多花黄精内生贝莱斯芽胞杆菌的分离鉴定及其抗菌与促生作用分析

Isolation and identification of antagonistic endophytic Bacillus velezensis from Polygonatum cyrtonema Hua and analysis of its antimicrobial and growth-promoting activity

为鉴定药用植物内生菌的种类,采用稀释涂布平板法从多花黄精Polygonatum cyrtonema Hua的根、茎、叶内分离出11株内生菌菌株,经平板对峙法筛选出对尖孢镰刀菌Fusarium oxysporum具有拮抗作用的菌株ZJU-3。经形态学、生理生化鉴定及16S rDNA序列分析,初步确定该菌株为贝莱斯芽胞杆菌Bacillus velezensis。通过基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)分析,发现该菌株能产生表面活性素(surfactin)、泛革素(fengycin)和伊枯草菌素(iturin)三类脂肽类化合物。利用盐酸沉淀和甲醇抽提法获得了菌株ZJU-3发酵液中脂肽粗提物。该脂肽粗提物可明显抑制尖孢镰刀菌菌丝的生长,抑制率达到51.6%。经液相色谱-质谱联用(LC-MS)分析表明菌株ZJU-3可产生吲哚乙酸、激动素、玉米素、赤霉素等多种植物激素。温室栽培试验结果发现该菌株对多花黄精具有显著的促生效果,根长和单株根数明显高于对照组。本研究可为进一步应用该菌株提供理论依据。

In order to identify the species of endophytic bacteria in medicinal plants,11 strains of endophytes were isolated from the roots,stems and leaves of Polygonatum cyrtonema Hua,and the strain ZJU-3 was found to be antagonistic to Fusarium oxysporum.The strain ZJU-3 was identified as Bacillus velezensis by morphological,physiological,biochemical and molecular methods.It could produce lipopeptides including surfactin,fengycin and iturin,based on analysis by MALDI-TOF-MS.The crude extracts of lipopeptide,obtained by the hydrochloric acid precipitation and methanol extraction,could inhibit the mycelial growth of F.oxysporum with an inhibition rate of 51.6%.The analysis by LC-MS showed that the strain ZJU-3 could produce plant hormones IAA,kinetin,zeatin and GA 3.In pot experiments,the strain ZJU-3 had a significant effect on the growth of P.cyrtonema,and the root length and number were significantly higher than those of the control group.This study provides a theoretical basis for further application of the strain.