Beck Kyler, Reyes Corral Cesar Alejandro, Rodriguez-Rodriguez Mariana, May Connie, Barnett Ryan, Thornton Mike, Bates Austin A, Woodhall James Warwick, Schroeder Brenda K
University of Idaho, Plant Sciences, Moscow, Idaho, United States;
University of Idaho, Entomology, Plant Pathology and Nematology, Moscow, Idaho, United States;
Plant Dis. 2020 Sep 15. doi: 10.1094/PDIS-06-20-1399-PDN.
In September 2014, a high rate of bulb rot (5-15% depending on producer) was reported across all cultivars developing early in the storage season in the onion producing region of southwestern Idaho. Spanish yellow onion bulbs cv. Vaquero displaying tan to light brown necrotic rot were obtained. The bulb rot originated in the neck and spread to successive scales (Figure 1). In August 2015, onion cv. Redwing and Vaquero were observed to have wet necrotic lesions developing on leaves in the field (Figure 2). Margins of necrotic tissue, 1-2 cm, were excised, surface sterilized, plated on water agar medium and incubated at 24°C. Hyphal growth was sub-cultured from eight strains (A- D in 2014; E-H in 2015) to fresh potato dextrose agar to obtain pure cultures. Cultures were characteristic of species as described by Nelson et al. (1983) with the presence of microconidia formed on polyphialides with macroconidia present. Primers ITS4-A1 and ITS5 primers (White et al. 1990); EF-1 and EF-2 (O'Donnell et al. 1998); and fRPB2-5F and fRPB2-7cR (Liu et al. 1999) were used to amplify regions of the ITS, elongation factor 1-α and the second largest subunit of DNA-directed RNA polymerase II. Amplicons were sequenced and analyzed using BLAST (https://www.ncbi.nlm.nih.gov/) and in combination using Pairwise DNA Alignment and Polyphasic Identification (http://www.westerdijkinstitute.nl/Fusarium/DefaultInfo.aspx?Page=Home) as described by O'Donnell et al. 2015. Analysis indicated that these strains are , which is part of the species complex (O'Donnell et al. 1998). Similarity (99.5%) was observed in pairwise analyses and the polyphasic identification clustering to representative strain NRRL 22944 and others. Sequences were submitted to Genbank and registered accession numbers are found in Table 1. To complete Koch's postulates, cv. Vaquero onion bulbs were surface sterilized and injected with 3 × 10 microconidia into the shoulder of each bulb. Five bulbs were inoculated for each isolate, placed in a mesh bag, and incubated at 30°C in the dark. Five bulbs injected with sterile water and five non-inoculated bulbs served as controls. After 14 days, each bulb was sliced vertically down the center and inspected for rot. All eight strains induced tan to light brown necrotic rot symptoms in each inoculated bulb. No symptoms were observed for the water inoculated and the non-inoculated onion bulbs. A fungus was isolated from the necrotic tissue and confirmed to be as described above. Ten µl aliquots containing 1 × 10 microconidia of strains (C, E-H) were applied to leaves in triplicate of 12-week-old onion plants (cv. Vaquero) wounded with a 21-gauge needle. Water controls were included. Within three days lesions, with light chlorosis, began to form and quickly spread on the leaves. A fungus was isolated and confirmed to be as described above. This is the first extensive description and identification of causing bulb rot in storage in Idaho (Mohan et al. 1997). In addition, this is the first report of the fungus causing leaf infection in the field. These findings confirm as the causal agent of the high incidence of bulb rot observed in 2014 and 2015. This bulb rot continues to occur in southwestern Idaho and since the pathogen can cause leaf infections growers are encouraged to be vigilant for both leaf lesions during the growing season and bulb rot in storage.
2014年9月,爱达荷州西南部洋葱产区报告称,在贮藏季节早期发育的所有品种中,鳞茎腐烂率很高(取决于生产商,为5%-15%)。获得了表现出棕褐色至浅褐色坏死腐烂的西班牙黄洋葱品种Vaquero的鳞茎。鳞茎腐烂始于颈部,并蔓延至连续的鳞片(图1)。2015年8月,观察到洋葱品种Redwing和Vaquero在田间叶片上出现湿坏死病斑(图2)。切除坏死组织边缘1-2厘米,进行表面消毒,接种于水琼脂培养基上,在24°C下培养。从8个菌株(2014年的A-D;2015年的E-H)的菌丝生长物转接至新鲜马铃薯葡萄糖琼脂上以获得纯培养物。培养物具有Nelson等人(1983年)所描述的该物种的特征,即有多育瓶梗上形成的小分生孢子以及大分生孢子。使用引物ITS4-A1和ITS5引物(White等人,1990年);EF-1和EF-2(O'Donnell等人,1998年);以及fRPB2-5F和fRPB2-7cR(Liu等人,1999年)来扩增ITS、延伸因子1-α和DNA定向RNA聚合酶II的第二大亚基区域。扩增产物进行测序,并使用BLAST(https://www.ncbi.nlm.nih.gov/)进行分析,并按照O'Donnell等人2015年所述,结合使用成对DNA比对和多相鉴定(http://www.westerdijkinstitute.nl/Fusarium/DefaultInfo.aspx?Page=Home)。分析表明这些菌株属于,是该物种复合体的一部分(O'Donnell等人,1998年)。在成对分析和与代表性菌株NRRL 22944及其他菌株的多相鉴定聚类中观察到相似性(99.5%)。序列提交至Genbank,登录号见表1。为了完成柯赫氏法则,对Vaquero洋葱鳞茎进行表面消毒,并向每个鳞茎肩部注射3×10个小分生孢子。每个分离株接种5个鳞茎,置于网袋中,在30°C黑暗条件下培养。注射无菌水的5个鳞茎和5个未接种的鳞茎作为对照。14天后,将每个鳞茎沿中心垂直切开并检查腐烂情况。所有8个菌株在每个接种的鳞茎中均诱导出棕褐色至浅褐色坏死腐烂症状。接种水和未接种的洋葱鳞茎未观察到症状。从坏死组织中分离出一种真菌,并如上所述确认为。将含有1×个菌株(C、E-H)小分生孢子的10微升等分试样一式三份施用于用21号针头致伤的12周龄洋葱植株(品种Vaquero)的叶片上。设置水对照。三天内,叶片上开始形成带有轻度褪绿的病斑并迅速蔓延。分离出一种真菌,并如上所述确认为。这是爱达荷州关于导致贮藏期鳞茎腐烂的首次详细描述和鉴定(Mohan等人,1997年)。此外,这是该真菌导致田间叶片感染的首次报道。这些发现证实是2014年和2015年观察到的鳞茎腐烂高发的致病因子。这种鳞茎腐烂在爱达荷州西南部仍持续发生,并且由于该病原菌可导致叶片感染,鼓励种植者在生长季节对叶片病斑和贮藏期鳞茎腐烂保持警惕。