棉花根部拮抗枯萎病菌或黄萎病菌的可培养内生细菌多样性分析

为解析棉花内生细菌的种群结构,更好地开发利用生防内生细菌,以棉花枯萎病菌、黄萎病菌为指示病原真菌,对常抗棉和泗棉3号根部分离获得的内生细菌进行拮抗活性筛选,获得87株至少对其中1个指示病原真菌具有拮抗作用的内生细菌。ARDRA分析(Amplified ribosomal DNA restriction analysis)和16S r DNA序列测定表明棉花根部可培养内生拮抗细菌可分为13个ARDRA类群,分属于芽孢杆菌属(Bacillus)、短芽孢杆菌属(Brevibacillus)、伯克霍尔德氏菌属(Burkholderia)、农杆菌属(Agrobacterium)、中华根瘤菌属(Sinorhizobium)、不动杆菌属(Acinetobacter)和假单孢菌属(Pseudomonas)7个属。其中常抗棉根部内生拮抗细菌分属于芽孢杆菌属、短芽孢杆菌属、伯克霍尔德氏菌属和农杆菌属,而泗棉3号根部内生拮抗细菌则分属于芽孢杆菌属、伯克霍尔德氏菌属、中华根瘤菌属、不动杆菌属和假单孢菌属,二者的优势种群都是芽孢杆菌属细菌。BOX-PCR分析显示这些内生拮抗细菌具有丰富的遗传多样性。通过测定这些内生拮抗细菌对棉花黄萎病菌和棉花枯萎病菌的拮抗力大小,以及是否具有产纤维素酶、几丁质酶、蛋白酶和嗜铁素的活性,表明这些内生拮抗细菌具有不同的生防作用机制。

Diversity Analysis of Culturable Endophytic Bacteria with Antagonistic Activity against Fusarium oxysporum f. sp. vasinfectum or Verticillium dahliae from the Roots of Gossypium hirsutum

The aim of this study was to understand the diversity and antagonistic potential of cotton endophytic bacteria and to utilize endophytic bacteria for biocontrol of cotton vascular disease. Through antagonistic testing of endophytic bacteria against Fusarium oxysporum f.sp. vasinfectum and Verticillium dahliae, 87 strains of culturable endophytic antagonistic bacteria from the roots of Changkangmian and Simian 3 cotton cultivars were identified. Based on the amplified ribosomal DNA restriction analysis(ARDRA), these culturable endophytic antagonistic bacterial strains could be classified into 13 ARDRA groups. The 16S rRNA gene fragments of representative strains from different groups were sequenced and subjected to a BLAST search to identify similar strains. The results showed that the endophytic antagonistic bacteria most likely belonged to genera Bacillus, Brevibacillus, Burkholderia, Agrobacterium, Sinorhizobium, Acinetobacter and Pseudomonas. The endophytic antagonistic bacteria from the roots of Changkangmian belonged to genera Bacillus, Brevibacillus, Burkholderia and Agrobacterium, while the endophytic antagonistic bacteria from the roots of Simian 3 belonged to genera Bacillus, Burkholderia, Sinorhizobium, Acinetobacter and Pseudomonas. The most prominent genus from both varieties was Bacillus. BOX-PCR was used to study the diversity of the endophytic antagonistic bacteria and showed that a broad genetic diversity existed even within the same ARDRA group. Strains were tested for antagonistic activity against Fusarium oxysporum f. sp. vasinfectum and Verticillium dahliae and production of antibiotic substances including cellulase, chitinase, protease and siderophores. The results indicated that the biocontrol mecha nisms of these culturable endophytic antagonistic bacteria were different.