萎缩芽胞杆菌LB6-2抑制立枯丝核菌活性物质的鉴定

Identification of Active Substances of Bacillus atrophaeus LB6-2 to Inhibit Rhizoctonia solani

本研究采用平板对峙法、菌丝生长速率法和菌核萌发抑制法筛选出一株细菌LB6-2,对立枯丝核菌的菌丝生长抑制率为71.76%,菌核萌发抑制率为100.00%。通过观察形态学特征、分析理化性质结合16S r DNA扩增序列分析对该菌株鉴定为萎缩芽胞杆菌Bacillusatrophaeus(OK639005.1)。采用大孔树脂法提取发酵产物并进行HPLC制备,对获得的14个组分进行活性测定确定其活性组分,从活性组分中制备获得3个单品化合物LBN4-2、LBN4-4、LBN4-5,经HPLC-MS、NMR分析和活性测定结果表明,化合物LBN4-2为Cyclo(Pro-Val)、LBN4-4为Cyclo(D-Leu-L-Pro)和LBN4-5为Cyclo(L-prolyl-D-phenylalanyl),3个化合物的浓度在4 mmol/L时对立枯丝核菌菌丝生长的抑制率分别为86.16%、84.87%和83.10%,浓度在2 mmol/L以上时,对立枯丝核菌菌核萌发的抑制率为100%。由此可见,萎缩芽胞杆菌LB6-2具有作为防治立枯丝核菌生防菌剂研发的潜力。

Bacteria LB6-2 was selected against Rhizoctonia solani by plate face-off method,mycelium growth rate method and sclerotium germination inhibition method.It showed the inhibition rate of mycelium growth and sclerotium germination of strain LB6-2 was 71.76%and 100.00%.The strain was identified Bacillus atrophaeus(OK639005.1)by observing morphological characteristics,analyzing physicochemical properties and 16S rDNA amplification.The fermentation products were extracted by macroporous resin method and prepared by HPLC.Fourteen active components were determined by activity determination,and the single compounds were prepared and analyzed by HPLC-MS,NMR and activity determination.The compounds LBN4-2,LBN4-4 and LBN4-5 were identified as Cyclo(Pro-Val),Cyclo(D-Leu-L-Pro)and Cyclo(L-prolyl-D-phenylalanyl),respectively.The activity assay of the three compounds showed that the inhibition rates of hyphal growth were 86.16%,84.87%and 83.10%at concentration was 4 mmol/L,respectively.While the concentration of the three compounds were above 2 mmol/L,the inhibition rates of hyphal germination were 100%.In summary,Bacillus atrophaeus LB6-2 has the potential to be used as a biocontrol agent against Rhizoctonia solani.