Chen Y, Wang W-X, Zhang A-F, Yang X, Xu Y-L
Institute of Plant Protection and Agro-products Safety, Anhui Academy of Agricultural Sciences, Hefei 230031, China.
Anhui Academy of Agricultural Sciences, Hefei 230031, China.
Plant Dis. 2013 Apr;97(4):558. doi: 10.1094/PDIS-08-12-0710-PDN.
Loquat, Eriobotrya japonica (Thunb.) Lindl., is an important fruit that is widely planted and used as an ornamental in Jingxian, Anhui Province, China. Loquat branches with severely spotted leaves and fruits were observed in this region in 2012. Symptoms on leaves consisted of small (0.5 to 1.2 cm in diameter), circular to oblong, greenish-brown lesions that coalesced to form isolated or confluent, dark brown spots. On fruit, the disease appeared as circular to elongated, sunken spots. Expanding lesions spread over the surface resulting in death of the fruit. Acervuli were observed within lesions. Isolations from symptomatic tissue onto potato dextrose agar (PDA) medium consistently yielded white fungal colonies of sparse aerial mycelium with acervuli containing black, slimy spore masses on the surface. The colony reached 8.0 cm diameter after 7-day culture on PDA at 24°C. Conidia produced in the culture were five-celled, narrow fusiform, straight or slightly curved, with a tapering base and 2 to 4 hyaline appentages (apical appentages measured 15 to 34 μm long and a single basal appentage was 5 to 9 μm long). Conidia were 24 to 32 × 5 to 8 μm with median cells 15 to 20 μm and two hyaline, cylindrical to conical apical cells typical of Pestalotiopsis spp. (3). A total of 12 isolates were obtained by isolation from the diseased fruit or leaves. Genomic DNA from the fungal isolates was purified using a DNA Gel Extraction Kit (AxyPrep, Hangzhou, China), and applied to a DNA Engine System PTC-200 (BIO-RAD, Watertown, MA) with ITS1 and ITS4 internal transcribed spacer (ITS) universal primers. The amplified sequences (533 bp) were analyzed together with other Pestalotiopsis sequences (1). ITS from all 12 of the fungal isolates were identical (99.5% similarity) to each other and to isolates of Pestalotiopsis theae, which infects tea trees in China (2). To demonstrate pathogenicity, suspensions (prepared in distilled water) of 10 conidia ml of each isolate were sprayed on the loquat leaves in vivo and mature fruits in vitro. Distilled water was used as the control. More than 20 leaves and 10 mature fruits were sprayed for the treated and control plants, respectively, and the inoculation tests were repeated twice. The inoculated plants and fruit were kept in a humidity chamber for 7 days. Approximately 50% of the inoculated leaves and fruits developed blight symptoms similar to natural infections. P. theae was reisolated from diseased plants to complete Koch's postulates. Control plants sprayed with distilled water remained symptomless. There is a previous study reporting that P. guepini infected loquat in Argentina (4); however, to our knowledge, this is the first report of P. theae causing leaf and fruit spots on loquat in China. References: (1) R. Jeewon et al. Mol. Phylogenet. Evol. 25:378, 2002. (2) J. Y. Lu. Diagnosis of plant diseases. Page 194 in: Pestalotiopsis. J. Y. Lu, Z. G. Xu, Y. X. Chen, D. R. Shen, X. B. Zheng, and Y. Q. Cao, eds. China Agriculture Press, Beijing, 1995. (3) T. R. Nag Raj. Coelomycetous Anamorphs with Appendage-Bearing Conidia. Mycologue Publications, Waterloo, Canada, 1993. (4) A. E. Perelló and S. Larran. Plant Dis. 83:695, 1999.
枇杷,学名Eriobotrya japonica (Thunb.) Lindl.,是一种重要的水果,在中国安徽省泾县广泛种植并用作观赏植物。2012年,该地区发现了枇杷枝叶和果实上出现严重斑点的情况。叶片上的症状表现为小的(直径0.5至1.2厘米)、圆形至椭圆形、绿褐色病斑,这些病斑会融合形成孤立的或连片的深褐色斑点。果实上,病害表现为圆形至长形的凹陷斑点。扩展的病斑蔓延至果实表面,导致果实死亡。在病斑内观察到分生孢子盘。将有症状的组织接种到马铃薯葡萄糖琼脂(PDA)培养基上,始终能长出白色真菌菌落,其气生菌丝稀疏,表面有含黑色、粘性孢子团的分生孢子盘。在24°C条件下于PDA上培养7天后,菌落直径达8.0厘米。培养物中产生的分生孢子为五细胞,窄梭形,直或稍弯曲,基部渐细,有2至4个透明附属物(顶端附属物长15至34μm,单个基部附属物长5至9μm)。分生孢子大小为24至32×5至8μm,中间细胞长15至20μm,有两个透明、圆柱形至圆锥形的顶端细胞,这是拟盘多毛孢属(Pestalotiopsis spp.)的典型特征。从患病果实或叶片上共分离得到12个菌株。使用DNA凝胶提取试剂盒(AxyPrep,中国杭州)从真菌菌株中纯化基因组DNA,并将其应用于带有ITS1和ITS4内转录间隔区(ITS)通用引物的DNA Engine System PTC - 200(BIO - RAD,美国马萨诸塞州沃特敦)。扩增的序列(533 bp)与其他拟盘多毛孢序列一起进行分析。所有12个真菌菌株的ITS彼此相同(相似度99.5%),并且与感染中国茶树的茶拟盘多毛孢(Pestalotiopsis theae)菌株相同。为证明致病性,将每个菌株10个分生孢子/毫升的悬浮液(用蒸馏水配制)分别喷雾接种到活体枇杷叶片和离体成熟果实上。以蒸馏水作为对照。处理组和对照组植株分别喷雾接种20多片叶子和10个成熟果实,接种试验重复两次。接种后的植株和果实置于湿度箱中7天。约50%的接种叶片和果实出现与自然感染相似的枯萎症状。从患病植株上重新分离得到茶拟盘多毛孢,以完成柯赫氏法则验证。喷洒蒸馏水的对照植株无症状。此前有研究报道阿根廷的枇杷被古氏拟盘多毛孢(P. guepini)感染;然而,据我们所知,这是中国关于茶拟盘多毛孢引起枇杷叶斑和果斑的首次报道。参考文献:(1) R. Jeewon等人,《分子系统发育与进化》25:378,2002年。(2) 卢耀增,《植物病害诊断》。载于《拟盘多毛孢》,卢耀增、徐宗刚、陈义勋、沈德儒、郑小波、曹以勤编,中国农业出版社,北京,1995年,第194页。(3) T. R. Nag Raj,《具附属物分生孢子的腔孢菌无性型》,Mycologue Publications,加拿大滑铁卢,1993年。(4) A. E. Perelló和S. Larran,《植物病害》83:695,1999年。