جداسازی باکتری‌های متحمل به مس از یک خاک آلوده، شناسایی و بررسی خصوصیات محرک رشدی آنها

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشجوی کارشناسی ارشد دانشکده کشاورزی دانشگاه ولی‌عصر (عج) رفسنجان

2 استادیار گروه علوم خاک دانشکده کشاورزی دانشگاه ولی‌عصر (عج) رفسنجان

3 دانشیار گروه علوم خاک دانشکده کشاورزی دانشگاه ولی‌عصر (عج) رفسنجان

چکیده

تلقیح گیاهان با باکتری­های ریزوسفری محرک رشد موثر و مقاوم به فلزات سنگین در خاک­های آلوده به منظور افزایش کارایی گیاه­پالایی و استخراج فلز سنگین از خاک توسط گیاه امری ضروری می­باشد. در این جهت این پژوهش به منظور جداسازی، شناسایی و صفات محرک رشدی باکتری­های مقاوم مس از یک خاک آلوده به مس انجام شد. ابتدا شش نمونه خاک ریزوسفری گیاهان بومی منطقه خاتون­آباد سرچشمه استان کرمان جمع­آوری شد و نهایتا شش جدایه باکتری از نمونه­های خاک ریزوسفری جمع­آوری و خالص­سازی شد. نتایج شناسی فنوتیپی، بیوشیمیایی و ژنتیکی نشان داد که پنج جدایه از جنس سودوموناس و یک جدایه از جنس باسیلوس بودند. سپس حداقل غلظت بازدارنده[1] مس، میزان تولید سیدروفور، تولید اکسین، سیانید هیدروژن، توانایی حل فسفات­های معدنی، در محیط جامد و مایع، اندازه‌گیری توان حل­کنندگی ترکیبات کم محلول روی در محیط جامد و مایع و تولید -ACC دآمیناز باکتری­ها بررسی شد. نتایج نشان داد تمام جدایه­ها نسبت به غلظت­های مختلف مس از خود مقاومت نشان دادند. بیشترین مقاومت نسبت به غلظت بازدارنده مس مربوط به سویه­های K4 و K5 در غلظت 400 میلی­گرم بر لیتر مس بود. همچنین در رابطه با تولید سیدروفور، بجز سویه­های K6 که توانایی تولید سیدروفور را در محیط CAS نداشت، پنج سویه دیگر توانایی تولید سیدروفور را در این محیط دارا بودند و سویه ی K5 بیشترین میزان تولید سیدروفور با نسبت هاله به کلنی 56/1 را به خود اختصاص داد. در بین جدایه­ها، دو جدایه K4 و K6 قادر به تولید اکسین نبودند. تولید اکسین توسط جدایه K1 تفاوت معنی­داری با دیگر جدایه­های مورد بررسی داشته و با مقدار 07/2 میلی­گرم در لیتر بیشترین میزان تولید اکسین در بین جدایه­ها را دارا بود. نتایج همچنین نشان داد که از بین جدایه­ها، تنها جدایه K4 توانایی تولید سیانید هیدروژن را با درجه چهار (نسبتا زیاد) دارا بوده و دیگر جدایه ها با اختصاص درجه یک مبنی بر عدم تولید سیانید هیدروژن گزارش شدند. نتایج ارزیابی انحلال تری­کلسیم فسفات[2] در محیط مایع PKV توسط جدایه­ها نشان داد که. سویه K1، K2، و K3 به ترتیب با میزان 706، 661 و 588 میلی­گرم در لیتر بیشترین حل­کنندگی فسفر را به خود اختصاص داده و با یکدیگر تفاوت معنی داری نداشتند. نتایج بررسی­ها نشان داد که در محیط جامد حاوی کربنات روی دو سویه K4 و K5 به ترتیب با مقادیر 910/0 و 850/0 نسبت هاله به کلونی تفاوت معنی­داری با دیگر سویه­ها داشته و بقیه سویه­ها شامل K1، K2، و K3 و K6 توانایی انحلال کربنات روی از خود نشان ندادند. همچنین انحلال اکسید روی در محیط جامد توسط سویه K4 با 11/2 نسبت هاله به کلونی بیشترین میزان انحلال اکسید روی و سویه­های K1، K2، و K3 و K6 قادر به انحلال اکسید روی در محیط جامد نبودند. سویه­های  K1، K2، K3 و K6 قادر به انحلال ترکیبات کم محلول اکسید و کربنات روی در محیط مایع نبودند. سویه K4 با 5/12 میلی­گرم در لیتر با بیشترین میزان انحلال کربنات روی در محیط مایع را داشت و همچنین بیشترین انحلال اکسید روی در محیط مایع به سویه K4 با 76/9 میلی­گرم در لیتر نسبت داده شد. با توجه به نتایج بدست آمده می‌توان از این جدایه‌ها در جهت افزایش کارایی گیاه‌پالایی این عنصر در خاک‌های آلوده استفاده کرد.



[1] .Minimum Inhibitory Concentration (MIC)


[2] .Tricalcium Phosphate (TCP)

کلیدواژه‌ها

عنوان مقاله [English]

Isolation, identification and plant growth promoting characteristics of Cu-tolerant bacteria in a contaminated soil

نویسندگان [English]

  • Z. Karimi 1
  • P. AbbaszadehDahaji 2
  • A. R. Akhgar 3
  • M Hamidpour 3
1 M.Sc. Student, Department of Soil Science, Vali-e-Asr University
2 Assistant professor, Department of Soil Science, Vali-e-Asr University
3 Associate professor, Department of Soil Science, Vali-e-Asr University
چکیده [English]

Plant inoculation with effective plant growth promoting and heavy metal resistant rhizobacteria is an efficient method to improve the phytoremediation efficiency and extraction of heavy metals from contaminated soils. With respect to this reason, astudy was conducted to isolate, identify and evaluate the growth stimulating traits of copper-resistant bacteria from a Cu-contaminated soil. Six rhizospheric soil samples of indigenous plants were collected in Sarcheshmeh Khatoonabad, Kerman province. After screening, six strains were selected. Phenotypic, biochemical and genetic identification data showed that five and one isolates were belong to Pseudomonas and Bacillus genera respectively. Then, the minimum inhibitory concentration of copper, siderophore production, auxin and hydrogen cyanide production, the ability to solubilize mineral phosphate in solid and liquid medium, the ability to solubilize Zn low soluble compounds in solid and liquid medium and production of ACC-deaminase were investigated. The results indicated that all of the isolates were resistant to various concentrations of copper. The maximum resistance to inhibitory concentration of copper was related to K4 and K5 strains at a concentration of 400 mg L-1 of copper. All of the strains were able to produce siderophore in CAS medium except for K6 strains and strain K5 was also the superior in siderophore production. Among the strains, K4 and K6 were not able to produce auxin. There was also a significant difference between auxin productions by isolates K1 with the other isolates, with the highest auxin production (2.07 mg L-1). The results also demonstrated that among the strains, only K4 strain was capable of producing hydrogen cyanide (degree: four=relatively high) and the other isolates were reported as non-production of hydrogen cyanide (degree: one). The results of Three Calcium Phosphate (TCP) solubilizing by isolates in PKV broth medium showed that K1, K2, and K3 strains had the most phosphorus solubility (706, 661 and 588 mg L-1) respectively and there wasn’t significant difference between them. The results demonstrated that K4 and K5 isolates with 0.910 and 0.850 halos to colony diameter had a significant difference with other isolates in solid medium containing ZnCO3 and the rest of the strains, K1, K2, K3 and K6 did not show the ability to solubilize the zinc carbonate as well. The solubilizing of solid zinc oxide in solid medium by K4 strain (halo:colony, 2.11) was the highest solubility of zinc oxide and K1, K2, K3 and K6 strains were not able to dissolve the zinc oxide in the solid medium. They did not show the ability to dissolve the low soluble zinc oxide nor the zinc carbonate compounds. K4 strain had the highest zinc carbonate solubility in the broth (12.5 mg L-1) and the highest solubility of zinc oxide in the broth was attributed to K4 strain (9.76 mg L-1). According to the results, these isolates can be used for enhancing phytoremediation efficiency of this element in contaminated soils.

کلیدواژه‌ها [English]

  • Auxin
  • Plant growth promoting rhizobacteria
  • Minimum Inhibitory Concentration
  • Siderophore

مراجع

  1. حجت نوقی، ف.، اخگر، ع.، اسفندیارپور، ع. و خاوازی، ک. 1392. ارزیابی جمعیت و خواص محرک رشدی باکتری­های اندوریزوسفری، ریزوسفری و غیر ریزوسفری در نهال­های پسته. نشریه دانش آب و خاک، جلد 23(4): 234-215.
  2. Abbas-Zadeh, P., Saleh-Rastin, N., Asadi-Rahmani, H., Khavazi, K., Soltani, A., Shoary-Nejati, A.R. and Miransari, M. 2010. Plant growth promoting activities of fluorescent pseudomonads, isolated from the Iranian soils. Acta Physiologiae Plantarum. 32: 281- 288.
  3. Ahemad, M. and Khan, M.S. 2012. Evaluation of plant growth promoting activities of rhizobacterium Pseudomonas putida under herbicide-stress. Annals of Microbiology. 62: 1531–1540.
  4. Ahemad, M. and Kibret, M. 2013. Mechanisms and applications of plant growth promoting rhizobacteria: current perspective. Journal of King Saud University Science. 26(1): 1-20.
  5. Ahemad, M. and Malik, A. 2011. Bioaccumulation of heavy metals by zinc resistant bacteria isolated from agricultural soils irrigated with wastewater. Bacteriology Journal. 2: 12–21.
  6. Alexander, D.B. and Zuberer, D.A. 1991. Use of chrome azurol S reagents to evaluate siderophore production by rhizosphere bacteria. Biology and Fertility of Soils. 12: 39-45.
  7. Amico, E.D., Cavalca, L. and Andreoni, V. 2005. Analysis of rhizobacterial communities in perennial Graminaceae from polluted water meadow soil, and screening of metal-risistant, potentially plant growth-promoting bacteria. FEMS Microbiology Ecology. 52: 153-162.
  8. Asghar, H.N., Zahir, Z.A., Arshad, M. and Khaliq, A. 2002. Relationship between in vitro production of auxins by rhizobacteria and their groth-promoting activities in Brassica juncea L. Biology Fertility Soils. 35: 231-237.
  9. Babalola, O.O. 2010. Beneficial bacteria of agricultural importance. Biotechnology Letters. 32(11): 1559–1570.
  10. Belimov, A.A., Hontzeas, N., Safronova, V.I., Demchinskaya, S.V., Piluzza, G., Bullitta, S. and Glick, B.R. 2005. Cadmium-tolerant plant growth-promoting bacteria associated with the roots of Indian mustard (Brassica juncea L. Czern.). Soil Biology Biochemistry. 37: 241–250.
  11. Bent, E., Tuzan, S., Chanway, C.P. and Enebak, S. 2001. Alteration in plant growth and in root hormone levels of lodgepole pines inoculated with rhizobacteria. Canadian Journal of Microbiology. 47: 793-800.
  12. Bhatt, P.V. and Vyas, B.R.M. 2014. Screening and Characterization of Plant Growth and Health Promoting Rhizobacteria. International Journal of Current Microbiology Applied Science. 3(6): 139-155.
  13. Chibuike, G.U. and Obiora, S.C. 2014. Heavy Metal Polluted Soils: Effect on Plants and Bioremediation Methods. Hindawi Publishing Corporation, Applied and Environmental Soil Science. 2014 (12 pages).
  14. Desai, S., Praveen Kumar, G., Sultana, U., Pinisetty, S., Ahmed, M.H., Amalraj, D. and Reddy, G. 2012. Potential microbial candidate strains formanagement of nutrient requirements of crops. African Journal of Microbiology Research. 6 (17): 3924–3931.
  15. Donate-Correa, J., Leon-Barrios, M. and Perez-Galdona, R. 2004. Screening for plant growth-promoting rhizobacteria in Chamaecytisus proliferus (tagasaste), a forage tree-shrub legume endemic to the Canary Island. Plant and Soil. 266: 261-272.
  16. Egamberdieva, D. 2009. Alleviation of salt stress by plant growth regulators and IAA producing bacteria in wheat. Acta Physiologiae Plantarum. 31: 861–864.
  17. Ganesan, V. 2008. Rhizoremediation of cadmium soil using a cadmium-resistant plant growth-promoting rhizopseudomonad. Current Microbiology. 56: 403–407.
  18. Glick, B.R., Penrose, D.M. and Li, J. 1998. A model for the lowering of plant ethylene concentrations by plant growth promoting bacteria. Journal of Theoretical Biology. 190: 63–68.
  19. Glick, B.R. 2003. Phytoremediation: synergistic use of plants and bacteria to clean up the environment. Biotechnology. 21: 383–393.
  20. Glick, B.R. 2012. Plant Growth-Promoting Bacteria: Mechanisms and Applications. Hindawi Publishing Corporation. Scientifica. 2015 (15 pages).
  21. Glick, B.R., Penrose, D.M. and Li, J. 1998. A model for the lowering of plant ethylene concentrations by plant growth promoting bacteria. Journal of Theoretical. Biology. 190: 63–68. ##
  22. Goteti, P.K., Emmanuel, L.D.A., Desai, S. and Shaik, M.H.A. 2013. Prospective Zinc Solubilising Bacteria for Enhanced Nutrient Uptake and Growth Promotion in Maize (Zea mays L.). International Journal of Microbiology. 2013 (7 pages).
  23. Han, J., Sun, L., Dong, X., Cai, Z., Sun, X., Yang, H., Wang, Y. and Song, W. 2005. Characterization of a novel plant growth-promoting bacteria strain Delftia tsuruhatensi s HR4 both as a diazotroph and a potential biocontrol agent against various plant pathogens. System Applied Microbiology. 28: 66–76.
  24. Haque, N., Peralta-Videa, J. R., Jones, G. L., Gill, T. E. and Gardea-Torresdey, J. L. 2008. Screening the phytoremediation potential of desert broom (Baccharis sarothroides Gray) growing on mine tailings in Arizona, USA. Environmental Pollution, 153: 362–368.
  25. He, L.Y., Zhang, Y.F., Ma, H.Y., Su, L.N., Chen, Z.J., Wang, Q.Y., Meng, Q. and Fang, S.X. 2010. Characterization of copper resistant bacteria and assessment of bacterial communities in rhizosphere soils of copper-tolerant plants. Applied Soil Ecology. 44: 49–55.
  26. Huang, X.D., El-Alawi, Y., Penrose, D.M., Glick, B.R. and Greenberg, B.M. 2004a. A Multi-process Phytoremediation System for Removal of Polycyc lic Aromatic Hydrocarbons from Contaminated Soils. Environmental Pollution. 130: 453-463.
  27. Huang, X.D., El-Alawi, Y., Penrose, D.M., Glick, B.R. and Greenberg, B.M. 2004b. Responses of Three Grass Species to Creosote during Phytoremediation. Environmental Pollution. 130: 465-476.
  28. Jeon, J.S., Lee, S.S., Kim, H.Y., Ahn, T.S and Song, H.G. 2003. Plant growth promoting in soil by some inoculated microorganism. The Journal of Microbiology. 41(4): 271-276.
  29. Jiang, C.Y., Sheng, X.F., Qian, M. and Wang, Q.Y. 2008. Isolation and characterization of a heavy metal resistant Burkholderia sp. From heavy metal-contaminated paddy field soil and its potential in promoting plant growth and heavy metal accumulation in metal polluted soil. Chemosphere. 72: 157–164.
  30. Khan, M.U., Sessitsch, A., Harris, M., Fatima, K., Imran, A., Arslan, M., Shabir, G., Khan, Q.M. and Afzal, M. 2015. Cr-resistant rhizo- and endophytic bacteria associated with Prosopis juliflora and their potential as phytoremediation enhancing agents in metal-degraded soils. Frontiers in Plant Science. 5: 755.
  31. Kloepper, J.W., Zablotowick, R.M., Tipping, E.M. and Lifshitz, R. 1991. Plant growth promotion mediated by bacterial rhizosphere colonizers. In: Keister, D.L., Cregan, P.B. (Eds.), The Rhizosphere and Plant Growth. Kluwer Academic Publishers, Dordrecht, Netherlands, p: 315–326.
  32. Madhaiyan, M., Poonguzhali, S. and Sa, T. 2007. Metal tolerating methylotrophic bacteria reduces nickel and cadmium toxicity and promotes plant growth of tomato (Lycopersicon esculentum L.). Chemosphere. 69: 220–228.
  33. Marchesi, J.R., Sato, T., Weightman, A.J., Martin, T.A., Fry, J.C., Hiom, S.J. and Wade, W.G. 1998. Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16S rRNA. Applied and Environmental Microbiology. 64: 795–799.
  34. Meunchang, S., Thongra, P., Sanoh, S., Kaewsuralikhit, S. and Ando, S. 2006. Development of rhizobacteria as a biofertilizer for rice production. International workshop on Sustained Management of the Soi- Rhizosphere System for Efficient Crop Production and Fertilizer Use. Bangkok, Thailand, p: 6-20.
  35. Prasad, M.N.V. and Freitas, H.M.O. 2003. Metal hyper-accumulation in plants-Biodiversity prospecting for phytoremedation technology. Electronic Journal of Biotechnology. 6(3): 285-321.
  36. Wuana, R.A. and Okieimen, F.E. 2011. HeavyMetals in Contaminated Soils: A Review of Sources, Chemistry, Risks and Best Available Strategies for Remediation. International Scholarly Research Network ISRN Ecology. 2011 (20 pages).
  37. Saharan, B.S. and Nehra, V. 2011. Plant Growth Promoting Rhizobacteria: A Critical Review. Life Sciences and Medicine Research. 2011 (30 pages).
  38. Saravanan, V.S., Madhaiyan, M. and Thangaraju, M. 2007. Solubilization of zinc compounds by the diazotrophic, plant growth promoting bacterium Gluconacetobacter diazotrophicus. Chemosphere. 66: 1794–1798.
  39. Saravanan, V.S., Subramoniam, S.R. and Raj, S.A. 2004. Assessing in vitro solubilization potential of different zinc solubilizing bacteria (ZSB) isolate. Brazilian Journal of Microbiology, 34: 121-125.
  40. Schaad N., Jones J.B. and Chun W., 2001: Laboratory guide for identification of plant pathogenic bacteria. APS Press, St. Paul, Minnesota, USA.
  41. Sevgi, E., Coral, G., Gizir, A.M. and Sangun, M.K. 2010. Investigation of heavy metal resistance in some bacterial strains isolated from industrial soils. Turkish Journal of Biology. 34: 423-431.
  42. Song Y., Ji, J., Mao, C., Yang, Z., Yuan, X., Ayoko, G.A. and Fros R.L. 2010. Heavy metal contamination in suspended soils of Changjiang River environmental implications. Geoderma. 159: 286–295.
  43. Sun, L., He, L., Zhang,. Y, Zhang, W., Wang, Q. and Sheng, X. 2009. Isolation and biodiversity of copper-resistant bacteria from rhizosphere soil of Elsholtzia splendens. Acta Microbiologica Sinica. 49(10): 1360-1366.
  44. Tchounwou, P.B., Yedjou, C.G., Patlolla, A.K. and Sutton, D.J. 2012. Heavy metal toxicity and the environment. Molecular, Clinical and Environmental Toxicology. 101: 133-164.
  45. VCI, Copper history/Future, Van Commodities Inc. 2011. http:// trademetalfutures.com /copperhistory.html.
  46. Verma, S.C., Ladha, J.K. and Tripathi, A.K. 2001. Evaluation of plant growth promoting and colonization ability of endophytic diazotrophs from deep water rice. Journal of Biotechnology. 91: 127–141.
  47. Walker, T.S., Bais, H.P., Grotewold, E. and Vivanco, J.M. 2003. Root exudation and rhizosphere biology. Plant Physiology. 132: 44–51.
  48. Zaki, S. and Farag, S. 2010. Isolation and molecular characterization of some copper biosorped strains. International Journal of Environmental Science and Technology. 7(3): 553-560.
زیست شناسی خاک
دوره 5، شماره 2
اسفند 1396
صفحه 95-107

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