台灣高鹽土壤中植物生長促進根系細菌 (PGPR) 的性狀分析

Characterization of Plant Growth Promoting Rhizobacteria (PGPR) from Saline Soil in Taiwan

陸布迪(Satria Prima Budi Utama)

指導教授 ： 張珮君

國立屏東科技大學/農學院/生物科技系/碩士(2019年)

https://doi.org/10.6346/NPUST202000022

摘要

農業部門目前面臨無數問題，例如：非生物脅迫導致的產量損失以及植物病原體的感染。有許多證據表明，使用促進植物生長的根際細菌（PGPR）將克服這些問題，進而改善植物的生長和作物的產量。在這項研究中，從植物的根部表面和內部組織中分離出細菌菌株，這些菌株的生長基於產生氨環丙烷-1-羧酸鹽（ACC）、吲哚乙酸（IAA）、鐵載體、磷酸鹽增溶、幾丁質酶以及抗真菌能力，這些能力被認為是促進植物生長的最可靠特徵。在本實驗中分離了32個菌株，其中包括16個表面分離和16個內生菌。分離出的30種細菌顯示出能夠產生ACC脫氨酶的能力，其值範圍在1.8 – 29.6 µmol / mg左右。分離的三烯產生的IAA含量為0.003 – 67.97 µg / ml。發現三十種分離物分泌鐵載體，導致7天后在1.5 mm-15 mm處形成暈圈。此外，通過增溶指數（SI）評估了溶解非生物利用的磷酸鹽的能力，SI為1.09至3.03mm。幾丁質酶測試顯示13株具有降解幾丁質能力的菌株，測試了所有菌株對灰葡萄孢（1.481-48.14％），炭疽菌（0.4）-46.25％和菜粉蝨（0.54-35.15％）的抗真菌活性。每個PGPR性狀的優良分離株分別為SLE1（肺炎克雷伯菌），SIE1（肺炎克雷伯菌），SGS3（陰溝腸桿菌），SJS3（嗜鹽氣單胞菌），SHS1（產氣克雷伯氏菌）,SCS1(金黃色葡萄球菌), 和SJS1（將粘質沙雷氏菌) 和基於16s RNA。鑑定最高的抗真菌和幾丁質酶測試結果，將粘質沙雷氏菌(Serratia marcescens）用50％硫酸銨進行培養和沈淀，然後通過CHT（陶瓷Hidroxyapatite）柱進行純化。該抗真菌蛋白顯示出一條帶，分子量約為50 kDa，該蛋白是通過LC-MS獲得的。通過使用MASCOT搜索引擎匹配數據庫，將抗真菌蛋白鑑定為假設蛋白。這些結果表明，在鹽脅迫條件的植物栽培中下，控制真菌病原體並改善植物生長是有希望的。

關鍵字

PGPR ； ACC脫氨酶 ； IAA ； 鐵載體 ； 磷酸鹽溶解 ； 抗真菌活性

並列摘要

The agricultural sector is continuously facing myriad of problems such as yield loss due to abiotic stress as well as phytopathogen infestation. There is well established evidence that the use of plant growth-promoting rhizobacteria (PGPR) would overcome those problems and in turn improve plant growth and crop yield. In this research bacteria strains were isolated from root surface and inner tissue of plants which grown in highly saline soils, based on their ability to produce aminocyclopropane-1-carboxylate (ACC), indole acetic acid (IAA), siderophore, phosphate solubilization, chitinase as well as their antifungal ability. Those abilities are considered the most reliable traits for promoting plant growth. A total of 32 strains were isolated and examined, with 16 surface isolates and 16 endophytes. Thirty bacteria isolates showed capability to produce ACC deaminase with a range of values around 1.8 – 29.6 µmol/mg. Thirtyone isolates produced IAA at levels of 0.003 – 67.97 µg/ml. Thirty isolates were found to secrete siderophore that results in halo zones around 1.5 mm - 15 mm after 7 days. Moreover, the ability to solubilize non- bioavailable phosphate was assessed by solubilization index (SI), with SI being 1.09 - 3.03 mm. Chitinase tests showed 13 isolates having the ability to degrade chitin, all isolates were tested the antifungal activity against Botrytis cinerea (1.481- 48.14 %), Colletotrichum gloeosporioides (0.44- 46.25%) and Pestaliotiopsis sp (0.54-35.15%). The superior isolates of each PGPR traits were identified as SLE1 (Klebsiella pneumoniae), SIE1 (Klebsiella pneumoniae),SGS3 (Enterobacter cloacae),SJS3 (Stenotrophomonas rhizophila), SHS1 (Klebsiella aerogenes), SCS1(Staphylococcus sp) and SJS1(Serratia marcescens) based on 16s RNA. The highest antifungal and chitinase test was identified the antifungal compounds, Serratia marcescens protein were precipitation by 50% ammonium sulfate and purified by CHT (Ceramic Hidroxyapatite) column. The antifungal protein showed band with molecular mass around 50 kDa, the protein was obtained by using LC- MS. The antifungal protein has been identified as hypothetical protein by matching databases using MASCOT search engine. These results suggest that it is promising to control the fungal pathogens and to improve plant growth under stress salt condition in plant cultivation.

並列關鍵字

PGPR ； ACC deaminase ； IAA ； Siderophore ； Phosphate solubilization ； Antifungal activity

參考文獻

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