Colonization of bacterial endophyte-Bacillus amyloliquefaciens SPX1 in tomato and its potential for controlling bacterial wilt

• Main Authors: Hui-Ru Pan, 潘蕙如
• Other Authors: Wen-Hsin Chung
• Format: Others
• Language: zh-TW
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/75801048771835723538

碩士 === 國立中興大學 === 植物病理學系所 === 103 === Endophytic bacteria (EB) invades the living plants tissues without causing apparent symptoms of diseases; meanwhile, several EBs can protect host from phytopathogens. The EB of Bacillus amyloliquefacines SPX1 isolated from sweet potato was used in this study. The results of introduction tests indicated that the populations and colonization area of SPX1 strain had no significant difference at 0-6 cm tissues site of tomato after inoculated into plant tissues by hypocotyl cutting inoculation method for 30 or 120 min. Besides, hypocotyl cutting inoculation method was better than drenching method for SPX1 strain colonizing in tomato tissues. Nutrient utilization showed that SPX1 preferred in the media added with carbon source such as sucrose and fructose, and nitrogen sources such as glutamate, ammonium chloride and ammonium sulfate. Moreover, SPX1 also can grow in lysogeny broth cantained NaCl ranged from 0 to 8 %. The pH test showed that pH 5 to 9 were suitable for the growth of SPX1. Host range tests pointed out that the reisolation rate of SPX1 strain could reach 80 to 100 % in sweet potato, cucumber, tomato, asparagine bean, pepper, and melon at 14-day after inoculation. Chemotaxis assays revealed that SPX1 is specifically attracted by sucrose, fructose, arginine and tyrosine. In greenhouse experiment, the SPX1 strain only showed the ability in controlling the bacterial wilt (BW) of tomato with hypocotyl cutting inoculation method, and could decrease the disease severity by 32 % at 28-day after transplated into the Ralstonia solanacearum-infested soil (106 cfu/g soil). Furthermore, the SPX1 strain treatments could reduce the disease severity of BW by 50 % by using hypocotyl cutting inoculation combined with drenching inoculation method. The ability effect of SPX1 in controlling BW also shown the same efficacy in different tomato cultivars (Santa and Hualien Asveg 21) with drenching and hypocotyl cutting inoculation method. The experiment of distribution efficacy of SPX1 strain showed that SPX1 could colonize in the whole stem tissues of vegetable sweet potato and tomato at 28-day after inoculation; however, the population of SPX1 could reached 103-104 cfu/g tissue at the basal parts of the inoculation site (0-3 or 4 cm) than in higher tissues site based on hypocotyl-cutting seedlings. Morever, the colonization ability of SPX1 on different tomato cultivar showed that the SPX1 more preferred colonized in Santa and Hualien Asveg 21 than Farmers 301. In order to observing the distribution of SPX1 in tomato plant tissues, the transformed line, SPX1/DH5α (pAD43-25) with green fluorescent protein marker, was used in this study. The SPX1/DH5α (pAD43-25) could be observed on root surface and colonize on axils of lateral roots dominantly. Furthermore, the SPX1/DH5α (pAD43-25) basically colonized in xylem tissues, and transmitted intercellarly from xylem to cortex, and trichome of epidermis at 7-day after inoculation with hypocotyl cutting inoculation method. Thus, the B. amyloliquefaciens SPX1 strain could be used as a biocontrol agent in controlling BW of tomato in greenhouse experiment and colonize in tomato tissue stably. On the other hand, the SPX1 could colonized in several crops, and the controlling ability of BW on each crop should be further studied.