Wang J-H, Li H-P, Zhang J-B, Wang B-T, Liao Y-C
Molecular Biotechnology Laboratory of Triticeae Crops and College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, P. R. China. This study was funded by the National Basic Research Program of China (2009CB118806) and in part supported by the Ministry of Education of China (20090146120013).
Plant Dis. 2014 Feb;98(2):279. doi: 10.1094/PDIS-05-13-0558-PDN.
From September 2009 to October 2012, surveys to determine population structure of Fusarium species on maize were conducted in 22 provinces in China, where the disease incidence ranged from 5 to 20% in individual fields. Maize ears with clear symptoms of Fusarium ear rot (with a white to pink- or salmon-colored mold at the ear tip) were collected from fields. Symptomatic kernels were surface-sterilized (1 min in 0.1% HgCl, and 30 s in 70% ethanol, followed by three rinses with sterile distilled water), dried, and placed on PDA. After incubation for 3 to 5 days at 28°C in the dark, fungal colonies displaying morphological characteristics of Fusarium spp. (2) were purified by transferring single spores and identified to species level by morphological characteristics (2), and DNA sequence analysis of translation elongation factor-1α (TEF) and β-tubulin genes. A large number of Fusarium species (mainly F. graminearum species complex, F. verticillioides, and F. proliferatum) were identified. These Fusarium species are the main causal agents of maize ear rot (2). Morphological characteristics of six strains from Anhui, Hubei, and Yunnan provinces were found to be identical to those of F. kyushuense (1), which was mixed with other Fusarium species in the natural infection in the field. Colonies grew fast on PDA with reddish-white and floccose mycelia. The average growth rate was 7 to 9 mm per day at 25°C in the dark. Reverse pigmentation was deep red. Microconidia were obovate, ellipsoidal to clavate, and 5.4 to 13.6 (average 8.8) μm in length. Macroconidia were straight or slightly curved, 3- to 5-septate, with a curved and acute apical cell, and 26.0 to 50.3 (average 38.7) μm in length. No chlamydospores were observed. Identity of the fungus was further investigated by sequence comparison of the partial TEF gene (primers EF1/2) and β-tubulin gene (primers T1/22) of one isolate (3). BLASTn analysis of the TEF amplicon (KC964133) and β-tubulin gene (KC964152) obtained with cognate sequences available in GenBank database revealed 99.3 and 99.8% sequence identity, respectively, to F. kyushuense. Pathogenicity tests were conducted twice by injecting 2 ml of a prepared spore suspension (5 × 10 spores/ml) into maize ears (10 per isolate of cv. Zhengdan958) through silk channel 4 days post-silk emergence under field conditions in Wuhan, China. Control plants were inoculated with sterile distilled water. The ears were harvested and evaluated 30 days post-inoculation. Reddish-white mold was observed on inoculated ears and the infected kernels were brown. No symptoms were observed on water controls. Koch's postulates were fulfilled by re-isolating the pathogen from infected kernels. F. kyushuense, first described on wheat in Japan (1), has also been isolated from rice seeds in China (4). It was reported to produce both Type A and Type B trichothecene mycotoxins (1), which cause toxicosis in animals. To our knowledge, this is the first report of F. kyushuense causing maize ear rot in China and this disease could represent a serious risk of yield losses and mycotoxin contamination in maize and other crops. The disease must be considered in existing disease management practices. References: (1) T. Aoki and K. O'Donnell. Mycoscience 39:1, 1998. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (3) F. Van Hove et al. Mycologia 103:570, 2011. (4) Z. H. Zhao and G. Z. Lu. Mycotaxon 102:119, 2007.
2009年9月至2012年10月,在中国22个省份开展了调查,以确定玉米上镰刀菌的种群结构,这些省份的个别田块病害发病率在5%至20%之间。从田间采集有明显镰刀菌穗腐症状的玉米穗(穗尖有白色至粉红色或鲑鱼色霉菌)。有症状的籽粒进行表面消毒(在0.1% HgCl中处理1分钟,在70%乙醇中处理30秒,然后用无菌蒸馏水冲洗三次),干燥后置于PDA培养基上。在28°C黑暗条件下培养3至5天后,将呈现镰刀菌形态特征的真菌菌落通过转移单个孢子进行纯化,并通过形态特征以及翻译延伸因子-1α(TEF)和β-微管蛋白基因的DNA序列分析鉴定到种水平。鉴定出大量镰刀菌物种(主要是禾谷镰刀菌复合种、轮枝镰孢和层出镰孢)。这些镰刀菌物种是玉米穗腐的主要病原菌。发现来自安徽、湖北和云南省的六个菌株的形态特征与九州镰孢相同,在田间自然感染中它与其他镰刀菌物种混合存在。菌落在PDA上生长迅速,有红白相间的絮状菌丝体。在25°C黑暗条件下平均生长速率为每天7至9毫米。背面色素沉着为深红色。小型分生孢子倒卵形、椭圆形至棒形,长度为5.4至13.6(平均8.8)微米。大型分生孢子直或稍弯曲,3至5分隔,顶端细胞弯曲且尖锐,长度为26.0至50.3(平均38.7)微米。未观察到厚垣孢子。通过对一个分离株的部分TEF基因(引物EF1/2)和β-微管蛋白基因(引物T1/22)进行序列比较,进一步研究了该真菌的同一性。对TEF扩增子(KC9641,33)和β-微管蛋白基因(KC964152)与GenBank数据库中可用的同源序列进行BLASTn分析,结果显示与九州镰孢的序列同一性分别为99.3%和99.8%。在中国武汉的田间条件下,在吐丝后4天通过花丝通道向玉米穗(每个郑单958品种分离株10个)注射2毫升制备好的孢子悬浮液(5×10孢子/毫升),进行了两次致病性试验。对照植株接种无菌蒸馏水。接种后30天收获玉米穗并进行评估。接种的玉米穗上观察到红白相间的霉菌,感染的籽粒为褐色。水对照未观察到症状。通过从感染的籽粒中重新分离病原菌,满足了柯赫氏法则。九州镰孢首次在日本的小麦上被描述,在中国也从水稻种子中分离到。据报道它会产生A型和B型单端孢霉烯族毒素,可导致动物中毒。据我们所知,这是九州镰孢在中国引起玉米穗腐的首次报道,这种病害可能对玉米和其他作物的产量损失以及霉菌毒素污染构成严重风险。在现有的病害管理措施中必须考虑这种病害。参考文献:(1) T. Aoki和K. O'Donnell。《真菌科学》39:1,1998年。(2) J. F. Leslie和B. A. Summerell。《镰刀菌实验室手册》。布莱克威尔出版社,艾姆斯,爱荷华州,2006年。(3) F. Van Hove等人。《真菌学》103:570,2011年。(4) Z. H. Zhao和G. Z. Lu。《真菌分类学报》102:119,2007年。