شناسایی اندوفیت‌های قارچی در کشت هوازی برنج استان گلستان

نوع مقاله : مقالات پژوهشی

نویسندگان

1 گروه گیاهپزشکی. دانشکده تولید گیاهی.دانشگاه علوم کشاورزی و منابع طبیعی گرگان

2 گروه گیاه‌پزشکی، دانشکده تولید گیاهی، دانشگاه علوم کشاورزی و منابع طبیعی گرگان

3 گروه زراعت، دانشکده تولید گیاهی، دانشگاه علوم کشاورزی منابع طبیعی گرگان

چکیده

برنج از مهم­ترین غلات مورد مصرف غذایی مردم جهان به­ویژه خاورمیانه می­باشد. ایران با داشتن نزدیک به 560 هزار هکتار زمین کشاورزی اختصاص یافته برای کشت برنج و تولید سالانه نزدیک به 5/2 میلیون تن، رتبه جهانی 23 در تولید و 26 در زمین­های تحت کشت و رتبه 13 در مصرف آن را به خود اختصاص داده است. در این پژوهش، به­منظور شناسایی قارچ­های اندوفیت در کشت هوازی برنج استان گلستان، بازدیدهایی از مزارع مختلف برنج هوازی استان گلستان شامل شهرستان­های گرگان، کردکوی، بندرگز، علی­آباد و کلاله طی ماه­های اردیبهشت تا شهریور سال 1398-1399 به­عمل آمد. پس از خالص­سازی، شناسایی جدایه­ها بر اساس خصوصیات ریخت­شناسی و توالی ناحیه ITS و TUB انجام گرفت. در این پژوهش، در مجموع 39 جدایه قارچی متعلق به 12 گونه از 6 جنس به­دست آمد که بیشترین تعداد جدایه متعلق به غلاف گیاه برنج با فراوانی 16 جدایه و کمترین آن 11 جدایه به ریشه گیاه برنج تعلق داشت. از بین 6 جنس شناسایی شده در این تحقیق، بیشترین تعداد جدایه متعلق به جنس­های Alternaria و Fusarium، به‌ترتیب هر یک با فراوانی 19 و 6 جدایه، و همچنین بیشترین تعداد گونه حاصل به جنس Fusarium با فراوانی شش گونه تعلق داشت. گونه­های قارچی شامل:  Fusarium chlamydosporum،F. proliferatum ، F. incarnatum، F. solani، F. fujikuroi، F. verticillioides، Sarocladium bactrocephalum، Nigrospora sphaerica، N. oryzae، Alternaria alternata، Epicoccum nigrum، Cladosporium cladosporioides هستند. در این میان گونه­های Nigrospora sphaerica، Epicoccum nigrum و Sarocladium bactrocephalum برای اولین­بار در این مطالعه به­عنوان اندوفیت گیاه برنج در ایران گزارش می­شوند.

کلیدواژه‌ها

موضوعات

عنوان مقاله [English]

Identification of Fungal Endophytes in Aerobic Rice Cultivation of Golestan Province

نویسندگان [English]

  • Z. Zare 1
  • S.E. Razavi 2
  • K. Rahnama 2
  • A. Taheri 2
  • J. Gherekhloo 3
1 Department of plant protection. faculty of plant production. Gorgan University of Agricultural sciences and Natural Resources
2 Department of Plant Protection, Plant Production Faculty, Gorgan University of Agricultural Sciences and Natural Resources, Iran
3 Department of Agriculture, Faculty of plant production, Gorgan University of Agricultural sciences and Natural Resources, Gorgan
چکیده [English]

Introduction
Rice is one of the most important grains in the world and large part of its consumption is related to Middle East. Iran ranks 23rd in production, 26th in cultivation, and 13th in rice consumption with nearly 560,000-hectare rice lands and an annual production of nearly 2.5 million tons. In natural and agricultural ecosystems, plants are associated with large populations of microorganisms in the rhizosphere, endosphere (inside the root), leaves (phyllosphere), as well as pollen and seeds. These microorganisms spend all or part of their life cycle inside the healthy tissues of host plant without any obvious sign of disease. The stable coexistence between endophytes and plants is due to production of biological secondary metabolites with a unique structure, which positively affects the survival of plant growth in adverse conditions. Studies have shown that endophytic fungi increase resistance to drought, insects, diseases and host tolerance level, especially in stress conditions. Considering the increase in the risk of dehydration and the lack of sufficient water resources and the development and adoption of aerobic rice cultivation, it is important to identify endophytic fungi that increase tolerance to drought stress and improve the growth factors of the host plant. There are relatively few studies on endophytic fungi that colonize healthy tissues of rice plants under aerobic conditions. Therefore, the aim of this study is to evaluate endophytic fungal collections in different parts of aerobically cultivated rice plants.
 
Materials and Methods
In this study, in order to identify endophytic fungi in aerobic cultivation of rice in Golestan province, sampling were carried out from several aerobic rice fields in Golestan province, including Gorgan, Kordkuy, Bandar Gaz, Aliabad and Kalaleh cities during the months of May to September in 2019-2020. Plant tissues were transferred to the laboratory (Department of Plant Protection) in separate paper bags along with specifications. Purification was performed on a 2% water-agar medium using Hyphal tip method. Based on investigations and morphological data, a number of isolates were selected as representatives for molecular identification. Genomic DNA was extracted from mycelia grown in PDB (Potato Dextrose Broth) culture medium according to the CTAB method with a slight modification, and the polymerase chain reaction (PCR) was performed according to Nemati and Abdulhazadeh method. The ITS region (ITS4-5.8S-ITS5) of the selected isolates was amplified during the PCR reaction using ITS5 (5'-GGAAGTAAAAGTCGTAACAAGG-3') and ITS4 (5'- TCCTCCGCTTATTGATATGC-3') primers and The β-tubulin region was amplified by T1 (5'-AACATGCGTGAGATTGTAAGT-3') and β Sandy-R (5'-GCRCGNGGVACRTACTTGTT-3') primers. Purification and sequencing of PCR products was done by Pishgam Biotechnology Company. The sequences of ITS and β-tubulin regions were modified and extracted using Chromas 2.6.6 software. Nucleotide sequences of the selected isolates were compared with the available sequences in the Gene bank by Blast algorithm and the sequences data were submitted in the NCBI database website. Phylogenetic trees were drawn using the Maximum likelihood method and MEGA 6.06 software.
 
Results and Discussion
Based on preliminary morphological studies, 20 isolates were selected as representatives of purified isolates and were identified by molecular analysis. In this study, a total of 39 fungal isolates were obtained, that the highest number of the isolates belonged to the pods of rice plants with the frequency of 17 isolates and the lowest number of isolates belonged to the roots of rice plants with 10 isolates. Among the 6 genera identified in this research, the largest number of isolates was belonged to Alternaria and Fusarium genera, with 18 and 6 isolates, respectively, and the highest number of species was belonged to Fusarium genus with 6 species. Fungal species include: F. chlamydosporum, F. proliferatum, F. incarnatum, F. solani, F. fujikuroi, F. verticillioides, S. bactrocephalum, N. sphaerica, N. oryzae, A. alternata, E. nigrum, C. cladosporioides. Among these species, Nigrospora sphaerica, E. nigrum and S. bactrocephalum are reported for the first time in this study as rice plant endophytes in Iran.
 
Conclusion
Investigation of endophytic fungi in the present study indicates that some of these isolates, including F. chlamydosporum, F. proliferatum, F. fujikuroi are pathogenic fungi. It means that range of the endophytic life and pathogenic life cannot be separated and determined and environmental conditions and nutritional status determine the type of interaction. F. chlamydosporum causes Fusarium root and crown rot, is reported in some sources as an endophytic fungus in various plants.
 

کلیدواژه‌ها [English]

  • Fusarium
  • Morphology
  • Molecular
  • Rice endophytic
  • Sarocladium bactrocephalum

مراجع

  1. Arnold, E.A. (2007). Understanding the diversity of foliar endophytic fungi: progress, challenges, and frontiers, Fungal Biology Reviews, 21(2-3), 51–66. https://doi.org/10. 1016/j.fbr.2007.05.003
  2. Arnold, A.E., & Herre, E.A. (2003). Canopy cover and leaf age affect colonization by tropical fungal endophytes: ecological pattern and process in Theobroma cacao (Malvaceae). Mycologia, 95, 388-398. https://doi.org/10.1080/15572536.2004.1183 3083
  3. Bacon, C.W., Glenn, A.E., & Yates, IE. (2008) .Fusarium verticillioides: managing the endophytic association with maize for reduced fumonisins accumulation. Toxin Rev, 27, 411–446. https://doi.org/10.1080/15569540802497889
  4. Barnett, H.L., & Hunter, B.B. (1998). Illustrated genera of imperfect fungi. Vol. Ed. 4. American Phytopathological Society (APS Press).
  5. Cao, L.X., You, J.L., & Zhou, S.N. (2002). Endophytic fungi from Musa acuminate leaves and roots in South China. World Journal of Microbiology and Biotechnology, 18(2), 169–171. https://doi.org/10.1023/A:1014491528811
  6. Chen, X.M., Dong, H.L., Hu, K.H., Sun, Z.R., Chen, J., & Guo, S.X. 2010. Diversity and antimicrobial and plant-growth-promoting activities of endophytic fungi in Dendrobium loddigesii Journal of Plant Growth Regulator, 29, 328–337. https://doi.org/10.1007/s00344-010-9139-y.
  7. Coombs, J.T., & Franco, C.M.M. (2003). Isolation and identification of actinobacteria from surface-sterilized wheat roots. Apply Environment Microbiology, 69, 5603–5608. http://dx.doi.org/10.1128/AEM.69.9.5603-5608.
  8. Debbab, A., Aly, A.H., & Proksch, P. (2011). Bioactive secondary metabolites from endophytes and associated marine derived fungi. Fungal Diversity, 49(1), 1-12. https://doi.org/10.1007/s13225-011-0114-0
  9. Domsch, K.H., Gams, W., & Anderson, T.H. (1980). Compendium of soil fungi. Volume 1. Academic Press (London) Ltd. 860 PP. https://doi.org/10.1111/j.1365-2389.010521.x
  10. Ellis, M.B. (1971). Dematiaceous Hyphomycetes. CMI, Kew, England, 608 pp.
  11. Estrada, A.E.R., Jonkers, W., Kistler, H.C., & May, G. (2012). Interactions between Fusarium verticillioides, Ustilago maydis, and Zea mays: an endophyte, a pathogen, and their shared plant host. Fungal Genetic Biology, 49, 578–587. http://dx.doi.org/10.1016/j.fgb.2012.05.001.
  12. Faeth, S.H., & Fagan, W.F. (2002). Fungal endophytes: Common host plant symbionts but uncommon mutualists, Integrative and Comparative Biology, 42(2), 360–368. https://doi.org/10.1093/icb/42.2.360
  13. Fisher, P.J., & Petrini, O. (1992). Fungal saprobes and pathogens as endophytes of rice (Oryza sativa), New Phytologist, 120(1), 137–143. https://doi.org/10.1111/j.1469-8137.1992.tb01066.x
  14. Ganley, R.J., Brunsfeld, S.J., & Newcombe, G. (2004). A community of unknown endophytic fungi in western white pine. Proceedings of the National Academy of Sciences of the United States of America. 101, 10107–10112. https://doi.org/10.1073/pnas.0401513101.
  15. Ganjali, R., Sharif Nabi, B., & Mirlouhi, A.F. (2004). Classical methods and specific primers in detection of endophytic fungi in some gramineous plants. Rostaniha, 5(1), 37-51. (In Persian)
  16. Hamilton, C.E., James, D.B., Labbe, J., & Yang,X. (2016). Mitigating climate change through managing constructed microbial communities in agriculture. Agriculture, Ecosystem & Environment, 216, 304–308. https://doi.org/10.1016/j.agee.2015.10.006
  17. Hatamzadeh, S., & Rahnama, K. (2016). Isolation and identification of some endophytic fungi of forest trees. Extension of Plant Protection, 10(13,14), 16-22. (In Persian with English abstract). www. ppext.ir.
  18. Hatamzadeh, S., Rahnama, K., White, J.F., Akbari Oghaz, N., Nasrollahnejad, S., & Hemati, KH. (2022). Investigation of some endophytic fungi from five medicinal plants with growth promoting ability on maize (Zea mays). Journal of Applied Microbiology, 0, 1–15.
  19. Hardoim, P.R., Van Overbeek, L.S., Berg, G., & et al. (2015). The hidden world within plants: ecological and evolutionary considerations for defining functioning of microbial endophytes. Microbiology and Molecular Biology Reviews, 79, 293–320. https://doi: 10.1128/MMBR.00050-14.
  20. Islam, S., Akanda, A.M., Prova, A., & et al. (2014). Growth promotion effect of Fusarium PPF1 from bermudagrass (Cynodon dactylon) rhizosphere on Indian spinach (Basella alba) seedlings are linked to root colonisation. Archiv Phytopathology Plant Protection, 47, 2319–31. https://doi.org/10.1080/03235408.2013.876745
  21. Jiménez-Gasco, M.M., Malcolm, G.M., Berbegal, M., Armengol, J., & Jiménez Díaz, R.M. (2014). Complex molecular relationship between vegetativecompatibility groups (VCGs) in Verticillium dahliae: VCGs do not always align with clonal lineages. Phytopathology, 104, 650–659. http://dx.doi.org/10.1094/PHYTO-07-13-0180-R.
  22. Jonbozorgi, S., Mehrabi-Koushki, M., & Farokhinejad, R. (2019). Isolation and Identification of Fungal Endophytes of the Cowpea in Khuzestan Province. Biological Journal of Microorganism, 8(29), 97-115. (In Persian)
  23. Köberl, M., Schmidt, R., Ramadan, E.M, Bauer, R., & Berg, G. (2013). The microbiome of medicinal plants: diversity and importance for plant growth, quality and health. Front Microbiology, l4, 400. http://dx.doi.org/10.3389/fmicb.2013.00400.
  24. Larran, S., Perello, A., Simon, M.R., & Moreno, V. (2007). The endophytic fungi from wheat (Triticium aestivum L). World Journal of Microbiology and Biotechnology, 23(4), 565–572. https://doi.org/10.1007/s11274-006-9266-6
  25. Leslie, J.F., & Summerell, B.A. (2006). The Fusarium Laboratory Manual. Blackwell Publishing. First Edition. 388pp. https://doi.org/10.1002/9780470278376
  26. Liu, H., Carvalhais, L.C., Crawford, M., Singh, E., Dennis, P.G., Pieterse, C.M., & Schenk, P.M. (2017). Inner plant values: diversity, colonization and benefits from endophytic bacteria. Frontiers in Microbiology 8, 2552. https://doi.org/10.3389/ fmicb.2017.02552
  27. Liu, H., Brettell, L.E., Qiu, Z., & Singh, B.K. (2020a). Microbiome-mediated stress resistance in plants. Trends in Plant Science, 25, 733–743. https://doi.org/1016/j .tplants.2020.03.014
  28. Moricca, S., Ginetti, B., & Ragazzi, A. (2012). Species- and organ-specificity in endophytes colonizing healthy and declining Mediterranean oaks. Phytopathologia Mediterranea, 51, 587–598. https://doi.org/10.14601/PhytopatholMediterr-11705
  29. Nair, D.N., & Padmavathy, S. (2014). Impact of endophytic microorganisms on plants, environment and humans. The Scientific World Journal, Article ID 250693, 11 pages. https://doi.org/10.1155/2014/250693
  30. Nemati, H., & Abdollahzadeh, A. (2009). Sweet and sourcherry- Production and Utilization. Jehad University of Mashhad Press, Iran. (In Persian)
  31. Porras-Alfaro, A., & Bayman, P. (2011). Emergent properties: endophytes and microbiomes, Annual Review of Phytopathology, 49, 291–315. https://doi.org/ 1146/annurev-phyto-080508-081831
  32. O’Donnell, , & Cigelnik, E. (1997). Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus Fusarium are Nonorthologous. Molecular Phylogenetics and Evolution, 7, 103-107. https://doi.org/10.1006/ mpev.1996.0376.
  33. Rao, A.N., Johnson, D.E., Sivaprasad, B., Ladha, J.K., & Mortimer, A.M. (2007). Weed management in direct-seeded rice. Advances in Agronomy, 93, 153–255. https://doi.org/10.1016/S0065-2113(06)93004-1
  34. Rodriguez, R.J., Redman, R.S., & Henson, J.M. (2005). The role of fungal symbioses in the adaptation of plants to high stress environments. Mitigation and Adaptation Strategies for Global Change, 9, 261–272. https://doi.org/10.1023/b:miti.0000029922. 31110.97
  35. Rodriguez, R., White, J., Arnold, A., & Redman, R. (2009). Fungal endophytes: diversity and functional roles. New Phytologist, 182(2), 314-330. https://doi.org/10.1111/j.1469-8137.2009.02773.x
  36. Solla, A., & Gil, L. (2012). Evaluating Verticillium dahliae for biological control of Ophiostoma novo-ulmiin Ulmus minor. Plant Pathology, 52, 579–585. http://dx.doi.org/10. 1046/j.1365-3059.2003. 00921.x
  37. Santamaria, J., & Bayman, P. (2005). Fungal epiphytes and endophytes of coffee leaves (Coffee arabica). Microbial Ecology, 50(1), 1–8. https://doi.org/1007/s00248-004-0002-1
  38. Schulz, B., & Boyle, C. (2005). Review: The endophytic continuum. Mycological Research, 109, 661-686. https://doi.org/10.1017/S095375620500273X
  39. Sheteiwy, M.S., Ali, D.F., Xiong, Y.C., Brestic, M., Skalicky, M., Hamoud, Y.A., Ulhassan, Z., Shaghaleh, H., AbdElgawad, H., Farooq, M., & Sharma, A. (2021b). Physiological and biochemical responses of soybean plants inoculated with arbuscular mycorrhizal fungi and Bradyrhizobium under drought stress. BMC Plant Biology, 21, 195. https://doi.org/1186/s12870-021-02949-z
  40. Sieber, T.N. (2007). Endophytic fungi in forest trees: are they mutualists? Fungal Biology Reviews, 21(2), 75-89. https:// doi:10.1016/j.fbr.2007.05.004
  41. Staniek, A., Woerdenbag, H.J., & Kayser, O. (2008). Endophytes: exploiting biodiversity for the improvement of natural product-based drug discovery. Journal Plant Interaction, 3, 75–93. https://doi.org/10.1080/17429140801886293
  42. Tamura, K., Stecher, G., Peterson, D., Filipski, A., & Kumar, S. (2013). MEGA6: Molecular evolutionary genetics analysis version 6.0. Molecular Biology and Evolutionment, 30, 2725-2729. https://doi.org/10.1093/molbev/mst197
  43. Tan, R.X., & Zou, W.X. (2001). Endophytes: a rich source of functional metabolites. Natural Product Reports, 18, 448-459. https://doi.org/10.1039/b100918o
  44. Tintjer, T., & Rudgers, J.A. (2006). Grass-Herbivore interactions altered by strains of a native endophyte. New Phytologist, 170, 513-521. https://doi.org/10.1111/j.1469-8137.2006.01720.x
  45. Weber, D. (2009). Endophytic fungi, occurrence and metabolites. In: Anke, T. & Weber, D. (eds). The Mycota. Vol. XV, Physiology and Genetics selected basis and applied aspects springer Ver lag, Berlin, Germany, pp. 153–195. https://doi.org/1007/978-3-642-00286-18
  46. White, T.J., Bruns, T., Lee, S., & Taylor, J. (1990). Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Inis, M.A., Gelfand, D.H., Sninsky, J.J. & White, T.J.(eds). PCR Protocols: A guide to Methods and Applications. Academic Press, San Diego, USA, pp. 315−322. http://dx.doi.org/10.1016/B978-0-12-372180-8.50042-1
  47. Vallino, M., Greppi, D., Novero, M., Bonfant, P., & Luppoto, E. (2009). Rice root colonization by mycorrhizal and endophytic fungi in aerobic soil. Annals of Applied Biology, 154, 195-204. https://doi.org/10.1111/j.1744-7348.2008.00286.x

 

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