Yin L F, Chen S N, Yuan N N, Zhai L X, Li G Q, Luo C X
Department of Plant Pathology, College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
Kunming Sijiqing Pesticide Co., Ltd., Kunming, 655000, China.
Plant Dis. 2013 Sep;97(9):1255. doi: 10.1094/PDIS-03-13-0310-PDN.
Brown rot of peach (Prunus persica) in China has been reported to be caused by at least three Monilinia species (1). In the present study, peaches with symptoms resembling brown rot caused by Monilinia species were collected from commercial orchards in the northwestern province of Gansu in August 2010, the southwestern province of Yunnan in July 2011, and in the central province of Hubei in July 2012. Affected fruit showed the typical symptoms of brown rot with zones of sporulation. Fungal isolates were single-spored and cultured on potato dextrose agar (PDA). Colonies showed grayness with concentric rings of sporulation after incubation at 25°C in the dark. Mean mycelial growth of isolates YHC11-1a and YHC11-2a from Yunnan, GTC10-1a and GTC10-2a from Gansu, and HWC12-14a and HWC12-23a from Hubei, was 4.6 ± 0.4 and 7.5 ± 0.7 cm after 3 and 5 days incubation, respectively. Conidia were lemon shaped and formed in branched monilioid chains, and the mean size was 9.3 (6.7 to 11.5) × 12.5 (7.9 to 17.8) μm, which was consistent with the characteristics of Monilinia fructicola (1,2). The species identification was confirmed by sequencing of the ribosomal ITS sequences. The ribosomal ITS1-5.8S-ITS2 region was amplified from each of the six isolates using primers ITS1 and ITS4 (3). Results indicated that the ITS sequences of these isolates were identical and showed the highest similarity (100%) with M. fructicola ITS sequences from isolates collected in China (GenBank Accession Nos. HQ893748, FJ515894, and AM887528), Slovenia (GU967379), Italy (FJ411109), and Spain (EF207423). The pathogen was also confirmed to be M. fructicola based on the detection of an M. fructicola- specific band (534 bp) using a PCR-based molecular tool developed for distinguishing Chinese Monilinia species affecting peach (1). Pathogenicity was tested on surface-sterilized, mature peaches (Shui Mi Tao) with representative isolates. Fruits were holed at three equidistant positions to a depth of 5 mm using a sterile cork borer. Mycelial plugs (5 mm in diameter) from the periphery of a 4-day-old colony of each isolate were placed upside down into each hole, control fruits received water agar. After 3 days of incubation at 22°C in a moist chamber, inoculated fruits developed typical brown rot symptoms while control fruits remained healthy. Pathogens from the inoculated fruit were confirmed to be M. fructicola based on morphological characteristics. To our knowledge, this is the first report of occurrence of M. fructicola in Gansu, Yunnan, and Hubei provinces, thousands of kilometers away from eastern China where occurrence of peach brown rot caused by M. fructicola has been confirmed (2,4). The results indicated the further geographical spread of the M. fructicola in China. References: (1) M. J. Hu et al. Plos One 6(9):e24990, 2011. (2) M. J. Hu et al. Plant Dis. 95:225, 2011. (3) T. J. White et al. Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. Academic Press, San Diego, 1990. (4) X. Q. Zhu et al. Plant Pathol. 54:575, 2005.
据报道,中国桃(Prunus persica)的褐腐病至少由三种链核盘菌属(Monilinia)真菌引起(1)。在本研究中,2010年8月从中国西北部甘肃省的商业果园、2011年7月从西南部云南省的商业果园以及2012年7月从中部湖北省的商业果园采集了有类似链核盘菌属真菌引起的褐腐病症状的桃子。受感染的果实呈现出带有产孢区域的典型褐腐病症状。真菌分离株进行单孢分离,并在马铃薯葡萄糖琼脂(PDA)上培养。在25°C黑暗条件下培养后,菌落呈现灰色并带有同心产孢环。来自云南的分离株YHC11 - 1a和YHC11 - 2a、来自甘肃的GTC10 - 1a和GTC10 - 2a以及来自湖北的HWC12 - 14a和HWC12 - 23a,在培养3天和5天后,平均菌丝生长分别为4.6±0.4厘米和7.5±0.7厘米。分生孢子呈柠檬形,形成于分枝状的念珠状链中,平均大小为9.3(6.7至11.5)×12.5(7.9至17.8)微米,这与桃褐腐病菌(Monilinia fructicola)的特征一致(1,2)。通过核糖体ITS序列测序确认了物种鉴定。使用引物ITS1和ITS4(3)从六个分离株中的每一个扩增核糖体ITS1 - 5.8S - ITS2区域。结果表明,这些分离株的ITS序列相同,并且与从中国(GenBank登录号HQ893748、FJ515894和AM887528)、斯洛文尼亚(GU967379)、意大利(FJ411109)和西班牙(EF207423)采集的桃褐腐病菌的ITS序列具有最高相似度(100%)。基于一种为区分影响中国桃树的链核盘菌属真菌而开发的基于PCR的分子工具检测到桃褐腐病菌特异性条带(534 bp),也证实了该病原菌为桃褐腐病菌(1)。使用代表性分离株对表面消毒的成熟桃子(水蜜桃)进行致病性测试。使用无菌打孔器在果实三个等距位置打孔至5毫米深。将每个分离株4日龄菌落边缘的菌丝块(直径5毫米)倒置放入每个孔中,对照果实接种水琼脂。在湿度箱中22°C培养3天后,接种果实出现典型的褐腐病症状,而对照果实保持健康。根据形态特征,确认接种果实中的病原菌为桃褐腐病菌。据我们所知,这是桃褐腐病菌在甘肃省、云南省和湖北省发生的首次报道,这些省份距离中国东部已确认有桃褐腐病菌引起的桃褐腐病发生的地区有数千公里之遥(2,4)。结果表明桃褐腐病菌在中国进一步的地理扩散。参考文献:(1)M. J. Hu等人,《公共科学图书馆·综合》6(9):e24990,2011年。(2)M. J. Hu等人,《植物病害》95:225,2011年。(3)T. J. White等人,《用于系统发育学的真菌核糖体RNA基因的扩增和直接测序》,学术出版社,圣地亚哥,1990年。(4)X. Q. Zhu等人,《植物病理学》54:575,2005年。