Document Type : Research Paper - Agronomy


1 Ph.D. Student of Soil Science, Department of Soil Science, Faculty Agriculture , Marvdasht Branch, Islamic Azad University, Marvdasht, Iran

2 Professor, Department of Soil Science, Faculty Agriculture, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran

3 Assistant Professor, Department of Biology, Marvdasht Branch, Islamic Azad University, Marvdasht, Iran

4 Assistant Professor, Department of Microbiology, Larestan Branch, Islamic Azad University, Larestan, Iran


Agriculture practices involve the extensive utilization of chemical fertilizers that lead to several impacts, namely groundwater pollution, eutrophication, and production of greenhouse gases. These ultimately lead to environmental pollution and pose several health risks. With this respect, plant growth promoting rhizobacteria (PGPR) consist of a diverse collection of rhizobacteria that occur in the rhizospheric zone and interact with plant roots. Moreover plant growth promoting rhizobacteria (PGPR) are a heterogeneous group of bacteria found in the rhizosphere, associated with roots and root surfaces or living freely in the soil and influencing plant growth directly or indirectly. A large number of bacteria is reported to contribute to plant growth, including species of Pseudomonas, Azospirillum, Azotobacter, Klebsiella, Enterobacter, Acinetobacter, Alcaligenes, Arthrobacter, Burkholderia, Bacillus, and Serratia. Additionally, several reports indicate the application of ACC-deaminase-producing PGPR in facilitating plant growth and inducing tolerance in various abiotic stresses, namely (1) salt stress, (2) drought stress, (3) heat stress, (4) chilling stress, etc. The present study was performed to screen PGPR strains with different PGP traits in calcareous soils under salinity condition and to use such PGPR strains as biological inoculation.
Materials and Methods
In this study, 45 rhizospheric soil samples were collected from different crops in agricultural farms and placed in plastic bags, and transported to a laboratory, where it was refrigerated at 4 ° C until further use. To obtain a solution of 10-4 to 10-6 dilutions, one milliliter of suspension was serially diluted and 100 microliters of this suspension solution were spread onto the sterile nutrient agar medium. plates were incubated at 28±2°C for 48 hours in a BOD incubator. Rhizobacterial isolates with a different morphological appearance and fast growth on agar plates were randomly selected and purified by repetitive streaking on the fresh medium. After purification, these PGPR isolates were screened for PGP traits. Isolates with phosphate solubilization, siderophore production, IAA production and ACC-deaminase production were screened and finally three isolates were selected for molecular identification and pot experiments.
Results and Discussion
In this research, a total of 45 rhizospheric soil samples were collected from the rhizosphere of different plants. 181 isolates were purified and isolated. In the initial screening, some of them were characterized by phosphate solubilization, siderophore production and indole acetic acid (IAA) production. In the secondary screening, superior isolates were selected in these three growth-promoting traits, and in the ACC-deaminase production test, all three isolates were able to produce this enzyme. Molecular identification of the isolates was performed by 16S rRNA gene sequencing technique and the isolates belonged to Bacillus Cereus, Pseudomonas Syringae and Pseudomonas Alcaliphila which were registered in the NCBI database. In greenhouse experiments, inoculation of Pseudomonas Alcaliphila showed the best performance shoot length, shoot dry weight, chlorophyll production, crude protein production of alfalfa (medicago sativa L.) and lowest proline production under salinity stress, while inoculation of Pseudomonas Syringae had better yield at root length and root dry weight.
Extremely cost-effective methods such as applications of microorganisms with PGPR activities can enhance plant growth, speed up seed germination, improve seedling emergence, and protect plants. In conclusion, the applied plant-growth-promoting rhizobacteria (PGPR) bacterial SF1050 strain (Pseudomonas Alcaliphila) proved to be a potential candidate for improving alfalfa crops in salinity conditions. Nevertheless, it is suggested to validate the current results by conducting field trials.


Main Subjects

Acevedo, E., Galindo-Castañeda, T., Prada, F., Navia, M., & Romero, H. M. (2014). Phosphate-solubilizing microorganisms associated with the rhizosphere of oil palm (Elaeis guineensis Jacq.) in Colombia. Applied Soil Ecology, 80, 26-33.
Ahemad, M., & Kibret, M. (2014). Mechanisms and applications of plant growth promoting rhizobacteria: current perspective. Journal of King saud University-Science, 26(1), 1-20.
Ahmad, F., Ahmad, I., & Khan, M. (2008). Screening of free-living rhizospheric bacteria for their multiple plant growth promoting activities. Microbiological Research, 163(2), 173-181.
Alexander, D., & Zuberer, D. (1991). Use of chrome azurol S reagents to evaluate siderophore production by rhizosphere bacteria. Biology and Fertility of Soils, 12, 39-45.
Bates, L. S., Waldren, R. P., & Teare, I. (1973). Rapid determination of free proline for water-stress studies. Plant and Soil, 39(1), 205-207.
Cavite, H. J. M., Mactal, A. G., Evangelista, E. V., & Cruz, J. A. (2021a). Biochemical characteristics and inoculation effects of multi-trait plant growth-promoting rhizobacteria on upland rice (Oryza sativa L. cv PSB Rc23) seedling growth. Archives of Microbiology, 203(6), 3533-3540.
Cavite, H. J. M., Mactal, A. G., Evangelista, E. V., & Cruz, J. A. (2021b). Growth and Yield Response of Upland Rice to Application of Plant Growth-Promoting Rhizobacteria. Journal of Plant Growth Regulation, 40(2), 494-508.
Egamberdieva, D., Wirth, S., Bellingrath-Kimura, S. D., Mishra, J., & Arora, N. K. (2019). Salt-tolerant plant growth promoting rhizobacteria for enhancing crop productivity of saline soils. Frontiers in Microbiology, 18, 2791.
Gordon, S. A., & Weber, R. P. (1951). Colorimetric estimation of indoleacetic acid. Plant Physiology, 26(1), 192-195.
Gupta, A., Bano, A., Rai, S., Kumar, M., Ali, J., Sharma, S., & Pathak, N. (2021). ACC deaminase producing plant growth promoting rhizobacteria enhance salinity stress tolerance in Pisum sativum. 3 Biotech, 11(12), 514.
Gyaneshwar, P., Kumar, G. N., Parekh, L., & Poole, P. (2002). Role of soil microorganisms in improving P nutrition of plants. Plant and Soil, 245(1), 83-93.
Halimursyadah, H., & Rizva, D. (2022). Exploration, isolation and characterization of indigenous rhizobacteria from patchouli rhizosphere as PGPR candidates in producing IAA and solubilizing phosphate. Paper presented at the IOP Conference Series, Earth and Environmental Science.
Indiragandhi, P., Anandham, R., Madhaiyan, M., Kim, G. H., & Sa, T. (2008). Cross-utilization and expression of outer membrane receptor proteins for siderophore uptake by Diamondback moth Plutella xylostella (Lepidoptera: Plutellidae) gut bacteria. FEMS Microbiology Letters, 289(1), 27-33.
Jeon, J. S., Lee, S. S., Kim, H. Y., Ahn, T. S., & Song, H. G. (2003). Plant growth promotion in soil by some inoculated microorganisms. Journal of Microbiology, 41(4), 271-276.
Karami, S., Yasrebi, J., Safarzadeh Shirazi, S., Whalen, J. K., Ronaghi, A., & Ghasemi-Fasaei, R. (2020). Sugar processing residuals as an iron source for grain crops grown in calcareous soil. Communications in Soil Science and Plant Analysis, 51(1), 60-69.
Khan, A., & Singh, A. V. (2021). Multifarious effect of ACC deaminase and EPS producing Pseudomonas sp. and Serratia marcescens to augment drought stress tolerance and nutrient status of wheat. World Journal of Microbiology and Biotechnology, 37(12), 198.
Liang, J. L., Liu, J., Jia, P., Yang, T., Zeng, Q. W., Zhang, S. C., … & Li, J.T. (2020). Novel phosphate-solubilizing bacteria enhance soil phosphorus cycling following ecological restoration of land degraded by mining. The ISME Journal, 14(6), 1600-1613.
Lichtenthaler, H. K. (1987). Chlorophylls and carotenoids: Pigments of photosynthetic biomembranes. Methods in Enzymology, 148, 350-382
Murali, M., Gowtham, H., Singh, S. B., Shilpa, N., Aiyaz, M., Niranjana, S., & Amruthesh, K. (2021). Bio-prospecting of ACC deaminase producing Rhizobacteria towards sustainable agriculture: A special emphasis on abiotic stress in plants. Applied Soil Ecology, 168, 104142.
Nekoyanfar, Z., Lack, Sh., & Abadouz, Gh. R. (2017). Assessment effect of cutting time and soil salinity on quality and quantity forage yield of five alfalfa (Medicago sativa L.) varieties under Ahvaz conditions. Plant Productions, 40(3), 113-127. [In Persian]
Pandey, S., & Gupta, S. (2020). Diversity analysis of ACC deaminase producing bacteria associated with rhizosphere of coconut tree (Cocos nucifera L.) grown in Lakshadweep islands of India and their ability to promote plant growth under saline conditions. Journal of Biotechnology, 324, 183-197.
Rodrı́guez, H., & Fraga, R. (1999). Phosphate solubilizing bacteria and their role in plant growth promotion. Biotechnology Advances, 17(4-5), 319-339.
Shrivastava, U. P., & Kumar, A. (2013). Characterization and optimization of 1-aminocyclopropane-1-carboxylate deaminase (ACCD) activity in different rhizospheric PGPR along with Microbacterium sp. strain ECI-12A. International Journal of Applied Sciences and Biotechnology, 1(1), 11-15.
Singh, A., Sharma, J., Paichha, M., & Chakrabarti, R. (2020). Achyranthes aspera (prickly chaff flower) leaves- and seeds-supplemented diets regulate growth, innate immunity, and oxidative stress in Aeromonas hydrophila-challenged Labeo rohita. Journal of Applied Aquaculture, 32(3), 250-267.
Tirry, N., Kouchou, A., Laghmari, G., Lemjereb, M., Hnadi, H., Amrani, K., … & El Ghachtouli, N. (2021). Improved salinity tolerance of Medicago sativa and soil enzyme activities by PGPR. Biocatalysis and Agricultural Biotechnology, 31, 101914.
Upadhyay, S. K., Saxena, A. K., Singh, J. S., & Singh, D. P. (2019). Impact of native ST-PGPR (Bacillus pumilus; EU927414) on PGP traits, antioxidants activities, wheat plant growth and yield under salinity. Climate Change and Environmental Sustainability, 7(2), 157-168.
Vazquez, P., Holguin, G., Puente, M., Lopez-Cortes, A., & Bashan, Y. (2000). Phosphate-solubilizing microorganisms associated with the rhizosphere of mangroves in a semiarid coastal lagoon. Biology and Fertility of Soils, 30(5-6), 460-468.