微生物學(xué)通報(bào) APR 20, 2008, 35(4): 512~518
Microbiology 2008 by Institute of Microbiology, CAS
曾慶桂 朱 篤* 顏日明 朱 飛 王志興
(江西師范大學(xué)生命科學(xué)學(xué)院 江西省亞熱帶植物資源保護(hù)與利用重點(diǎn)實(shí)驗(yàn)室 南昌 330022)
摘 要: 從蛇足石杉(Huperzia serrata)葉片中分離到一株對(duì)小麥赤霉(Fusarium graminearum)等多種植物病原真菌有強(qiáng)拮抗作用的內(nèi)生細(xì)菌H-6。通過(guò)形態(tài)和培養(yǎng)特性觀察,、生理生化實(shí)驗(yàn)及16S rDNA序列分析, 初步鑒定該菌屬于伯克霍爾德屬, 命名為Burkholderia sp. H-6,。同時(shí)還對(duì)該菌培養(yǎng)條件進(jìn)行了優(yōu)化, 得出馬鈴薯浸出液中添加2.5%的甘露醇和0.1% NaNO3有利于細(xì)胞生長(zhǎng)和抑菌活性的產(chǎn)生, 最適培養(yǎng)溫度為28℃, 最佳初始pH4.0。
關(guān)鍵詞: 蛇足石杉, 內(nèi)生細(xì)菌, 拮抗真菌活性, 鑒定
Isolation and Identification of Endophytic Bacteria H-6 with High Antifungal Activity from Huperzia serrata
ZENG Qing-Gui ZHU Du* YAN Ri-Ming ZHU Fei WANG Zhi-Xing
(College of Life Sciences, Jiangxi Normal University, Jiangxi Provincial Key Lab of Protection and Utilization
of Subtropic Plant Resources, Nanchang 330022)
Abstract: In this study, nearly 200 endophytic bacteria were isolated from different part of Huperzia serrata, over 60 bacterium with clear antifungal activity were selected from those cultures. Among them, strain H-6 exhibited the highest antifungal activity which was strongly inhibits the growth of many plant pathogenic fungi such as Sclerotinia scleroliorum, Fusarium graminearumt, Sclerotinia libertiana, Phytophthora capsici Leonia and Sesame fusarium wilt. According to the characteristics of morphology, physiology and biochem-istry tests and the comparison of 16S rDNA sequence, the strain H-6 was similar to the Burkholderia. So strain H-6 was identified as Burkholderia sp. H-6. The results also showed that Burkholderia sp. H-6 was markedly different from Burkholderia cepacia that was applied widely in agriculture as antagonistic bacte-ria.
The medium and culture conditions of the strain all were optimized by single factor and orthogonal ex-periments. In the investigation of the culture condition, growth was carried out in a basal medium (potato juice) and gradually supplemented with the various ingredients to be investigated. The major ingredients be-ing investigated included carbon sources and nitrogen sources. The optimal antifungal activity production condition is growth in a medium (potato juice with 2.5% mannitol and 0.1% NaNO3), initial pH 4.0 at 28℃.
Keywords: Huperzia serrata, Endophytic bacterium, Antifungal activity, Identification
植物病害80%以上是由真菌引起的, 控制真菌病害對(duì)于提高農(nóng)作物產(chǎn)量極其重要,。目前真菌防治主要以化學(xué)農(nóng)藥為主, 而化學(xué)農(nóng)藥引起的環(huán)境污染已經(jīng)對(duì)人類健康造成潛在的威脅, 因而從中尋找可用作生物農(nóng)藥或者生物肥料的菌種成為當(dāng)前的研究熱點(diǎn)[1],。
植物內(nèi)生菌是一個(gè)多樣性十分豐富的生物類群。在植物內(nèi)生菌中, 有對(duì)植物病原菌產(chǎn)生拮抗而保護(hù)植物免受病原菌侵害的衛(wèi)士, 有對(duì)宿主體內(nèi)某些活性成分的形成有重要影響者, 也有能產(chǎn)生與宿主相同或相似的生理活性成分者, 還有與宿主只是共生或伴生關(guān)系而作用不明者等等[2],。迄今, 從植物中分離可用作生物農(nóng)藥菌種已有較多的研究, 已從裸子植物和被子植物中分離到較多能產(chǎn)拮抗病害真菌或細(xì)菌次級(jí)代謝產(chǎn)物的內(nèi)生菌株[3-6],。但相對(duì)而言, 蕨類植物中的內(nèi)生菌資源研究報(bào)道較少。
蛇足石杉( Huperzia serrata (Thunb.) Trev.) 是一種珍貴的藥用蕨類植物, 屬石杉科石杉屬[7],。迄今有關(guān)蛇足石杉資源,、藥用成分等方面已有較多報(bào)道, 但有關(guān)蛇足石杉內(nèi)生菌資源的研究方面, 僅見(jiàn)2005年石瑋等從蛇足石杉分離出4株內(nèi)生真菌的報(bào)道[8], 且尚未有對(duì)其價(jià)值進(jìn)一步研究的報(bào)道。本實(shí)驗(yàn)室從蛇足石杉中分離到了近200株內(nèi)生細(xì)菌,、150株內(nèi)生真菌,。在內(nèi)生菌分離的過(guò)程中發(fā)現(xiàn), 在近三分之一的經(jīng)過(guò)表面消毒的蛇足石杉碎片中都長(zhǎng)出了周圍有一黃綠色色素圈的細(xì)菌, 而且在分離平板上對(duì)長(zhǎng)出的內(nèi)生真菌有較強(qiáng)拮抗作用, 命名為H-6。對(duì)其進(jìn)行了病原真菌的拮抗對(duì)峙實(shí)驗(yàn), 發(fā)現(xiàn)它對(duì)小麥赤霉(Fusarium graminearum)等多種植物病原菌都有強(qiáng)拮抗作用,。為此筆者對(duì)其進(jìn)行了形態(tài)及生理生化特征分析以及16S rDNA分子鑒定, 并對(duì)其培養(yǎng)條件進(jìn)行了初步優(yōu)化, 為進(jìn)一步研究H-6和將其應(yīng)用于生物防治提供依據(jù),。
全文下載：一株拮抗真菌的蛇足石杉內(nèi)生細(xì)菌分離鑒定及培養(yǎng)條件優(yōu)化
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