| Summary: | 碩士 === 國立臺灣大學 === 生物科技研究所 === 105 === Bacterial wilt, which is caused by the soil-borne pathogen Ralstonia solanacearum (Rs), is one of the most serious diseases in tomato plant. Rs can invade plant roots and spread quickly into the xylem vessels toward the whole plant tissue through the vascular system. Symptoms of this disease include stunting, vascular browning, wilting, and even causing plant death. In Taiwan, the disease caused by Rs is managed only by using beneficial microorganisms. Nowadays, the study of endophytic bacteria in plants and their applications on biological control of plant pathogens are gaining a lot of interest. Bacillus amyloliquefaciens BPD1 (Ba-BPD1) has been shown to have plant-growth-promoting effects and the ability to produce several types of antimicrobial compounds against plant pathogens. In this study, we found that Ba-BPD1 has the biggest inhibition zone against Rs when incubated in LMS medium. Furthermore, after treatment with 5 μl/ml of the culture supernatant of Ba-BPD1, the Rs population decreased to 20% and their motility activity also decreased as well as the morphology of Rs cells. To identify the antimicrobial compounds probably present in the supernatant, six gene fragments of antimicrobial compounds, including bacillaene, macrolactin, difficidin, bacillomycin D, bacillibactinc, and bacillycin, were amplified from Ba-BPD1 through PCR analysis. Moreover, we also found that volatile organic compounds (VOCs) produced by Ba-BPD1 had negative effects on the colony size, morphology, and motility activity of Rs. In the pot experiments, we found that treatment with Ba-BPD1 could reduce the Rs disease severity by 39.6% in tomato cv. Santa. To further understand the mechanism of Ba-BPD1 inhibition of tomato plant wilting, we evaluated the pathogen populations in soil and the expression levels of the defense-related genes in leaf after Ba-BPD1 treatment. The results revealed that the soil pathogen populations were not significantly decreased after Ba-BPD1 treatment. On the other hand, gene expression levels of the AOC and ACO genes, which are involved in the JA and ET pathways, respectively, were increased after pathogen inoculation in Ba-BPD1 treatment. On the other hand, the growth of tomato seedling was influenced by Ba-BPD1 inoculation. Last but not the least, our experiments showed that Ba-BPD1 could colonize into the tomato plant and the populations reached up to 3.7×104 CFU/g tissue at the basal parts of the inoculation site (0-2 cm). Otherwise, composition of the endophyte community in root and diversity of the endophyte community in root and stem tissues changed in response to inoculation were correlated with tomato resistance to the disease. Thus, this study demonstrated that Ba-BPD1 has a potential to be used as a biocontrol agent in controlling tomato plant wilting caused by Rs as it is able to produce antimicrobial compounds, induce plant systemic resistance, colonize plant tissues in a stable manner, modulate the endophyte community of the host and also promote plant growth.
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