Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/31992
標題: 蕈狀芽孢桿菌防治番茄萎凋病之相關機制分析
Related mechanisms analyses for controlling tomato Fusarium wilt with Bacillus mycoides
作者: 湯佳蓉
Tang, Jia-Rong
關鍵字: 番茄鐮孢菌萎凋病
Tomato Fusarium wilt
誘導抗病
Bacillus mycoides
induced resistance
Bacillus mycoides
出版社: 植物病理學系所
引用: 丁姵分. 2006. 番茄萎凋病之生物防治菌的鑑定與防病潛力評估. 國立中興大學植物病理學系碩士論文. 51 pp. 台灣農家要覽. 1944. 農作物病害. 臺灣總督府農業試驗所. 1109 pp. 行政院農業委員會. 2012. 民國 99 年農業統計年報. 行政院農業委員會出版. 65 pp. 孫守恭、黃振文. 1996. 台灣鐮孢菌病害. 世維出版社. 170 pp。 黃振文、孫守恭. 1982. 台灣番茄萎凋病. 植保會刊 24:265-270。 黃振文、孫守恭. 2001. 蔬菜有機栽培非農藥病害防治專輯. 財團法人台北市瑠公產銷基金會編印. 台北市. 153 pp. 黃久菱. 2005. 萵苣萎凋病的生理小種鑑定與防治試驗. 國立中興大學植物病理學系碩士論文. 57 pp. 黃靜淑. 2008. Bacillus mycoides 防治甘藍幼苗病害之效果評估. 國立中興大學植物病理學系碩士論文. 58 pp. 黃涵、洪立. 1988. 台灣蔬菜彩色圖說. 國立台灣大學園藝系編印. 台北. 210 pp。 陳正次. 1995. 番茄. P.326-337. 台灣農家要覽:農作篇 (二). 農業委員會台灣農家要覽增修訂再版策劃委員會編. 財團法人豐年社出版. 698 pp. 陳泊菘. 2009. 產氣細菌防治柳橙綠黴病的效果與其抑菌氣體成分的分析. 國立中興大學植物病理學系碩士論文. 63 pp. 鄭安秀、王仕賢、黃山內. 2001. 番茄嫁接茄子根砧防治土傳病害. 台南區農業專訊 35:1-3 pp. Aime, S., Cordier, C., Alabouvette, C., and Olivain C. 2008. Comparative analysis of PR gene expression in tomato inoculated with virulent Fusarium oxysporum f. sp. lycopersici and the biocontrol strain F. oxysporum Fo47. Physilogical and Molecular Plant Pathology 73: 9-15. Alejandro, P. G., Diego, R. and Antonio, D. V. 2011. Plant protection and growth stimulation by microorganisms: biotechnological applications of Bacilli in agriculture. Biotechnology 22: 187-193. Bargabus, R. L., Zidack, N. K., Sherwood, J. W., and Tacobsen, B. J. 2002. Characterization of systemic resistance in sugar beet elicited by a non-pathogenic, phyllosphere-colonizing Bacillus mycoides, biological control agent. Physiol. Mol. Plant Pathol. 61: 289-298. Benhaumou, N., Joseph, W. K. and Tuzun, S. 1998. Induction of resistance against Fusarium wilt of tomato by combination of chitosan with an endophytic bacterial strain: ultrastructure and cytochemistry of the host response. Planta 204: 153-350. Choudhary, D. K., and Johri, B. N. 2009. Interactions of Bacillus spp. and plants-with special reference to induced systemic resistance (ISR). Microbiol. Res. 164(5): 493-513. Duijff B. J., Gianinazzi-pearson V., and Lemanceau P. 1997. Involvement of the outer membrane lipopolysaccharides in the endophytic colonization of tomato roots by biocontrol Pseudomonas fluorescens strain WCS417r. New Phytol. 135: 325-334. Francis, I., Holsters, M. 2010. The gram-positive side of plant-microbe interactions. Environ. Microbiol. 1: 1-12. Jacobsen, B. J., Zidack, N. K., and Larson, B. J. 2004. The role of Bacillus-based biological control agents in intergrated pest management system: Plant Disease. Phytopathology 94: 1272-1275. Jetiyanon, K., and Kloepper, J.W. 2002. Mixtures of plant growth-promoting rhizobacteria for induction of systemic resistance against multiple plant diseases. Biol. Control. 24: 285-291. John, A. R., Urs, H. N., Michael, G. W., Antonio, M., Henry-york, S. 1996. Systemic acquired resistance. The plant cell 8: 1809-1819. Kloepper, J. W., Ryu, C. M., and Zhang, S. 2004. The nature and application of biocontrol microbes: Bacillus spp. -induced systemic resistance and promotion of plant growth by Bacillus spp. Phytopathology 94: 1259-1266. Kumar, S., Chandra, A., and Pandey, K. C. 2008. Bacillus thuringiensis (Bt) transgenic crop: an environmentally friendly insect-pest management strategy. Environ. Biol. 29: 641-653. Kϋhn, B. N. and Brooks, D. M. 2002. Cross talk between signaling pathway in pathogen defense. Plant Biol. 5: 325-331. Liu, L., Kloepper, J. W., and Tuzun, S. 1995. Induction of systemic resistance in cucumber against Fusarium wilt by plant growth-promotin-rhizobacteria. Phytopathology 85: 695-698. Murashige, T., and Skoog, F. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiology 15: 473-497. Nash, S. M. and W. C. Snyder. 1962. Quantitative estimations by plate counts of propagules of the bean root rot Fusarium in field soils. Phytopathology 52: 567-572. Olivan, C., and Alabouvette, C. 1999. Process of tomato root colonization by a pathogenic strain of Fusarium oxysporum f. sp. lycopersici in comparison with a non-pathogenic strain. New Phytol. 141: 497-510. Pareja-Jaime, Y., Roncero, M. I. G.,and Ruiz-Roldan, M. C. 2008. Tomatinase from Fusarium oxysporum f. sp. lycopersici is required for full virulence on tomato plants. Molecular Plant-Microbe Interactions 21: 728-736. Perez-Garcia, A., Romero, D., and Vicente, A. 2011. Plant protection and growth stimulation by microorganisms: biotechnological applications of Bacilli in agriculture. Biotechnology 22: 187-193. Robert, P. L., and Deborah R. F. 1998. Efficacy of various fungal and bacterial biocontrol organisms for control of Fusarium wilt of tomato. Plant Dis. 82: 1022-1028. Samia, M. EL. 2007. Induction and modulation of resistance in tomato plants against Fusarium wilt disease by bioagent fungi (arbuscular mycorrihiza) and/or hormonal elicitors (jasmonic acid & salicylic acid): 2-Changes in the antioxidant enzymes, phenolic compounds and pathogen related-proteins. African Journal of Basic & Applied Sciences 1(4): 717-732. Van Wees, S. C. M., Luijendijk, M., Smoorenburg, I., van Loon, L. C., Pieterse, C. M. J., 2008. Rhizobacteria-mediated induced systemic resistance (ISR) in Arabidopsis is not associated with a direct effect on expression of known defence-inducible gene Atvsp upon challenge. Plant Molecular Biology 41: 537-549.
摘要: 在溫室利用 Bacillus mycoides CHT2402 及 NP02 細菌懸浮液分別拌入栽培介質後,培育番茄幼苗 14 天,再移植種於番茄萎凋病菌 (Fusarium oxysporum f.sp. lycopersici) 土中,結果發現兩菌株分別可降低植株罹病度 48.1% 及 46.2%。為了降低環境影響植株之抗病機制的表現,因此本研究於三角瓶中建立栽培番茄實生苗及接種生防菌及病原菌的技術,以了解生防菌誘導番茄植株抗病的相關機制。首先以 Biolog GP Microplate 分析番茄萎凋病菌 Fol-04 菌株及 B. mycoides CHT2402 和 NP02 兩菌株對碳、氮素源利用的需求,隨後比較生防菌及病原菌對營養需求的差異性,以便確立分析平台的基礎培養基配方,結果發現在 Murashige 和 Skoog 兩氏 (MS) 培養基中蔗糖的濃度調為 1% (w/v),最適合番茄植株、生防菌及病原菌等三者間的交互作用。在三角瓶與溫室栽培環境下,B. mycoides CHT2402 與 NP02 均可於處理 9 天後完全纏據番茄根部。將番茄種子於 CHT2402 及 NP02 菌液中催芽後培養於 MS 培養基二週,再以本病原菌單孢接種於番茄根部附近,5 天後發現對照組的植株已受病原菌感染且出現倒伏的現象,惟處理組植株尚維持挺立且未表現病徵。自三角瓶及溫室栽植處理及未處理過 B. mycoides CHT2402 及 NP02 菌株之番茄根部取樣後,以環氧樹脂包埋後切片,再以光學顯微鏡觀察它們的根組織之細胞結構,發現處理過 CHT2402 及 NP02 之植株根部於表皮細胞下方的細胞壁較對照組分別增厚 0.1-0.16 μm 及 0.12-0.17 μm,至於皮層細胞下方的細胞壁則是分別增厚 0.05-0.1 μm 及0.12-0.18 μm。利用穿透式電子顯微鏡觀察的結果,發現處理過 B. mycoides 番茄根部細胞大多於細胞間隙以非結晶物質累積於細胞壁,造成組織增厚的現象。在抗病基因表現方面,是於處理 B. mycoides 後第 9、11、13、15 天取樣後萃取其 mRNA 後,反轉錄成 cDNA 後以 qPCR 分析 PAL 及 LOX 基因的表現量,發現於三角瓶及溫室栽培環境中處理組植株的 LOX 與 PAL 基因均有被較高量誘導表現的現象。其中於三角瓶中處理 NP02 後第 11 天及處理 CHT2402 後第 13 天兩基因均有相同趨勢的高量表現;至於溫室中處理過 NP02 的番茄根部於處理後 11-13 天 PAL 基因有受誘導表現的現象且於處理後第 13-15 天 LOX 基因亦受誘導表現,而處理過 CHT2402的番茄根部則於處理後第 13 天僅 LOX 基因有受高量誘導表現的現象。綜合本研究結果証明 B. mycoides 須先於病原菌感染前纏據於番茄根部及維管束組織,並誘導番茄植株產生抗病反應,才能有效扺抗萎凋病菌的感染。
Bacillus mycoides CHT2402 and NP02 were effective in respectively reducing 48.1% and 46.2% disease severity of Fusarium wilt of tomato plants caused by Fusarium oxysporum f. sp. lycopersici Fol-04 in the greenhouse. In order to explore related mechanisms for inducing tomato plants resistant to Fusarium wilt disease by the biocontrol agent, a platform was set up for simultaneously culturing tomato seedlings, B. mycoides, and F. oxysporum f. sp. lycopersici Fol-04 in the flask cultivation system. Biolog GP Microplate was used to analyze the utilization of carbon and nitrogen sources by F. oxysporum f. sp. lycopersici Fol-04, B. mycoides CHT2402 and NP02. The results showed that sucrose concentration of Murashige’s and Skoog’s (MS) medium was adjusted to 1% (w/v) had more suitable interaction among tomato seedlings, the pathogen and biocontrol agents. The roots of tomato seedlings could be completely colonized by B. mycoides CHT2402 and NP02 nine days after the seeds were treated with cell suspension of the biocontrol agents and grown in the flask and greenhouse cultivation systems. Tomato seeds were incubated in the cell suspension (108 cfu/ ml) of B. mycoides CHT2402 and NP02 for 3 days, and then they were transplanted to the modified MS medium in the flask. Two weeks later, each of tomato seedlings was inoculated with single spore of F. oxysporum f. sp. lycopersici Fol-04 near the root. It was found that tomato seedlings could be protected from the pathogen by B. mycoides CHT2402 and NP02 for five days in the flask cultivation system. To study mechanisms for controlling tomato Fusarium wilt by the biocontrol agent, the root tissues of tomato plants treated with B. mycoides CHT2402 and NP02 were analyzed by tissue section and qPCR techniques. The results of Spurr’s resin block section indicated that the cell wall thickness of epidermis cells of tomato plant treated with B. mycoides CHT2402 and NP02 did increase 0.1-0.16 μm and 0.12-1.17 μm, and the cell wall thickness of cortex cells increased 0.05-0.1 μm and 0.12-0.18 μm. In addition, the results of qPCR showed that expression of LOX and PAL genes of tomato plant were induced by B. mycoides CHT2402 and NP02 in both cultivation conditions. According to above results, it was found that B. mycoides CHT2402 and NP02 were able to control tomato Fusarium wilt if they could colonize the roots and vascular tissues of tomato plants prior to the pathogen infection.
URI: http://hdl.handle.net/11455/31992
其他識別: U0005-0708201213385800
文章連結: http://www.airitilibrary.com/Publication/alDetailedMesh1?DocID=U0005-0708201213385800
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