جداسازی و شناسایی باکتری‌های حل‌کننده روی و بررسی کارایی چند حامل در زنده‌مانی جدایه برتر

نوع مقاله : مقاله پژوهشی

نویسندگان

1 گروه علــوم خاک دانشگاه ارومیه

2 استادیار

3 موسسه تحقیقات حفظ نبات، سازمان تحقیقات، آموزش و ترویج کشاورزی، تهران

4 - دانشیار گروه گیاهپزشکی، دانشکده کشاورزی، دانشگاه ارومیه

چکیده

روی یکی از عناصر کم‌مصرف است که در خاک‌های آهکی عمدتا به شکل‌های کم‌محلول وجود دارد. یکی از راه‌های فراهمی روی مورد نیاز گیاه در این شرایط، استفاده از میکروارگانیسم‌ها در قالب کود زیستی است. برای عرضه این کودها نیاز به حامل مناسب می‌باشد. لذا، این تحقیق به منظور جداسازی و شناسایی باکتری‌های حل‌کننده روی از خاک‌ها و بررسی زنده‌مانی سویه برتر در حامل‌های مختلف به مدت نه ماه در قالب طرح پایه کاملا تصادفی دو فاکتوره (حامل و زمان) با سه تکرار اجرا گردید. حامل‌های جامد، شامل پرلیت، پیت ماس، خاک اره، تفاله چغندرقند، کود دامی، ورمی‌کمپوست، آزولا، بیوچار (آزولا و سیب)، سبوس و تالک بودند. در این تحقیق، زادمایه‌های باکتری‌یایی تهیه شده با جمعیت اولیه یکسان پس از نگهداری در دمای اتاق از نظر توان زنده‌مانی باکتری مورد مقایسه قرار گرفتند. برای شمارش باکتری‌های زنده در حامل‌های میکروبی، در هر ماه از روش شمارش MPN استفاده شد. نتایج نشان داد که از 24 نمونه خاک، تعداد 15 جدایه باکتری‌ حل‌کننده روی جداسازی و خالص‌سازی شد. پنج جدایه‌ی برتر از لحاظ انحلال ترکیب نامحلول روی، جهت شناسایی مولکولی مورد بررسی قرار گرفتند. بر اساس توالی ژن 16S rRNA، سه جدایه‌ به جنس سودوموناس و دو جدایه به انتروباکتر تعلق داشتند. از بین باکتری‌های شناسایی شده باکتری سودوموناس فلورسنس (Ur 22) توان کمی (2/75 میلی‌گرم در لیتر) و توان کیفی (28/2) بالایی در انحلال‌ ترکیب نامحلول فسفات روی داشت. همچنین نتایج نشان داد که بیشترین لگاریتم جمعیت شمارش شده بعد از گذشت نه ماه در حامل بیوچار سیب ( log cfu g-107/5) و بیوجار آزولا ( log cfu g-107/5) و کمترین جمعیت شمارش شده در حامل تالک ( log cfu g-111/2) به دست آمد. بطور کلی می‌توان بیان داشت که از بین حامل‌های مورد آزمایش، بیوچار بعلت ویژگی‌های خوب و مقرون به‌صرفه بودن از نظر اقتصادی توصیه می‌شود.

کلیدواژه‌ها


عنوان مقاله [English]

Isolation and Identification of Zn solubilising bacteria and Evaluation of various Carrier Efficacy in Superior Isolate Survival

نویسندگان [English]

  • Fatemeh Hashemnejad 1
  • mohsen barin 2
  • Maryam Khezri 3
  • youbert ghoosta 4
1 Department of Soil Science, Urmia University, Urmia, Iran.
2
3 Iranian Research Institute of Plant Protection, Agricultural Research, Education and Extension Organization (AREEO), Tehran, Iran.
4 Associate Prof. of Mycology and Plant Pathology, Department of Plant Pathology, Urmia University, Urmia, Iran
چکیده [English]

Zinc (Zn) is one of the micronutrient that is in calcareous soils in low solubility forms. One of the ways of supplying Zn of the plant in these conditions is the use of microorganisms as biofertilizer. A suitable carrier is needed to supply these fertilizers. Therefore, this study was performed to isolation and identification of Zn solubilizing bacteria from soil and to evaluate the survival of the superior strain in various carriers during nine months in a completely randomized two-factor design (carrier and Time) was performed with three replications. Solid carriers included perlite, peat moss, sawdust, sugar beet waste, manure, vermicompost, azolla, biochar (azolla and apple), bran and talc. In this study, bacterial inoculants prepared with the same initial population after storage at room temperature were compared for the survival of the bacteria. For counting the bacteria in microbial carriers, MPN counting method was used in each month. The results showed that out of 24 samples of the rhizosphere soil, 15 strains of Zn-solubilizing bacteria were isolated and purified. Five isolates found to be the most efficient in solubilizing insoluble Zn compounds were examined for molecular identification. Based on gene 16S rRNA sequencing, three of the isolates belonged to the genus of Pseudomonas, and two to Enterobacter. The highest solubility index (2.28) was related to Pseudomonas fluorescens strains which were used for inoculation of microbial carriers by Ur 22 strain. The results of bacterial count in carriers showed that among the tested carriers, the highest population counted after nine months in carrier of apple biochar (5.07 log cfu g-1) and azolla biochar (5.07 log cfu g-1), and the lowest population was obtained in talc carrier (2.11 log cfu g-1). In general, it can be said that among the tested carriers, Biochar is recommended due to its good properties and cost-effectiveness.

کلیدواژه‌ها [English]

  • Biofertilizer
  • Pseudomonas fluorescens
  • Solid carrier
  • Survival
Alexander M. 1982. Most probable number method for microbial populations. In: Page, A.L., Miller, R.H., Keeney, D.R. (Eds.), Methods of Soil Analysis, Part 2, 2nd Ed. American Society of Agronomy, Madison, WI, pp. 815-820.
Andreini C., Bertini I., and Rosato A. 2009. Metalloproteomes: a bioinformatic approach. Accounts of Chemical Research, 42: 1471–1479.
Araujo J., Díaz-Alcántara C.A., Urbano B., and González-Andrés F. 2020. Inoculation with native Bradyrhizobium strains formulated with biochar as carrier improves the performance of pigeonpea (Cajanus cajan L.). European Journal of Agronomy, 113: p.125985.
Arora N.K., Khare E., and Maheshwari D.K. 2010. Plant growth promoting rhizobacteria: constraints in bioformulation, commercialization, and future strategies. In Plant growth and health promoting bacteria Springer, Berlin, Heidelberg. pp. 97-116.
Badr M.A. 2006. Efficiency of K-feldspar combined with organic materials and silicate dissolving bacteria on tomato yield. Journal of Applied Sciences Research, 2:1191–1198.
Bashan Y. 1998. Inoculants of plant growth-promoting bacteria for use in agriculture. Biotechnology advances, 16(4): 729-770.
Bonnier, C. (1960). Symbiose Rhizobium-légumineuses: Aspects particuliers aux régions tropicales. Annales de l'Institut Pasteur, 98: 537-556. ‏
Brenner D.J., Krieg N.R., and Statey J.T. 2005. Editors. Bergie's manual systematic bacteriology. 2nd ed. New York: Springer; Part C.
Broadley M.R., White P.J., Hammond J.P., Zelko I., and Lux, A. 2007. Zinc in plants. New Phytologist. 173: 677–702.
Castaldi S., Riondino M., Baronti S., Esposito F., Marzaioli R., Rutigliano F., and Miglietta F. 2011. Impact of biochar application to a Mediterranean wheat crop on soil microbial activity and greenhouse gas fluxes. Chemosphere, 85(9): 1464-1471.
Chao W.L. and Alexander M. 1984. Mineral soils as carriers for Rhizobium inoculants. Applied and Environmental Microbiology, 47(1): 94-97. 
Chen, J. H. 2006. The combined use of chemical and organic fertilizers and/or biofertilizer for crop growth and soil fertility. In International workshop on sustained management of the soil-rhizosphere system for efficient crop production and fertilizer use. Land Development Department Bangkok Thailand. 16: 20.
Costerousse B., Schönholzer-Mauclaire L., Frossard E., and Thonar C. 2018. Identification of Heterotrophic Zinc Mobilization Processes among Bacterial Strains Isolated from Wheat Rhizosphere (Triticum aestivum L.). Applied and Environmental Microbiology, 84: 1-16.
Date R. A. 1988. Colonization of the rhizosphere by root-nodule bacteria. Microbiology in Action. Research Studies Press, Chichester. Eds. I Kennedy and W Murrell.
Egamberdiyeva D., Juraeva D., Poberejskaya S., Myachina O., Teryuhova P., Seydalieva L., and Aliev A. 2004. Improvement of wheat and cotton growth and nutrient uptake by phosphate solubilizing bacteria. In Proceeding of 26th annual conservation tillage conference for sustainable agriculture, Auburn. Pp. 58-65.
Fages J. 1992. An industrial view of Azospirillum inoculants: formulation and application technology. Symbiosis, 13: 15-26.
Fasim F., Ahmed N., Parsons R., and Gadd G. M. 2002. Solubilization of zinc salts by a bacterium isolated from the air environment of a tannery. FEMS microbiology letters, 213(1):1-6.
Ferreira E.M. 2005. Residues of the cork industry as carriers for the production of legume inoculants. Silva Lusitana, 13(2):159-167.
Gandhi A., Muralidharan G., Sudhakar E., and 2Murugan A. 2014. Screening for elite zinc solubilizing bacterial isolate from rice rhizosphere environment. International Journal of Recent Scientific Research. 5:2201-2204
Ghasemi Piranloo, F., Sarikhani, M. R., Najafi, N. 2019. Study of Enterobacter cloacae viability in several solid carriers and the effect of prepared offspring on wheat germination and growth. Agricultural Knowledge and Sustainable Production, 29:167-180. (In Persian)
Girgis M.G.Z., Khalil H.M., and Sharaf M.S. 2008. In vitro evaluation of rock phosphate and potassium solubilizing potential of some Bacillus strains. Australian Journal of Basic and Applied Sciences, 2(1): 68-81.
Huber D., El-Nasshar H., Moore L., Mathre D., and Wagner J. 1989. Interaction between a peat carrier and bacterial seed treatments evaluated for biological control of the take-all diseases of wheat (Triticum aestivum L.). Biology and fertility of soils, 8(2): 166-171.
Khavarzi K., and Rajali F. 2000. Use of some inexpensive materials as a carrier of Bradyrhizubium japonica. Journal of Soil and Water, 14: 45-36. (In Persian)
Khavazi K., Asgharzadeh A., Rajali F., Asadi Rahmani H., Besharti H., Fallah Nusratabad A.R., Afshari M., Khosravi H., Khademi Z., Bazarga K.  2013. Instructions on how to study biofertilizers. Soil and Water Research Institute, SADES Publications, Pp. 44. (In Persian)
Leach A., and Mumford J. 2008. Pesticide Environmental Accounting: A method for assessing the external costs of individual pesticide applications. Environmental pollution, 151(1): 139-147.
Lelliot R.A., and Stead D.E. 1987. Methods for The Diagnosis of Bacterial Diseases of Plants. Blackwell Scientific Publications.
Llop P., Caruso P., Cubero J., Morente C. and Lopez M.M. 1999. A simple extraction procedure for efficient routine detection of pathogenic bacteria in plant material by polymerase chain reaction. Journal of Microbiological Methods, 37(1): 23-31.
Mashhadi Asghari S., and Ali Asgharzadeh N. 2003. Comparison of the efficacy of several carriers of Sinorizobium meliloti to produce alfalfa inoculum. Journal of Science and Technology Agricultural and Natural Resources, 8:63-75. (In Persian)
Meena V.S., Maurya B.R., and Bahadur I. 2014. Potassium solubilization by bacterial strain in waste mica. Bangladesh Journal of Botany, 43(2):235–237.
Motsara M.R., Bhattacharyya B., and Srivastava B. 1995. Biofertiliser technology, marketing and usage: a sourcebook-cum-glossary. Fertiliser Development and Coordination Organization, New Delhi.
Muhammad N., Dai Z, Xiao K., Meng J., Brookes P.C., Liu X., and Xu J. 2014. Changes in microbial community structure due to biochars generated from different feedstocks and their relationships with soil chemical properties. Geoderma, 226: 270-278.
Mukhtar S., Shahid I., Mehnaz S., and Malik K.A. 2017. Assessment of two carrier materials for phosphate solubilizing biofertilizers and their effect on growth of wheat (Triticum aestivum L.). Microbiological research, 205:107-117.
Mumtaz, M.Z., Ahmad, M., Jamil, M., Hussain, T. 2017. Zinc solubilizing Bacillus spp. potential candidates for biofortification in maize. Microbiological Research, 202: 51–60.
Myers S.S., Wessells K.R., Kloog I., Zanobetti A., Schwartz J. 2015. Effect of increased concentrations of atmospheric carbon dioxide on the global threat of zinc deficiency: a modelling study. The Lancet Global Health, 3: 639–645.
Shaikh S.S., and Saraf M.S. 2017. Optimization of growth conditions for zinc solubilizing plant growth associated bacteria and fungi. Journal of Advanced Research in Biotechnology, 2(1):1–9.
Page A.L., Miller R.H., and Keeney D.R. 1982. Method of soil analysis. part 2- chemical and microbiological properties. 2nd ed., ASA pub. Buxton, Madison, WI, 159-166.
Rivera‐Utrilla J., Bautista‐Toledo I., Ferro‐García M.A. and Moreno‐Castilla C. 2001. Activated carbon surface modifications by adsorption of bacteria and their effect on aqueous lead adsorption. Journal of Chemical Technology and Biotechnology, 76(12): 1209-1215.
Salem M., Kohler J., Wurst S., and Rillig M.C. 2013. Earthworms can modify effects of hydrochar on growth of Plantago lanceolata and performance of arbuscular mycorrhizal fungi. Pedobiologia, 56(4): 219-224.
Samonin V., and Elikova E. 2004. A study of the adsorption of bacterial cells on porous materials. Microbiology, 73(6): 696-701.
Saravanan V.S., Subramoniam S.R., and Raj S.A. 2003. Assessing in vitro solubilization potential of different zinc solubilizing bacterial (ZSB) isolates. Brazilian Journal of Microbiology, 35(1-2):121-125.
Schaad N.W., Jones, J.B., and Chun W. 2001. Laboratory Guide for Identification of Plant Pathogenic Bacteria, (Third Ed.) St. Plant Pathology, 50(6):812-814.
Shaikh S.S., Saraf M.S. 2017. Optimization of growth conditions for zinc solubilizing plant growth associated bacteria and fungi. Journal of Advanced Research in Biotechnology, 2(1):1–9
Shariati S., Alikhani H.A., and Pourbabaei A. 2013. Application of vermicompost as a carrier of phosphate solubilizing bacteria (Pseudomonas fluorescens) in increase growth parameters of maize. International Journal of Agronomy and Plant Production, 4(8):2010-2017.
Somasegaran P., and Hoben H. J. 1994. Handbook for Rhizobia. Methods in Legume-Rhizobium Technology, Springer Science & Business Media. Pp.332-341.
Stephens J., and Rask H. 2000. Inoculant production and formulation. Field Crops Research, 65(2): 249-258.
Tallapragada P., Seshachala U. 2012. Phosphate-solubilizing microbes and their occurrence in the rhizospheres of Piper betel in Karnataka, India. The Turkish Journal of Biology, 36: 25-35.
Tamboli R.R. 2019. Isolation and Characterization of Zinc Solubilizing Bacteria from Rhizosphere soil of Latur District, Marathwada, India. Biosciences biotechnology research asia, 16: 797-803
Vahedi R, and Rasoili-Sadaghiani M.H. 2020. Bioavailability of Selected Micronutrients as Affected by Biochar and Compost of Trees Pruning in the Presence of Mycorrhiza at Wheat Rhizosphere. Journal of Water and Soil Science, 23 (4) :155-170. (In Persian)
Vahedi R., Rasouli-Sadaghiani M., Barin M., Vetukuri R.R. 2021. Interactions between Biochar and Compost Treatment and Mycorrhizal Fungi to Improve the Qualitative Properties of a Calcareous Soil under Rhizobox Conditions. Agriculture, 11: 993.
Van Elsas J.D., and Heijnen C. 1990. Methods for the introduction of bacteria into soil: a review. Biology and fertility of soils, 10(2): 127-133.  
Vessey J. K., 2003. Plant growth promoting rhizobacteria as biofertilizers. Plant and soil, 255(2): 571-586.
Weisburg W.G., Barns S.M., Pelletier D.A., and Lane D. J. 1991. 16S ribosomal DNA amplification for phylogenetic study. Journal of bacteriology, 173(2): 697-703.
Xavier I.J., and Holloway Leggett M. 2004. Development of rhizobial inoculant formulation. Crop Management 3(1):1-6.
Zaheer A., Malik A., Sher A., Mansoor Qaisrani M., Mehmood A., Ullah Khan S., Ashraf M., Mirza Z., Karim S., and Rasoo M. 2019. Isolation, characterization, and effect of phosphate-zinc-solubilizing bacterial strains on chickpea (Cicer arietinum L.) growth. Saudi Journal of Biological Sciences, 26: 1061–1067.