花生(ArachishypogaeaL.)是一种广泛种植的油料作物,在全球范围内具有重要的农业意义。2022年7月,莱西(36º85\'N,120º54\'E),山东省,中国。大约25%的植物表现出各种症状,包括茎和根腐烂和变黑,茎上的微菌核,叶子变黄和枯萎,甚至死亡。收集20株患病植物以确认病原体。有症状的根被切成小块,用75%乙醇消毒1分钟,用0.5%NaClO消毒2分钟,用无菌水冲洗三次,在无菌滤纸上干燥,然后涂布在补充有100μg/mL氯霉素的马铃薯葡萄糖琼脂(PDA)上,并在25°C下在黑暗中孵育。在成长的初期,真菌稀疏形成,白色菌丝体,哪个白色,然后随着年龄的增长变暗,5天后在培养基中形成微菌核。菌丝体聚集成黑色,圆形至椭圆形或不规则形状的微菌核长84至163μm,宽54至125μm(n=40)。这些形态特征与Macrophominaphaseolina(Holliday和Punithalingam,1970).通过用代表性分离株SXY183的ITS1和ITS4对内部转录间隔区(ITS)和翻译延伸因子1-α(TEF)进行测序来进行分子鉴定。SXY183的ITS(OR056369)和TEF(OR098356)显示出100%和97.74%的相似性,与绿豆菜(KF951622,KF951997),分别。使用基于ITS和TEF的基因序列的相邻连接(NJ)分析进行系统发育分析。根据分子分析和形态特征,将真菌鉴定为M.phaseolina。在温室条件下对花生测试了代表性分离株(SXY183)的致病性。两周龄花生(华宇号9115)的幼苗在每株植物的根基部接种菌丝栓(直径8mm),并在温室中培养(白天30℃,晚上25℃,一个12小时的光周期,和80%RH)。用未定殖的PDA的插塞接种10株植物作为对照。接种后7天在所有接种幼苗的茎和根上观察到棕色病变,但不是对照植物。该实验重复三次。从有症状的根中重新分离并基于ITS和TEF的形态特征和DNA序列分析进行确认。M.phaseolina是一种土壤传播的真菌,分布在世界各地,具有广泛的寄主范围。以前已经报道了几种寄主植物如小豆,蚕豆,西瓜,Plukenetiavolbilis,中国的苍术和姜黄(蔡等。,2020;Sun等人。2016;Sun等人。,2019年;Sun等人。,2020年;王等人。,2020年;吴等人。,2022年)。然而,这是在山东省首次发现根腐病引起花生根腐病的报告,中国。我们的报告将为研究这种疾病的流行病学和管理提供重要信息。
Peanut (Arachis hypogaea L.) is a widely grown oilseed crop of great agricultural importance worldwide. In July 2022, disease symptoms were observed on peanut roots in Laixi (36º85\' N, 120º54\' E), Shandong Province,
China. About 25% of the plants showed various symptoms, including stem and root rot and blackening, microsclerotia on the stem, yellowing and wilting of leaves, and even death. Twenty diseased plants were collected to confirm the pathogen. Symptomatic roots were cut into small pieces, disinfested with 75% ethanol for 1 min and 0.5% NaClO for 2 min, rinsed three times with sterile water, dried on sterile filter paper, and then spread on potato dextrose agar (PDA) supplemented with 100 μg/mL chloramphenicol and incubated at 25°C in the dark. At the beginning of growth, the fungus formed sparse, white mycelia, which white, then darkened with age and microsclerotia were formed in the medium after 5 days. The mycelium aggregated into black, round to oblong or irregularly shaped microsclerotia 84 to 163 μm long and 54 to 125 μm wide (n=40). These morphological characteristics were consistent with the description of Macrophomina phaseolina (Holliday and Punithalingam, 1970). Molecular identification was performed by sequencing the internal transcribed spacer (ITS) region with ITS1 and ITS4 and translation elongation factor 1-alpha (TEF) with EF1-728F/EF1-986R (Glass and Donaldson 1995) of a representative isolate SXY183. ITS (OR056369) and TEF (OR098356) of SXY183 showed 100% and 97.74% similarity with M. phaseolina (KF951622, KF951997), respectively. Phylogenetic analysis was performed using Neighbor-Joining (NJ) analysis based on the gene sequences of ITS and TEF. The fungus was identified as M. phaseolina based on molecular analysis and morphological characteristics. The pathogenicity of a representative isolate (SXY183) was tested on peanuts under greenhouse conditions. Two-week-old peanut (Huayu No. 9115) seedlings were inoculated with a mycelial plug (8 mm diameter) at the root base of each plant and cultured in a greenhouse (30°C during the day and 25°C at night, a 12-h photoperiod, and 80% RH). Ten plants were inoculated with a plug of non-colonized PDA as a control. Brown lesions were observed on the stem and root of all inoculated seedlings 7 days after inoculation, but not on the control plants. The experiment was repeated three times. M. phaseolina was re-isolated from the symptomatic root and confirmed based on morphological characteristics and DNA sequence analysis of ITS and TEF. M. phaseolina is a soil-borne fungus that is distributed worldwide and has a broad host range. Disease agent has previously been reported on several host plants such as adzuki bean, faba bean, watermelon, Plukenetia volubilis, Atractylodes lancea and Curcuma longa in
China (Cai et al., 2020; Sun et al. 2016; Sun et al., 2019; Sun et al., 2020; Wang et al., 2020; Wu et al., 2022). However, this is the first report in which M. phaseolina was found to cause peanut root rot in Shandong Province,
China. Our report will provide important information for studying the epidemiology and management of this disease.