نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشگاه بوعلی سینا همدان

2 دانشگاه بوعای سینا همدان

3 دانشیار گروه علوم خاک دانشگاه تبریز

4 دانشگاه تبریز

چکیده

ازتوباکتر از باکتری‌های آزادزی تثبیت‌کننده نیتروژن در خاک بوده که از راه‌های گوناگونی مایه افزایش رشد گیاه می‌شود. فراوانی و فعالیّت آن‌ها در خاک وابسته به ویژگی‌های خاک و کاربری آن است. بر این پایه برای جداسازی و شناسایی برخی از ازتوباکترهای افزآینده رشد گیاه، 50 نمونه خاک به گونه بختانه ‌از کاربری‌های گوناگون در سه استان آذربایجان شرقی، اردبیل و گیلان از لایه صفر تا 25 سانتی‌متری خاک نمونه‌برداری شد. برای جداسازی ازتوباکترها از روش خمیره خاک بهره‌گیری شد و ناب‌سازی آن‌ها با استفاده از کشتگاه LG و Nutrient Agar انجام گرفت. در این بررسی در آغاز 50 جدایه جداسازی شد که بر پایه ویژگی‌های فنوتیپی و ریخت‌شناسی جدایه‌ها و چگونگی رشد آن‌ها در کشتگاه‌های یادشده، جدایه‌های همانند کنار گذاشته شده و شناسایی نه جدایه به نام‌های 2SP-5، 14SPI، 14SP2-1، 16SP-2، 34SPIII، 35SP، 35SP-2،43SP-2 و 44SP-2 همراه با آزمون توانایی افزایندگی رشد آنها انجام شد. نتایج شناسایی مولکولی (16S rDNA) نشان داد که از میان نه جدایه، پنج جدایه 14SPI، 14SP2-1، 16SP-2، 35SP و 44SP-2 ازتوباکتر کروکوکوم بوده و جدایه 34SPIII و 35SP-2 سودوموناس و جدایه 2SP-5 و 43SP-2 بیجرینکیا شناخته شدند. یافته‌های بررسی ویژگی‌های افزایندگی رشد گیاه نیز نشان داد که بیش‌ترین تثبیت نیتروژن در ازتوباکتر کروکوکوم جدایه‌ 14SP2-1 به‌دست آمد و بالاترین توانایی حل کنندگی فسفر و ساخت اکسین در باکتری سودوموناس جدایه 34SPIII دیده شد. از دیدگاه آزادسازی پتاسیم نیز باکتری‌های ازتوباکتر کروکوکوم جدایه‌ 44SP-2 و 14SP2-1 توانایی بالاتری نشان دادند. ازتوباکترهای به‌دست آمده همگی متعلق به گونه ازتوباکتر کروکوکوم بودند که از نمونه خاک‌های دو استان آذربایجان شرقی و گیلان با کاربری پوشش مرتعی و شالیزار به دست آمدند.

کلیدواژه‌ها

عنوان مقاله [English]

Isolation, identification, and determination of plant growth promoting properties of Azotobacteria isolated from soil samples North-west of Iran under different land-use

چکیده [English]

Azotobacter is free-living nitrogen-fixing bacteria that it leading to promoting plants growth through various ways. Frequency and distribution of Azotobacter is affected by soil use and its properties. Accordingly, this study aimed to isolate and identify some of plant growth promoting (PGP) Azotobacters. To this end, 50 soil samples were taken randomly from depth 0-25-cm under different land-uses in three provinces viz. Eastern-Azerbaijan, Ardabil and Gilan. Soil paste method was used for isolation of these bacteria and purification was done in LG and NA media. We first isolated 50 bacterial isolates based on the phenotypic and morphological properties. And then we selected nine isolates of them for detailed and deep studies (including 2SP-5, 14SPI, 14SP2-1, 16SP-2, 34SPIII, 35SP, 35SP-2, 43SP-2 and 44SP-2). Results of molecular identification of bacteria (16S rDNA) revealed that among nine isolates, the five isolates (viz. 14SPI, 14SP2-1, 16SP-2, 35SP and 44SP-2) belonged to Azotobacter chroococcum, and isolates 34SPIII and 35SP-2 belonged to Pseudomonas sp. while 2SP-5 and 43SP-2 were identified as Beijerinckia sp. We also found the highest N fixation, and P solubility and auxin production were recorded by A. chroococcum 14SP2-1 and Pseudomonas sp. 34SPIII, respectively. Maximum potassium releasing was observed by A. chroococcum 14SP2-1 and 44SP-2. In this study, all the identified Azotobacteria belonged to A. chroococcum which isolated from the pasture, corn and rice lands of East Azerbaijan and Gilan provinces.

کلیدواژه‌ها [English]

  • Molecular identification of bacteria
  • N-fixing bacteria
  • Soil paste
  • Plant growth promoting properties
Reference
Ahmad F., Ahmad I. and Khan M. S. 2008. Screening of free-living rhizospheric bacteria for their multiple plant growth promoting activities. Microbiological research, 163(2): 173-181.
Altschul SF., Madden T.L., Schaffer A.A., Zhang J., Zhang Z., Miller W. and Lipman D.J. 1997. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Research. 25: 389-402.
Aquilantia L., Favillib F. and Clementia F. 2004. Comparison of different strategies for isolation and preliminary identification of Azotobacter from soil samples. Soil Biology & Biochemistry, 36: 1475-1483.
Behrooz A., Olamaee M., Movahedi Naeeni S.A.R. and Ghorbani Nasrabadi R. 2016. Evaluation of plant growth promotion chracteristics  of native soil Azotobacter isolates from Golestan province. Juranl of Soil Management and Sustainable, 6(1): 233-246. (In Persian).
Brahmaprakash G. P. and Sahu P. K. 2012. Biofertilizers for sustainability. Journal of the Indian Institute of Science, 92(1): 37-62.
Cotteni A. 1980. Methods of plant analysis. In: Soil and Plant Testing FAO Soils Bulletin. 38/2, pp. 64-100.
Dastager S. G., Deepa C. K. and Pandey A. 2010. Isolation and characterization of novel plant growth promoting Micrococcus sp NII-0909 and its interaction with cowpea. Plant Physiology and Biochemistry, 48(12): 987-992.
Deaker R., László Kecskés M. Timothy Rose M. Amprayn K. Krishnen G. Thi Kim Cuc T. Thuy Nga V. Thi Cong P. Thanh Hien N. and Robert Kennedy I. 2011. Practical methods for the quality control of inoculant biofertilizers. ACIAR.
Diep C. N. and Hieu T. N. 2013. Phosphate and potassium solubilizing bacteria from weathered materials of denatured rock mountain, Ha Tien, Kiên Giang province Vietnam. American Journal of Life Sciences, 1(3): 88-92.
Fageria N. K., Baligar V. C. and Li Y. C. 2009. Differential soil acidity tolerance of tropical legume cover crops. Communications in Soil Science and Plant Analysis, 40(7-8): 1148-1160.
Farajzadeh D., Yakhchali B., Aliasgharzad N., Sokhandan-Bashir N. and Farajzadeh M. 2012. Plant growth promoting characterization of indigenous Azotobacteria isolated from soils in Iran. Current microbiology, 64(4): 397-403.
Gee G.W. and Bauder J.W. 1986. Particle size analysis. Methods of Soil Analysis. Part1, Physical and
mineralogical methods. American Society Agronomy Publication. 383-411.
Giongo A., Beneduzi A., Gano K., Vargas L. K., Utz L. and Passaglia L. M. P. 2013. Characterization of plant growth-promoting bacteria inhabiting Vriesea gigantea Gaud. and Tillandsia aeranthos (Loiseleur) LB Smith (Bromeliaceae). Biota Neotropica, 13(3): 80-85.
Gravel V., Antoun H. and Tweddell R. J. 2007. 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(8): 1968-1977.
Han H. S. and Lee K. D. 2005. Phosphate and potassium solubilizing bacteria effect on mineral uptake, soil availability and growth of eggplant. Research journal of agriculture and biological sciences, 1: 176-180.
Harris J.O. 1973. Azotobacter of the Konza Prairie. In L.C. Hulbert (ed): Third Midwest Prairie Conference Proceedings. Manhattan, KS: Division of Biology, Kansas State University, pp. 53-54.
Hu X. and Boyer G. L. 1996. Siderophore-mediated aluminum uptake by Bacillus megaterium ATCC 19213. Applied and Environmental Microbiology, 62(11): 4044-4048.
Jasim B., Joseph A. A., John C. J., Mathew J. and Radhakrishnan E. K. 2014. Isolation and characterization of plant growth promoting endophytic bacteria from the rhizome of Zingiber officinale. 3 Biotechnology, 4(2): 197-204.
Jiménez D. J., Montaña J. S. and Martínez M. M. 2011. Characterization of free nitrogen-fixing bacteria of the genus Azotobacter in organic vegetable-grown Colombian soils. Brazilian Journal of Microbiology, 42(3): 846-858.
Kanimozhi K. and Panneerselvam A. 2010. Studies on isolation and nitrogen fixation ability of Azospirillum spp. isolated from Thanjavur district. Der Chemica Sinica, 1(3):138-145.
Kennedy C., Rudnick P., MacDonald M. and Melton T. 2005. “Genus III:Azotobacter,”in Bergey’s Manual of Systematic Bacteriology. The Proteobacteria, Part B, the Gammaproteobacteria, G. M. Garrity, Ed.,vol.2,pp.384–402, Springer, NewYork, NY, USA, 2ndedition.
Khan A.A., Jilani G., Akhtar M.S., SaqlanNaqvi S.M. and Rasheed M. 2009. Phosphorus Solubilizing Bacteria: Occurrence, Mechanisms and their Role in Crop Production, Journal of Agricultural and Biological Science, 1(1): 48-58.
Khosravi H. 2013. Application of biofertilizers containing free-living nitrogen-fixing microorganisms in agriculture. Journal Management System, 2(1): 51-63.(In Persian).
Khosravi H. 2013. Azotobacter and its role in the management of soil fertility. Journal Management System, 2(2): 79-94. (In Persian).
Kizilkaya R. 2008. Yield response and nitrogen concentrations of spring wheat (Triticum aestivum) inoculated with Azotobacter chroococcum strains. Ecological Engineering, 33(2): 150-156.
Kizilkaya R. 2009. Nitrogen fixation capacity of Azotobacter spp. strains isolated from soils in different ecosystems and relationship between them and the microbiological properties of soil. Journal of Environmental Biology, 30(1): 73-82.
Lenart A. 2012. Occurrence, characteristic and genetic diversity of Azotobacter chroococcum in various soils of southern Poland. Polish Journal of Environmental Studies, 21(2): 415-424.
Luo H., Chang R., Wang S., Xu J., Zhou X. and Zhang J. 2011. Screening of highly effective potassium bacteria in rhizosphere soil of high- end brand tobacco in Yunnan, South-west china. Journal of gricultural Science, 24: 1816-1817.
Milagres A. M., Machuca A. and Napoleao D. 1999. Detection of siderophore production from several fungi and bacteria by a modification of chrome azurol S (CAS) agar plate assay. Journal of Microbiological Methods, 37(1): 1-6.
Mishustin E.N., Smirnova G.A. and Lokhmachea R.A. 1981. The decomposition of silicates by microorganisms and the use of silicate bacteria fertilizer. Biologic Bull Academic Science, 8: 400-409.
Motsara M. R. and Roy R. N. 2008. Guide to laboratory establishment for plant nutrient analysis (Vol. 19). Rome: Food and Agriculture Organization of the United Nations.
 
Noreen S., Ali B. and Hasnain S. 2012. Growth promotion of Vigna mungo (L.) by Pseudomonas spp. exhibiting auxin production and ACC-deaminase activity. Annals of microbiology, 62(1): 411-417.
Öborn I., Andrist Rangel Y., Askekaard M., Grant C. A., Watson C. A. and Edwards A. C. 2005. Critical aspects of potassium management in agricultural systems. Soil Use and Management, 21(1): 102-112.
Rajaie S., Raiesi F., Alikhani, H. and Givi J. 2005. Potential  use of native Azotobacter Chroococcum Strains as a Plant Growth Promoting Biofertilizer (PGPB) in some Wheat Fields of Chaharmahal  va Bakhtiari Province. A thesis, Master of Science.University of Sharekord.
Rodrı́guez H. and Fraga R. 1999. Phosphate solubilizing bacteria and their role in plant growth promotion. Biotechnology advances, 17(4): 319-339.
Rubio E. J., Montecchia M. S., Tosi M., Cassán F. D., Perticari A. and Correa O. S. 2013. Genotypic Characterization of Azotobacteria Isolated from Argentinean Soils and Plant-Growth-Promoting Traits of Selected Strains with Prospects for Biofertilizer Production. The Scientific World Journal, 2013.
Safari Sinegani A. A. and Rashidi T. 2011. Changes in phosphorus fractions in the rhizosphere of some crop species under glasshouse conditions. Journal of Plant Nutrition and Soil Science, 174(6): 899-907.
Safari Sinegani A.A., Sharifi Z. and Safari Sinegani M. 2010. Methods in Applied Microbiology. Bu- Alisina university press. Iran, 457p. (In Persian).
Sarikhani M.R., Khoshru B. and Oustan Sh. 2016. Efficiency of some bacterial strains in potassium release from mica and phosphate solubilization under in vitro conditions. Geomicrobiology Journal, 33: 832-838.
Schwyn B. and Neilands JB. 1987. Universal chemical assay for the detection and determination of siderophores. Analytical Biochemistry, 160: 47–56.
Seeley H.W. and Vandemark P.J. 1970. Microbes in action: A laboratory manual of microbiology. D. P. Tarapo Revale Sons and Company Ltd., Bombay, pp. 86-95.
Sperber J. I. 1958. The incidence of apatite-solubilizing organisms in the rhizosphere and soil. Crop and Pasture Science, 9(6): 778-781.
Steenhoudt O. and Vanderleyden J. 2000. Azospirillum, a free-living nitrogen-fixing bacterium closely associated with grasses: genetic, biochemical and ecological aspects. FEMS microbiology reviews, 24(4): 487-506.
Sugumaran P. and Janarthanam B. 2007. Solubilization of potassium-containing minerals by bacteria and their effect on plant growth. World Journal of Agricultural Science, 3: 350-355.
Tejera N., Lluch C., Martinez-Toledo M. V. and Gonzalez-Lopez J. 2005. Isolation and characterization of Azotobacter and Azospirillum strains from the sugarcane rhizosphere. Plant and soil, 270(1): 223-232.
Verma L.N. 1993. Biofertiliser in agriculture. In: Organics in soil health and crop production (ed. P.K. Thampan). Peekay Tree Crops Development Foundation, Cochin, India, pp. 152–183.
Vessey J.K. 2003. Plant growth promoting rhizobacteria as biofertilizers. Plant Soil, 255: 571-586.
Whitelaw M.A. 2000. Growth promotion of plants inoculated with phosphate-solubilizing fungi. Advances in Agronomy, 69: 99–151.
Wilson M. K., Abergel  R. J., Arceneaux  J. E., Raymond  K. N. and  Byers B. R. 2010. Temporal production of the two Bacillus anthracis siderophores, petrobactin and bacillibactin. Biometals, 23(1): 129-134.
Winkler U. and Zotz G. 2009. Highly efficient uptake of phosphorus in epiphytic bromeliads. Annals of botany, 103(3): 477-484.
Zeng X., Liu X., Tang J., Hu S., Jiang P., Li W. and Xu L. 2012. Characterization and Potassium-Solubilizing Ability of Bacillus Circulans Z 1–3. Advanced Science Letters, 10(1): 173-176.