草酸青霉A1菌株的鉴定及对苹果4种镰孢病菌的拮抗作用

从连作条件下的平邑甜茶健康根系分离得到1株内生真菌A1,通过形态学特征、培养特性观察鉴定该菌属于青霉属,将其ITS r DNA序列和β-tubulin序列与Gen Bank中已知标准菌株的ITS r DNA序列和β-tubulin序列进行比对,并利用MEGA5.05软件构建青霉菌A1菌株进化树。结果表明,青霉菌A1与草酸青霉Penicillium oxalicum(KF152942.1,KC344990.1)序列同源性最高,为100%,最终将青霉菌A1鉴定为草酸青霉A1。在马铃薯培养基(PDA)和玉米粉培养基上分别将内生真菌A1与尖孢镰孢菌、串珠镰孢菌、层出镰孢菌和腐皮镰孢菌进行对峙试验,结果在PDA培养基上对菌丝生长抑制率分别为42.0%、74.0%、47.0%和54.5%,拮抗系数达到Ⅱ级或Ⅰ级,在玉米粉培养基上菌丝生长抑制率分别为60.5%、82.2%、68.4%和53.4%,拮抗系数达到Ⅱ级或Ⅰ级,说明草酸青霉A1对4种苹果连作障碍病原镰孢属真菌具有较好的抑制作用。平邑甜茶盆栽试验结果表明,与连作土对照相比,A1菌肥显著促进了连作平邑甜茶幼苗鲜样质量、干样质量的增加,分别增加2.53倍和2.35倍,说明草酸青霉A1菌肥能在一定程度上减轻苹果连作障碍。使用实时荧光定量技术对土壤中的尖孢镰孢菌基因拷贝数进行检测,发现A1菌肥处理的土壤中尖孢镰孢菌的基因拷贝数量大幅下降,说明草酸青霉菌A1可以抑制其生长。A1菌株可作为防控苹果连作障碍的拮抗菌进一步研究。

Identification of Penicillium oxalicum A1 Strain and Antagonistic Effects on Four Species of Fusarium Pathogen of Apple

A strain of endophytic fungi A1 was isolated from the roots of Malus hupehensis Rehd. seedlings by using dilution separation methods after surface disinfection and grinding，and was identified as Penicillium oxalicum by observation of morphology，cultural characteristics compared the ITS rDNA sequences and β-tubulin sequences with the known standard strains sequences that of GenBank，and the MEGA5.05 software was used to construct the phylogenetic tree of the A1 strain. The similarity of the ITS rDNA and β-tubulin sequences between A1 strain and Penicillium oxalicum（KF152942.1，KC344990.1）were 100%，so it was finally identified as the nearest genetic relationship with Penicillium oxalicum. By using a potato medium and a corn meal medium，endophytic fungi A1 was respectively confrontation test with Fusarium oxysporum，F. moniliforme，F. proliferatum and F. solani. It could inhibit growth of hypha of F. oxysporum，F. moniliforme，F. proliferatum and F. solani to a variable extent，and increased by 42.0%，74.0%，47.0% and 54.5% on the potato medium，and the coefficient of antagonism could be reached Ⅱ or Ⅰ. Similarly，growth of hypha was increased by 60.5%，82.2%，68.4% and 53.4% and the coefficient of antagonism could be reached Ⅱ or Ⅰon the corn meal medium. That Penicillium oxalicum A1 had better inhibitory effect on four kinds of Fusarium sp. of the apple replant disease. Pot experimental results showed that fresh weight and dry weight were obviously increased by function of A1 manure，and were 2.53 and 2.35 times respectively higher than the control，so A1 manure could reduce the apple replant disease to a certain extent. At the same time soil F. oxysporum gene copy number was detected by using real-time fluorescence quantitative technique and found that Penicillium oxalicum A1 could inhibit the growth of F. oxysporum，and could be ulteriorly studied as a antagonism strain to control the apple replant disease.