further genetic analysis of strain 8-3-1 was performed targeting informative housekeeping genes, i.e., acnA (GenBank Accession No. JX196704), gabA (JX196706), icdA (JX196707), mdh (JX196708), mtlD (JX196709), pgi (JX196710), and proA (JX196711) (2). These sequences from strain 8-3-1 were 99 to 100%, homologous to sequences of multiple strains of P. carotovorum subsp. carotovorum. Therefore, strain 8-3-1 grouped with P. carotovorum subsp. carotovorum on the phylogenetic trees (neighbor-joining method, 1,000 bootstrap values) of seven concatenated housekeeping genes when compared with 60 other strains, including Pectobacterium spp. and Dickeya spp. (3). Pathogenicity of four strains (7-3-1, 7-3-2, 8-3-1, and 8-3-2) was evaluated by depositing a bacterial suspension (10 CFU/ml) on the potato slices of cultivar 'Favorita' and incubating at 30 ± 1°C. Slices inoculated with just water served as non-inoculated checks. The strains caused soft rot within 72 h and the checks had no rot. Bacteria were reisolated from the slices and were shown to be identical to the original strains based on morphological, cultural, and biochemical tests. Although this pathogen has already been reported in northern China (1), to our knowledge, this is the first report of P. carotovorum subsp. carotovorum causing bacterial soft rot of potato in Guangdong Province of China. References: (1) Y. X. Fei et al. J. Hexi Univ. 26:51, 2010.(2) B. Ma et al. Phytobacteriology 97:1150, 2007. (3) S. Nabhan et al. Plant Pathol. 61:498, 2012. (4) W. G. Weisbury et al. J. Bacteriol. 173:697, 1991.
马铃薯(Solanum tuberosum L.)是中国的主要作物,2010年在330万公顷的土地上产量达8000万吨。胡萝卜软腐果胶杆菌胡萝卜软腐亚种Jones 1901;Hauben等人1999年报道该菌在全球范围内可导致包括马铃薯、胡萝卜和卷心菜在内的多种寄主发生软腐病。2010年春季,在广东省不同品种的贮藏马铃薯块茎中发现了一种伴有恶臭味的软腐病。块茎上的症状表现为黄褐色、水渍状区域并有似水的渗出物。腐烂组织呈白色至奶油色。在收获前的田间也能发现感染植株的茎部出现典型的墨黑色症状。对三块不同的马铃薯田进行了调查,13%的植株有这些症状。采集了27个样本(每个样本三个有症状的块茎)。在添加5%蔗糖的营养琼脂培养基上,从所有患病组织中成功分离出细菌,并在26±1°C下培养36小时。在胰蛋白胨大豆琼脂培养基上纯化后,通过以下确定性试验鉴定出四株典型菌株(7-3-1、7-3-2、8-3-1和8-3-2):革兰氏阴性杆菌、氧化酶阴性、兼性厌氧、能够降解果胶酸盐、对红霉素敏感、磷酸酶阴性、不能从α-甲基葡萄糖苷产酸、能从海藻糖产酸。Biolog分析(版本4.20.05,加利福尼亚州海沃德)将这些菌株鉴定为胡萝卜软腐果胶杆菌胡萝卜软腐亚种(SIM分别为0.808、0.774、0.782和0.786)。菌株7-3-1(GenBank登录号JX258132)、7-3-2(JX258133)和8-3-1(JX196705)的身份通过16S rRNA基因测序得到确认(4),因为使用BLASTn与其他胡萝卜软腐果胶杆菌胡萝卜软腐亚种菌株(GenBank登录号JF926744和JF926758)相比,它们具有99%的序列同一性。针对信息丰富的管家基因,即acnA(GenBank登录号JX196704)、gabA(JX196706)、icdA(JX196707)、mdh(JX196708)、mtlD(JX196709)、pgi(JX196710)和proA(JX196711)(2),对菌株8-3-1进行了进一步的遗传分析。菌株8-3-1的这些序列与胡萝卜软腐果胶杆菌胡萝卜软腐亚种多个菌株的序列具有99%至100%的同源性。因此,与包括果胶杆菌属和迪基氏菌属在内的其他60个菌株相比,在七个串联管家基因的系统发育树(邻接法,1000次自展值)上,菌株8-3-1与胡萝卜软腐果胶杆菌胡萝卜软腐亚种归为一类(3)。通过将细菌悬浮液(10 CFU/ml)接种在‘费沃瑞它’品种的马铃薯切片上,并在30±1°C下培养,对四株菌株(7-3-1、7-3-2、8-3-1和8-3-2)的致病性进行了评估。仅接种水的切片作为未接种对照。这些菌株在72小时内导致软腐,而对照没有腐烂。从切片中重新分离出细菌,并根据形态、培养和生化试验证明与原始菌株相同。尽管这种病原菌在中国北方已有报道(1),但据我们所知,这是胡萝卜软腐果胶杆菌胡萝卜软腐亚种在中国广东省导致马铃薯细菌性软腐病的首次报道。参考文献:(1)Y. X. Fei等人,《河西学院学报》26:51,2010年。(2)B. Ma等人,《植物细菌学》97:1150,2007年。(3)S. Nabhan等人,《植物病理学》61:498,2012年。(4)W. G. Weisbury等人,《细菌学杂志》173:697,1991年。