苹果盘二孢的分离培养研究

苹果盘二孢Marssonina coronaria引起的褐斑病是造成我国苹果树早期落叶的主要原因,由于该病菌分离培养困难,阻碍了对其生物学特性的研究,进而影响了对其防治机理和流行规律的研究。本研究应用4种培养基质,探索了3种方法对苹果盘二孢的分离效果。结果表明,3种方法均可分离到病原菌,但组织块分离法和分生孢子团分离法成功率仅有10%左右,而单孢分离法污染少,成功率高达到90%以上,明显优于其他两种方法。不同培养基上菌落形态、大小和产孢情况差异也很大,培养1个月(25℃)后PDA上菌落黑褐色隆起,表面蚯蚓粪状,无气生菌丝,无子实体和基内菌丝;10%V8培养基上菌落中央隆起,黑褐色,表面生少量气生菌丝,边缘放射状,基内菌丝深褐色,有子实体;苹果叶片葡萄糖琼脂培养基(LDA)上菌落平坦,黄褐色,表面生茂密的金黄色气生菌丝,基内菌丝深褐色,有子实体;苹果叶片煎汁葡萄糖琼脂培养基(LEDA)上菌落有明显的不规则隆起,黄褐色至黑褐色,表面生少许气生菌丝,菌落生长缓慢,无基内菌丝,分生孢子盘菌落表面生,菌落直径仅2mm左右,而在其他培养基上的菌落直径可达6-8mm,说明培养基质、分离方法均对苹果盘二孢的分离培养和生长发育有明显的影响。

Culture study of Marssonina coronaria from diseased apple leaves

Marssonina coronaria is the causal agent of Marssonina leaf blotch of apple which leads severe defoliation starting at early summer. It is difficult to obtain the pure culture of this fungus because of its slow growth. Three isolation methods were compared by using 4 culture media in this study. The results showed that each isolation method was successful to obtain the pure culture of fungus. However, single spore isolation was much easier, less contamination and more successful than isolation using diseased leaf segment and conidia mass from aeervuli. The isolation methods of the latter only showed successful rate of 10%. Additionally, the growth of the pathogen was obviously affected by cultural media. The morphological characters such as colony color and diameter, and production of fruit body of the fungus growing on PDA, 10% V8, apple leaves dextrose agar (LDA) and apple leaves extraction dextrose agar (LEDA) media were different. The fungus formed colonies of about 8ram in diameter and did not produce fruit bodies and aerial hyphae in 1 month incubation on PDA. However it formed the similar size colonies as on PDA and produced fruit bodies and aerial hyphae on 10% V8 and LDA media in the same incubation period. Very small colonies (2mm in diameter) and fruit bodies were found on LEDA media in the same incubation condition. These results indicated that successful isolation of M. coronaria from apple leaves depended on suitable isolation method and cultural media as well as fresh samples.