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甘肃省首次报道由立枯丝核菌AG4 HGII引起的马铃薯茎溃疡病

First Report of Potato Stem Canker Caused by Rhizoctonia solani AG4 HGII in Gansu Province, China.

作者信息

Yang Y G, Wu X H

机构信息

Department of Plant Pathology, China Agricultural University, Beijing 100193, China.

出版信息

Plant Dis. 2013 Jun;97(6):840. doi: 10.1094/PDIS-09-12-0896-PDN.

Abstract

Black scurf and stem canker on potato (Solanum tuberosum L.), caused by Rhizoctonia solani, is an important disease throughout the world. Isolates of R. solani AG3 are the principal cause of these diseases on potato (2). In August 2011, at the tuber bulking growth stage, symptoms typically associated stem canker, including dark brown stem lesions, were observed on 20% of potato plants collected from 23 locations (about 2,000 ha) in Gansu Province, northwest China. Stem pieces (each 5 mm long) taken from the margins of the healthy and diseased tissues were surface-disinfected with 0.5% NaOCl for 2 min, rinsed with sterilized water, dried, then placed on potato dextrose agar (PDA) at 25°C in the dark. Twenty-nine fungal isolates taken from single hyphal tips were identified as R. solani based on morphological traits, including mycelium branched at right angles with a septum near the branch and a slight constriction at the branch base. Hyphal cells were determined to be multinucleate (4 to 10 nuclei/cell) when stained with 4'-6-diamidino-2-phenylindole (DAPI). Anastomosis groups were determined by pairing with reference strains (kindly provided by N. Kondo, Hokkaido University, Japan), and three isolates (designed GS-15, GS-24, and GS-25) anastomosed with isolates of R. solani AG4. The internal transcribed spacer (ITS) region of rDNA was amplified from genomic DNA of each of the three isolates with primers ITS1 and ITS4. The resulting sequences (GenBank Accession Nos. JX843818, JX843819, and JX843820) were 100% identical to those of >10 R. solani AG4 HGII isolates (e.g., HQ629873.1; isolate ND13). Therefore, based on the anastomosis assay and molecular characteristics, the three isolates were identified as R. solani AG4 HGII. To determine pathogenicity of the AG4 HGII isolates, potato seed tubers (cv. Favorita) with 3 to 5 mm long sprouts were inoculated with wheat seeds (sterilized by autoclaving twice at 121°C for 1 h with a 24 h interval between autoclavings) colonized with each isolate (1). One sprouted tuber was planted in a sterilized plastic pot (1 liter) with a single colonized wheat seed placed 10 mm above the uppermost sprout tip in a sand/sawdust mixture (1:2 v/v, with dry heat sterilization at 161°C for 4 h before use). Plants were incubated in a glasshouse maintained at 25 to 27°C. The test was performed on 20 plants for each isolate, and the experiment was repeated. After 3 weeks, control plants inoculated with sterilized wheat seeds remained asymptomatic, and no Rhizoctonia spp. were isolated from these plants, whereas all inoculated plants showed symptoms of stem canker. R. solani AG4 HGII was reisolated consistently from symptomatic stems, and the identity of the reisolates confirmed by the morphological and molecular characteristics mentioned above, fulfilling Koch's postulates. Potato stem canker caused by R. solani AG4 HGII was reported previously in the United States (3). To our knowledge, this is the first report of R. solani AG4 HGII causing stem canker on potato in Gansu Province, the main potato-producing area of China. R. solani AG4 HGII can cause sheath blight on corn in China (4), which is commonly grown in rotation with potato. This rotation could increase the risk of soilborne infection to either crop by R. solani AG4 HGII. References: (1) M. J. Lehtonen et al. Plant Pathol. 57:141, 2008. (2) L. Tsror. J. Phytopathol. 158:649, 2010. (3) J. W. Woodhall et al. Plant Dis. 96:1701, 2012. (4) X. Zhou et al. J. Shenyang Agric. Univ. 43:33, 2012.

摘要

由立枯丝核菌引起的马铃薯(Solanum tuberosum L.)黑痣病和茎溃疡病是一种在全球范围内的重要病害。立枯丝核菌AG3菌群的分离物是马铃薯上这些病害的主要病因(2)。2011年8月,在块茎膨大生长阶段,在中国西北部甘肃省23个地点(约2000公顷)采集的马铃薯植株中,20%的植株出现了通常与茎溃疡病相关的症状,包括深褐色的茎部病斑。从健康和患病组织边缘取下的茎段(每段5毫米长)用0.5%次氯酸钠进行表面消毒2分钟,用无菌水冲洗,干燥,然后置于马铃薯葡萄糖琼脂(PDA)上,于25°C黑暗条件下培养。根据形态特征,从单个菌丝尖端分离得到的29个真菌分离物被鉴定为立枯丝核菌,其特征包括菌丝体呈直角分支,分支附近有隔膜,分支基部有轻微缢缩。用4′,6-二脒基-2-苯基吲哚(DAPI)染色后,菌丝细胞被确定为多核(4至10个核/细胞)。通过与参考菌株(由日本北海道大学的近藤信提供)配对来确定融合群,三个分离物(编号为GS-15、GS-24和GS-25)与立枯丝核菌AG4的分离物融合。用引物ITS1和ITS4从这三个分离物的基因组DNA中扩增核糖体DNA的内转录间隔区(ITS)。所得序列(GenBank登录号:JX843818、JX843819和JX843820)与10多个立枯丝核菌AG4 HGII分离物(如HQ629873.1;分离物ND13)的序列100%相同。因此,根据融合试验和分子特征,这三个分离物被鉴定为立枯丝核菌AG4 HGII。为了确定AG4 HGII分离物的致病性,将带有3至5毫米长芽的马铃薯种薯(品种为费沃瑞它)接种上被各分离物定殖的小麦种子(通过在121°C下高压灭菌两次,每次1小时,两次灭菌间隔24小时进行灭菌)(1)。将一个发芽的种薯种植在一个灭菌的塑料盆(1升)中,盆中装有一份定殖的小麦种子,放置在最上部芽尖上方10毫米处,盆中为沙子/锯末混合物(体积比1:2,使用前在161°C下干热灭菌4小时)。植株在保持在25至27°C的温室中培养。对每个分离物的20株植株进行试验,并重复该实验。3周后,接种灭菌小麦种子的对照植株没有症状,且未从这些植株中分离到立枯丝核菌属的任何菌株,而所有接种植株都表现出茎溃疡病的症状。从有症状的茎中持续重新分离到立枯丝核菌AG4 HGII,并且通过上述形态和分子特征确认了重新分离物的身份,从而满足了柯赫氏法则。先前在美国报道过由立枯丝核菌AG4 HGII引起的马铃薯茎溃疡病(3)。据我们所知,这是立枯丝核菌AG4 HGII在中国主要马铃薯产区甘肃省引起马铃薯茎溃疡病的首次报道。立枯丝核菌AG4 HGII在中国可引起玉米纹枯病(4),玉米通常与马铃薯轮作。这种轮作可能会增加立枯丝核菌AG4 HGII对两种作物土传感染的风险。参考文献:(1)M. J. Lehtonen等人。植物病理学。57:141,2008。(2)L. Tsror。植物病理学杂志。158:649,2010。(3)J. W. Woodhall等人。植物病害。96:1701,2012。(4)X. Zhou等人。沈阳农业大学学报。43:33,2012。

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