Pruvost O, Robène-Soustrade I, Ah-You N, Jouen E, Boyer C, Wuster G, Hostachy B, Napoles C, Dogley W
CIRAD-Université de la Réunion, UMR PVBMT, Saint Pierre, La Réunion, F-97410 France.
LNPV, Pôle de Protection des Plantes, Saint Pierre, La Réunion, F-97410 France.
Plant Dis. 2009 Jun;93(6):671. doi: 10.1094/PDIS-93-6-0671B.
Bacterial leaf spot of cucurbits caused by Xanthomonas cucurbitae (4) can be a harmful disease of several cucurbit species in tropical environments, mainly within the Cucumis, Cucurbita, and Citrullus genera. The bacterium induces angular, water-soaked leaf spots, which sometimes become necrotic and have a chlorotic halo. Scab-like lesions on fruit can also be observed (2). Water-soaked, angular leaf lesions were collected from approximately 15 watermelon plants (Citrullus lanatus) in a production field located in Mahé, Seychelles in 2003. Yellow-pigmented Xanthomonas-like bacterial colonies were isolated on KC semiselective medium (yeast extract 7 g, peptone 7 g, glucose 7 g, agar 18 g, distilled water 1,000 ml, propiconazole 20 μg ml, cephalexin 40 mg liter, and kasugamycin 20 mg liter) from all isolation attempts (3). Amplified fragment length polymorphism (AFLP) analysis was performed on four watermelon strains together with reference strains of Xanthomonas cucurbitae (LMG 690 [type strain] and LMG 8663) and the type strain of all other valid Xanthomonas species using SacI/MspI and four primer pairs (unlabeled MspI + 1 [A, C, T, or G] primers and 5'-labeled - SacI + C primer for the selective amplification step) (1). The four strains from watermelon showed identical fingerprints and were most closely related to X. cucurbitae. One strain from diseased watermelon (JZ88-1) was further analyzed by MultiLocus Sequence Analysis (MLSA) using portions of three housekeeping genes (atpD, dnaK, and gyrB) as described previously (1). This strain displayed a very high relatedness (99.8 and 98.9% with strain LMG 690 and LMG 8663, respectively) to the two reference strains of X. cucurbitae. AFLP and MLSA were useful for identifying strains at the species level that were consistent with previous results (1). Bottle-gourd (Lagenaria siceraria), pumpkin (Cucurbita maxima), squash cv. Aurore (Cucurbita pepo), cucumber cv. L-04 (Cucumis sativus), cantaloupe melon cv. Cezanne (Cucumis melo), and watermelon cv. Fou-nan (C. lanatus) leaves were infiltrated (10 inoculation sites per leaf and three replicates) with bacterial suspensions (JZ88-1, LMG 690 and LMG 8663) containing approximately 1 × 10 CFU ml (approximately 1 × 10 CFU per inoculation site). Typical water-soaked lesions that developed into necrotic spots were observed 6 to 8 days after inoculation for all inoculated strains on all inoculated plant species. One month after inoculation, Xanthomonas was recovered from lesions and population sizes determined on KC semiselective medium (3) ranging from 1 × 10 to 9 × 10 CFU per lesion were typical of a compatible interaction. Bacterial leaf spot has appeared sporadically in Mahé, Seychelles since 2003, most often with limited incidence. However, growers need to be aware of the potential negative effect of this disease in tropical environments. References: (1) N. Ah-You et al. Int. J. Syst. Evol. Microbiol. 59:306, 2009. (2) J. F. Bradbury. Page 309 in: Guide to Plant Pathogenic Bacteria. CAB International, Slough, UK, 1986. (3) O. Pruvost et al. J. Appl. Microbiol. 99:803, 2005. (4) L. Vauterin et al. Int. J. Syst. Bacteriol. 45:472, 1995.
由西瓜黄单胞菌（Xanthomonas cucurbitae）（4）引起的葫芦科细菌性叶斑病，在热带环境中，对几种葫芦科作物来说，可能是一种有害病害，主要发生在黄瓜属（Cucumis）、南瓜属（Cucurbita）和西瓜属（Citrullus）中。该细菌会引发角状、水渍状叶斑，这些叶斑有时会坏死，并带有褪绿晕圈。果实上也会出现疮痂状病斑（2）。2003年，从塞舌尔马埃岛一个生产地里的大约15株西瓜植株（Citrullus lanatus）上，采集了水渍状、角状叶斑。在KC半选择性培养基（酵母提取物7克、蛋白胨7克、葡萄糖7克、琼脂18克、蒸馏水1000毫升、丙环唑20微克/毫升、头孢氨苄40毫克/升、春雷霉素20毫克/升）上，从所有分离试验中均分离出了黄色色素沉着的类黄单胞菌菌落（3）。对4个西瓜菌株以及西瓜黄单胞菌的参考菌株（LMG 690 [模式菌株]和LMG 8663）和所有其他有效黄单胞菌物种的模式菌株，使用SacI/MspI和4对引物（未标记的MspI + 1 [A、C、T或G]引物以及用于选择性扩增步骤的5'-标记的 - SacI + C引物）进行了扩增片段长度多态性（AFLP）分析（1）。来自西瓜的4个菌株显示出相同的指纹图谱，并且与西瓜黄单胞菌关系最为密切。使用之前描述的方法（1），对一株患病西瓜菌株（JZ88-1），通过多位点序列分析（MLSA），利用3个管家基因（atpD、dnaK和gyrB）的部分序列进行了进一步分析。该菌株与西瓜黄单胞菌的两个参考菌株显示出非常高的相关性（分别与菌株LMG 690和LMG 8663的相关性为99.8%和98.9%）。AFLP和MLSA对于在物种水平上鉴定菌株很有用，与之前的结果一致（1）。用含有约1×10⁸CFU/毫升（每个接种位点约1×10⁸CFU）的细菌悬液（JZ88-1、LMG 690和LMG 8663）对瓠瓜（Lagenaria siceraria）、南瓜（Cucurbita maxima）、“奥罗雷”南瓜品种（Cucurbita pepo）、“L-04”黄瓜品种（Cucumis sativus）、“塞尚”哈密瓜品种（Cucumis melo）和“福南”西瓜品种（C. lanatus）的叶片进行浸润接种（每片叶10个接种位点，3次重复）。接种后6至8天，在所有接种的植物品种上，所有接种菌株均观察到典型的水渍状病斑发展成坏死斑。接种1个月后，从病斑中分离出黄单胞菌，并在KC半选择性培养基（3）上测定菌量，每个病斑1×10⁶至9×10⁶CFU的菌量是亲和性互作的典型特征。自2003年以来，细菌性叶斑病在塞舌尔马埃岛偶尔出现，发病率大多有限。然而，种植者需要意识到这种病害在热带环境中的潜在负面影响。参考文献：（1）N. Ah-You等人，《国际系统与进化微生物学杂志》59:306，2009年。（2）J. F. Bradbury，《植物病原细菌指南》第309页，CAB国际，英国斯劳，1986年。（3）O. Pruvost等人，《应用微生物学杂志》99:803，2005年。（4）L. Vauterin等人，《国际系统细菌学杂志》45:472，1995年。
