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標題: Characteristics of Azotobacter species isolated from rice rhizosphere soils of organic and conventional farmings in Taiwan
作者: Meng-Ke Tsai
關鍵字: Azotobater;rice;free-living nitrogen fixation bacterium;固氮菌屬;水稻;有益菌;有機與慣行水稻田
引用: 趙震慶、蘇楠榮、王銀波。1996。有機農耕法之土壤肥力的變遷。中華農學會報 新 173:85–102。 張彥麗。2007。低磷土壤接種微生物壯秧劑對水稻秧苗吸收氮磷鉀的影響。東北農林大學環境與資源學院。 陳仁炫。 2010 。植物營養與土壤環境之相互效應。有機土壤改良技術研討會。國立高雄第一科技大學。 陳睦鈞、林宗俊、黃振文。2010。影響根圈細菌產生乙酸之因子。植病會刊 19: 201-212。 簡宣裕、林素禎、張明暉。 2011 。游離固氮菌 Azotobacter sp. 之施用對洋香瓜合理化施肥的效益。豐年社。 簡宣裕、張明暉、林素禎。 2012 。多功能非共生固氮細菌Azotobacter屬之介 紹與應用。健康農業。32:37-45 Ahmad, F., I. Ahmad and M. S. Khan. 'Screening of Free-Living Rhizospheric Bacteria for Their Multiple Plant Growth Promoting Activities.' Microbiol Res 163, no. 2 (2008): 173-81. Anjum, F.H., A. Tanveer, R. Ahmad, A. Ali, M.A. Nadeem, and M. Tahir. 2007. Response of cotton (Gossypium hirsutum) to split application of nitrogen and weed control methods. Indian Journal of Agricultural Sciences. 77:224-229. Aquilanti, L., F. Favilli, and F. Clementi. 2004. Comparison of different strategies for isolation and preliminary identification of Azotobacter from soil samples. Soil Biology & Biochemistry. 36:1475-1483. Aquilanti, L., I. Mannazzu, R. Papa, L. Cavalca, and F. Clementi. 2004. Amplified ribosomal DNA restriction analysis for the characterization of Azotobacteraceae: a contribution to the study of these free-living nitrogen-fixing bacteria. Journal of Microbiological Methods. 57:197-206. Arnow, L.E. (1937) Colorimetric determination of the components of 3,4-dihydroxyphenylalanine-tyrosine mixtures. J Biol Chem 118: 531–537. Awasthi, C.P., A. Kumar, N. Singh, and R. Thakur. 2011. Biochemical composition of grain amaranth genotypes of Himachal Pradesh. Indian Journal of Agricultural Biochemistry. 24:141-144. Bali, A., G. Blanco, S. Hill, and C. Kennedy. 1992. Excretion of ammonium by a nifl mutant of Azotobacter vinelandii fixing nitrogen. Applied and Environmental Microbiology. 58:1711-1718. Banerjee, A., S. Supakar and R. Banerjee. 'Melanin from the Nitrogen-Fixing Bacterium Azotobacter Chroococcum: A Spectroscopic Characterization.' PLoS One 9, no. 1 (2014): e84574. Becking, G.C. 1992. Methodology in neurotoxicology – activities within the world-health-organization and international program on chemical safety. Toxicology Letters. 64-5:203-208. Bhatia, R., S. Ruppel and N. Narula. 'Nifh-Based Studies on Azotobacterial Diversity in Cotton Soils of India.' Arch Microbiol 191, no. 11 (2009): 807-13. Bhattacharjee, R. B., A. Singh and S. N. Mukhopadhyay. 'Use of Nitrogen-Fixing Bacteria as Biofertiliser for Non-Legumes: Prospects and Challenges.' Appl Microbiol Biotechnol 80, no. 2 (2008): 199-209. Brick et al., 1990. J.M. Brick, R.M. Bostock, S.E. Silverstone. Rapid in situ assay for indoleacetic acid production by bacteria immobilized on nitrocellulose membrane. Appl. Environ. Microbiol., 57, pp. 535–538 Chauhan, Sapna, Kunal Wadhwa, Manjula Vasudeva and Neeru Narula. 'Potential Ofazotobacterspp. As Biocontrol Agents Againstrhizoctonia Solaniandfusarium Oxysporumin Cotton (Gossypium Hirsutum), Guar (Cyamopsis Tetragonoloba) and Tomato (Lycopersicum Esculentum).' Archives of Agronomy and Soil Science 58, no. 12 (2012): 1365-1385. Chennappa, G., C. R. Adkar-Purushothama, M. K. Naik, U. Suraj and M. Y. Sreenivasa. 'Impact of Pesticides on Pgpr Activity of Azotobacter Sp. Isolated from Pesticide Flooded Paddy Soils.' Greener Journal of Agricultural Sciences 4, no. 4 (2014): 117-129. Choudhury, A., and I.R. Kennedy. 2004. Prospects and potentials for systems of biological nitrogen fixation in sustainable rice production. Biology and Fertility of Soils. 39:219-227. Gaind, S., and A.C. Gaur. 1991. Thermotolerant phosphate solubilizing microorganisms and their interaction with mung bean. Plant and Soil. 133:141-149. Gauri, S. S., S. M. Mandal and B. R. Pati. 'Impact of Azotobacter Exopolysaccharides on Sustainable Agriculture.' Appl Microbiol Biotechnol 95, no. 2 (2012): 331-8. Gravel, Valerie, Hani Antoun and Russell J. Tweddell. 'Growth Stimulation and Fruit Yield Improvement of Greenhouse Tomato Plants by Inoculation with Pseudomonas Putida or Trichoderma Atroviride: Possible Role of Indole Acetic Acid (Iaa).' Soil Biology and Biochemistry 39, no. 8 (2007): 1968-1977. Gupta, A., J.M. Meyer, and R. Goel. 2002. Development of heavy metal-resistant mutants of phosphate solubilizing Pseudomonas sp NBRI 4014 and their characterization. Current Microbiology. 45:323-327. He, Y., K. Xie, P. Xu, X. Huang, W. Gu, F. Zhang and S. Tang. 'Evolution of Microbial Community Diversity and Enzymatic Activity During Composting.' Res Microbiol 164, no. 2 (2013): 189-98. Javier Jimenez, D., J. Salvador Montana, and M. Mercedes Martinez. 2011. Characterization of free nitrogen fixing bacteria of the genus Azotobacter in organic vegetable-grown Colombian soils. Brazilian Journal of Microbiology. 42:846-858. Jones, D. L., T. Eldhuset, H. A. de Wit, and B. Swensen (2001) Aluminium effects on organic acid mineralization in a Norway spruce forest soil. Soil Biol. Biochem. 33: 1259-1267. Kannan T, Ponmurugan P (2010) Response of paddy (Oryza sativa L.) varieties to Azospirillum brasilense inoculation. J Phytol 2:8–13 Kennedy, C., and D. Dean. 1992. The nifU, nifS and nifV gene-products are required for activity of all 3 nitrogenases of Azotobacter vineladii. Molecular & General Genetics. 231:494-498. Khush, G.S. 1997. Origin, dispersal, cultivation and variation of rice. Plant Molecular Biology. 35:25-34. Kim, Y., K. Mikawa, T. Saito, K. Tanaka, and H. Emori. 1997. Development of novel anaerobic/aerobic filter process for nitrogen removal using immobilized nitrifier pellets. Water Science and Technology. 36:151-158. Kumar, Vivek, Rishi Kumar Behl and Neeru Narula. 'Establishment of Phosphate-Solubilizing Strains of Azotobacter Chroococcum in the Rhizosphere and Their Effect on Wheat Cultivars under Green House Conditions.' Microbiological Research 156, no. 1 (2001): 87-93. Kumar, P., I. Mehrotra, and T. Viraraghavan. 1998. Temperature response of biological phosphorus-removing activated sludge. Journal of Environmental Engineering-Asce. 124:192-196. Lee, S.E., K.S. Kim, J.H. Ahn, and C.W. Kim. 1997. Comparison of phosphorus removal characteristics between various biological nutrient removal processes. Water Science and Technology. 36:61-68. Lo, W., P. Zhang, C.-C. Ling, S. Huang, and R.H. Holm. 2012. Formation, Spectroscopic Characterization, and Solution Stability of an Fe4S4 (2+) Cluster Derived frorn beta-Cyclodextrin Dithiolate. Inorganic Chemistry. 51:9883-9892. Melody SC (1997) Plant molecular biology—a laboratory manual. Springer, New York Miranda, Laura, Khalid Boulahya, Aurea Varela, Jose M. Gonzalez-Calbet, Marina Parras, Maria Hernando, M. Teresa Fernandez-Diaz, Antonio Feteira and Derek C. Sinclair. 'Structure-Property Relationships of the 10h Hexagonal-Type Perovskite Bamn(0.4)Fe(0.6)O(2.73).' Chemistry of Materials 19, no. 14 (2007): 3425-3432. Nag, P. and S. Pal. 'Fe Protein over-Expression Can Enhance the Nitrogenase Activity of Azotobacter Vinelandii.' J Basic Microbiol 53, no. 2 (2013): 156-62. Neilands, J. B. (1981). 'IRON-ABSORPTION AND TRANSPORT IN MICROORGANISMS.' Annual Review of Nutrition 1: 27-46. Patil, A.A., and K.M. Bojappa. 1984. Effects of cultivars and graded levels of nitrogen and phosphorus on certain quality attributes of tomato (Lycopersicon esculentum Mill). I. TSS, acidity, ascorbic acid and puffiness. Mysore Journal of Agricultural Sciences. 18:35-38. Paul, S., B. Paul, M. Aslam Khan, C. Aggarwal, J. K. Thakur and M. S. Rathi. 'Effects of Lindane on Lindane-Degrading Azotobacter Chroococcum; Evaluation of Toxicity of Possible Degradation Product(S) on Plant and Insect.' Bull Environ Contam Toxicol 90, no. 3 (2013): 351-6. Rai, U. N., K. Pandey, S. Sinha, A. Singh, R. Saxena and D. K. Gupta. 'Revegetating Fly Ash Landfills with Prosopis Juliflora L.: Impact of Different Amendments and Rhizobium Inoculation.' Environment International 30, no. 3 (2004): 293-300. Sahoo, R. K., M. W. Ansari, T. K. Dangar, S. Mohanty and N. Tuteja. 'Phenotypic and Molecular Characterisation of Efficient Nitrogen-Fixing Azotobacter Strains from Rice Fields for Crop Improvement.' Protoplasma 251, no. 3 (2014): 511-23. Shavrukov, Y., P. Langridge, M. Tester, and E. Nevo. 2010. Wide genetic diversity of salinity tolerance, sodium exclusion and growth in wild emmer wheat, Triticum dicoccoides. Breeding Science. 60:426-435. Shenker, M., I. Oliver, M. Helmann, Y. Hadar and Y. Chen. 'Utilization by Tomatoes of Iron Mediated by a Siderophore Produced by Rhizopus-Arrhizus.' Journal of Plant Nutrition 15, no. 10 (1992): 2173-2182. Shieh, W.Y., U. Simidu, and Y. Maruyama. 1989. Enumeration and characterization of nitrogen-fixing bacteria in an eelgrass(zostera-marina). Microbial Ecology. 18:249-259. Shokri, D. and G. Emtiazi. 'Indole-3-Acetic Acid (Iaa) Production in Symbiotic and Non-Symbiotic Nitrogen-Fixing Bacteria and Its Optimization by Taguchi Design.' Curr Microbiol 61, no. 3 (2010): 217-25. Smedt, P.d., P.A.P. Liddle, and G. Cavazza. 1980. Analysis of different types of rum. Annales des Falsifications et de l'Expertise Chimique. 73:385-397. Takahashi, H., M. Rai, T. Kitagawa, S. Morita, T. Masumura, and K. Tanaka. 2004. Differential localization of tonoplast intrinsic proteins on the membrane of protein body type II and aleurone grain in rice seeds. Bioscience Biotechnology and Biochemistry. 68:1728-1736. Tester, Mark and Peter Langridge. 'Breeding Technologies to Increase Crop Production in a Changing World.' Science 327, no. 5967 (2010): 818-822. Tian, Wei, Qian Sun, Dabing Xu, Zhenhua Zhang, Da Chen, Chunyu Li, Qirong Shen and Biao Shen. 'Succession of Bacterial Communities During Composting Process as Detected by 16s Rrna Clone Libraries Analysis.' International Biodeterioration & Biodegradation 78, (2013): 58-66. Tung, S. K., L. J. Teng, M. Vaneechoutte, H. M. Chen and T. C. Chang. 'Identification of Species of Abiotrophia, Enterococcus, Granulicatella and Streptococcus by Sequence Analysis of the Ribosomal 16s-23s Intergenic Spacer Region.' J Med Microbiol 56, no. Pt 4 (2007): 504-13. Yanni, Y.G., and F.K. Abd El-Fattah. 1999. Towards integrated biofertilization management with free living and associative dinitrogen fixers for enhancing rice performance in the Nile delta. Symbiosis. 27:319-331. You, M., T. Nishiguchi, A. Saito, T. Isawa, H. Mitsui and K. Minamisawa. 'Expression of the Nifh Gene of a Herbaspirillum Endophyte in Wild Rice Species: Daily Rhythm During the Light-Dark Cycle.' Appl Environ Microbiol 71, no. 12 (2005): 8183-90. Zhang, X., H. Zhao, J. Zhang and Z. Li. 'Growth of Azotobacter Vinelandii in a Solid-State Fermentation of Technical Lignin.' Bioresour Technol 95, no. 1 (2004): 31-3.
Azotobacter, a free-living nitrogen fixation bacterium, has been proved in promoting rice growth through releasing some beneficial compounds. However, the function of Azotobacter spp. are highly specific, and rarely studies focus on the comparisons of their status in Taiwan organic and conventional paddy rice fields. The purposes of this study were: 1. to screen and classify with 16S rDNA-based of the Azotobacter isolates from 8 organic and 8 conventional paddy rice fields distribution around different Taiwan area. 2. to determine and compare the characters of the isolates between organic and conventional fields. 3. to select the highly potential isolates and evaluate their apparent beneficial function on the growth of rice seedlings. Results showed that there were 40 Azotobacter isolates identified, based on 16S rDNA sequence analysis.There are five Azotobacter spp. been found, including A. armeniacus, A. beijerinckii, A. chroococcum, A. tropicalis and A. vinelandii. There were 12.5% isolates from conventional farms, and 87.5% from organic farms. In acid soils A. beijerinckii (53.3%, 43.8%) is the most popular and A chroococcum (40.8%, 43.8%) is the second at soil pH 4.5 - 5.5 and pH 5.5 - 6.5, respectively. As the soil pH increased, the percentage of A. Beijerinckii isolates decreased. In soil pH ranged 6.5 - 7.5 the most popular species are A. chroococcum (33.3%), A. vinelandii (33.3%) and followed by A. beijerinckii (22.2%). The ability of Pyoverdine production, tricalcium phosphate dissolution and gelatin hydrolysis of Azotobacter isolates which screened from organic farms is more than from conventional farms. In this study, a total of 14 Azotobacter isolates produce Pyoverdine after subculture, 16 isolates can use the carboxymethyl cellulose as carcon source, the colonies of 17 isolates will turn into brown after 3 days, and 5 isolates (CHB202, CHB214, CHB251, CHB257 and CHB278) have the ability to use lignin. In addition, CHB194, CHB213, CHB216 and CHB251 have been identified as the nif H genes owner, but their ethylene reduction ability is too low to be considered as nitrogen fixer. All isolates could grow at tricalcium phosphate medium, but there are more than half of the isolates (22/40) have no transparent ring around the colonies. There were 5 potential Azotobacter isolates, CHB196, CHB198, CHB188, CHB197, CHB260 have been evaluated as potential beneficial microorganisms by thier performances on nitrogen fixation, IAA production and tricalcium phosphate solubilization. In bioassay of these five isolates on the growth of rice seedlings, results showed that all of the inoculants increased the growth of rice seedling shoot and root. Furthermore, the promotion effect of CHB188, CHB196, CHB197 isolates reached significant level compared to the control. The present work provided the information that the Azotobacter species screened from Taiwan paddy rice fileds have potential in increasing the production of rice.

接種游離固氮菌屬Azotobacter被證實可增加土壤中氮含量及促進水稻生長。然而Azotobacter spp.的功能依據地區而有高度特異性且台灣水稻田之栽培農法與土壤性質對分離之 Azotobacter 菌種的影響研究較少。本研究的目的可分為1.分離與以16S rDNA 序列鑑定自台灣本土水稻田中分離之Azotobacter spp.。2.探討台灣本土有機與慣行農法水稻根圈土壤中Azotobacter 分離菌株種類及其生理、生化特性與土壤性質的相關性。3.評估Azotobacter 分離株對於水稻苗生長之影響。
本研究從8處有機耕種及8處傳統耕種水田土壤樣品中分離共40株Azotobacter菌株,依據分離株之16S rDNA 序列分析結果,所分離菌株屬A. armeniacus (亞美尼亞短桿固氮細菌)、A. beijerinckii (倍傑林客短桿固氮細菌)、A. chroococcum (色球短桿固氮細菌)、A. tropicalis (熱帶短桿固氮細菌) 和A. vinelandii (葡萄園短桿固氮細菌)等5種。分離之菌株12.5% 來自慣行田區、87.5% 來自有機田區。土壤pH 4.5 - 5.5及pH 5.5 - 6.5兩範圍中,Azotobacter分離株菌種組成以A. beijerinckii為主(53.3%及43.8%),A. chroococcum(40.8%及43.8%)次之。隨著土壤pH值上升,A. beijerinckii之比例下降。在土壤樣品pH 6.5 - 7.5主要菌種組成以A. chroococcum (33.3%)及A. vinelandii (33.3%),其次才是A. beijerinckii (22.2%)。另慣行農法田區之Azotobacter分離株生成載鐵物質- Pyoverdine的比例(56%) 高於有機農法田區 (32%),在載鐵物質、磷酸三鈣溶解能力及水解明膠的能力測試也有同樣的趨勢。
本研究所篩選之 Azotobacter 有14菌株在繼代培養後仍能生成Pyoverdine並於235nm (UVC)波長下產生生物性螢光。有17株菌在經過繼代培養三天後會轉成褐色。CHB202、CHB214、CHB251、CHB257及CHB278等5分離株有分解木質素之透明環。有16分離株具分解羧甲基纖維素後形成之透明圈。另外,CHB194、CHB213、CHB216 及CHB251等4菌株具有nif H固氮基因,在繼代培養後所測試固氮活性數值太低無法視為具有游離固氮活性。所有分離株均能在磷酸三鈣培養基上生長良好,然而卻有半數以上分離株 (22/40) 無法在菌落周圍形成透明環。所篩選Azotobacter分離株所合成IAA濃度範圍0.49~15.59 μg/ml。綜合評估游離固氮活性、溶解磷酸三鈣能力及IAA生成量等有益特性,有明確表現者由高到低為CHB196、CHB198、CHB188、CHB197、CHB260。這5菌株可提高稻穀發芽率及水稻苗地上部及根部生長且顯著高於對照組。從本研究之結果證明可以由本土土壤篩選有利水稻栽培的Azotobacter菌株。
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