Plant Protection (Scientific Journal of Agriculture)

Plant Protection (Scientific Journal of Agriculture)

Efficacy of effective microorganisms, humic Acid, and Peribacillus frigoritolerans on reducing bacterial leaf blight of rice under field conditions

Document Type : Research paper-Persian

Authors
1 M.Sc. graduate of Plant Pathology, Department of Plant Protection, Faculty of Agriculture, Yasouj University, Iran
2 Associate Professor, Department of Plant Protection, Faculty of Agriculture, Yasouj University, Iran
Abstract
Background and Objective
Bacterial leaf blight of rice, caused by Xanthomonas oryzae pv. oryzae (Xoo), is one of the most destructive and economically significant diseases of this globally strategic crop. The disease is widely distributed in major rice-growing regions and can cause severe yield losses, especially under warm and humid environmental conditions that favor rapid pathogen multiplication and spread. Given the essential role of rice in global food security, developing sustainable and effective management strategies for bacterial leaf blight remains a key priority in modern plant protection. Even though chemical control measures have traditionally been used to lower disease incidence, their repeated application is associated with limitations such as inconsistent efficacy, emergence of resistant pathogen strains, environmental contamination, and negative impacts on non-target beneficial microorganisms. Thus, increasing attention has been directed toward alternative approaches that boost plant defense capacity and promote biological suppression of pathogens. In this context, the present study aimed to ascertain the efficacy of natural growth stimulants and biological agents in the management of bacterial leaf blight under field conditions.
Materials and Methods
In this study, the effects of various treatments, including humic acid, effective microorganisms (EM), and the antagonistic bacterium Peribacillus frigoritolerans, were appraised on four rice cultivars (Fajr, Hashemi, Setayesh, and Shirudi) under natural field conditions. These cultivars represent commonly cultivated varieties with different levels of susceptibility and agronomic characteristics. The experiment was undertaken as a factorial arrangement based on a randomized complete block design with four replications to ensure reliable statistical comparison of treatment effects. The treatments were applied ten days after transplanting the seedlings to allow proper establishment of the plants. The pathogenic bacterium was inoculated seven days after treatment application to ascertain the preventive and protective effects of the applied treatments against subsequent pathogen challenge. The population dynamics of the bacterium within leaf tissues were measured on days 1, 5, 10, 15, and 20 post-inoculation to monitor bacterial colonization, multiplication, and persistence over time. Disease severity was assessed 30 days following inoculation based on the mean length of necrotic lesions on infected leaves, providing a quantitative indicator of disease development as well as host response.
Results
The results revealed that treatment efficacy varied depending on both cultivar and assessment time. The combined treatment of humic acid and EM consistently and significantly (p≤0.01) diminished the Xoo population and necrotic lesion length compared to the pathogen-inoculated control in the Fajr, Hashemi, and Setayesh cultivars. This reduction was sustained across multiple sampling intervals, suggesting a stable suppressive influence on bacterial growth within plant tissues. The combined application was therefore identified as the most effective treatment strategy among those appraised in this study. Humic acid treatment alone also exhibited significant efficacy in the Fajr and Hashemi cultivars, reflecting its potential role in stimulating plant growth, improving nutrient uptake, and enhancing defense-related physiological responses. EM treatment alone presented an inhibitory effect only at specific time points and primarily in the Setayesh cultivar, suggesting that its performance may depend on the host genotype or environmental conditions. The P. frigoritolerans treatment failed to control the disease significantly in any of the cultivars under the conditions of this experiment, as no meaningful reduction was observed in bacterial population or lesion length when compared to the inoculated control.
Discussion
The integrated application of humic acid and EM, as a sustainable and environmentally friendly strategy, presents high potential for managing bacterial leaf blight in susceptible rice cultivars. The enhanced effectiveness of the combined treatment may be attributed to ameliorated plant vigor, stimulation of host defense mechanisms, as well as favorable modulation of the rhizosphere microbial community, which together may limit pathogen establishment and disease progression under field conditions. Further, the synergistic interaction between these components may boost physiological resilience and improve overall plant performance during pathogen stress. This approach, by enhancing overall plant and rhizosphere health, can serve as a suitable alternative for lowering reliance on chemical methods in rice disease management programs and may contribute to more resilient as well as sustainable production systems in intensive rice-growing regions.
Keywords
Subjects

Akbari, M., Rezaei, R., & Charehgani, H. (2019). Evaluation of resistance in some rice cultivars to bacterial blight disease caused by Xanthomonas oryzae pv. oryzae. Plant Protection (Scientific Journal of Agriculture), 42(3), 59-70. https://doi.org/10.22055/ppr.2019.15095
Asli, S., & Neumann, P. M. (2010). Rhizosphere humic acid interacts with root cell walls to reduce hydraulic conductivity and plant development. Plant and Soil, 336, 313–322. https://doi.org/10.1007/s11104-010-0483-2
Bajwa, R., Javaid, A., & Tasneem, Z. (1995). Response of indigenous soil microflora to EM inoculation in Pakistan. Biological Science, 1, 73–79.
Bardin, M., Ajouz, S., Comby, M., Lopez-Ferber, M., Graillot, B., Siegwart, M., & Nicot, P. C. (2015). Is the efficacy of biological control against plant diseases likely to be more durable than that of chemical pesticides? Frontiers in Plant Science, 6, 566. https://doi.org/10.3389/fpls.2015.00566
Bonaterra, A., Badosa, E., Daranas, N., Francés, J., Roselló, G., & Montesinos, E. (2022). Bacteria as biological control agents of plant diseases. Microorganisms, 10(9), 1759. https://doi.org/10.3390/microorganisms10091759
Boonsrangsom, T., Boondech, A., Chansongkram, W., Suachaowna, N., Buddhachat, K., Rungrat, T., Jumpathong, J., Pongcharoen, P., Inthima, P., Aeksiri, N., Ratanasut, K., & Sujipuli, K. (2025). Molecular characterization and pathogenicity of Xanthomonas oryzae pv. oryzae isolates from lower northern Thailand, the causal agent of rice bacterial blight. Physiological and Molecular Plant Pathology, 136, 102550. https://doi.org/10.1016/j.pmpp.2024.102550
Buddhachat, K., Ritbamrung, O., Sripairoj, N., Inthima, P., Ratanasut, K., Boonsrangsom, T., & Sujipuli, K. (2021). One-step colorimetric LAMP (cLAMP) assay for visual detection of Xanthomonas oryzae pv. oryzae in rice. Crop Protection, 150, 105809. https://doi.org/10.1016/j.cropro.2021.105809
Charehgani, H. (2020). Effect of wood vinegar, humic acid and Effective Microorganisms against Meloidogyne javanica on tomato. Plant Pathology Science, 9(2), 73–84.
Civáň, P., & Brown, T. A. (2018). Role of genetic introgression during the evolution of cultivated rice (Oryza sativa L.). BMC Evolutionary Biology, 18, 57. https://doi.org/10.1186/s12862-018-1180-7
Daiss, N., Lobo, M. G., Socorro, A. R., Brückner, U., Heller, J., & Gonzalez, M. (2008). The effect of three organic pre-harvest treatments on Swiss chard (Beta vulgaris L. var. cycla L.) quality. European Food Research and Technology, 226, 345–353. https://doi.org/10.1007/s00217-006-0543-2
Dungait, J. A., Cardenas, L. M., Blackwell, M. S., Wu, L., Withers, P. J., Chadwick, D. R., & Whitmore, A. P. (2012). Advances in the understanding of nutrient dynamics and management in UK agriculture. Science of the Total Environment, 434, 39–50. https://doi.org/10.1016/j.scitotenv.2012.06.004
Filippi, M. C. C., Da Silva, G. B., Silva-Lobo, V. L., Côrtes, M. V. C. B., Moraes, A. J. G., & Prabhu, A. S. (2011). Leaf blast (Magnaporthe oryzae) suppression and growth promotion by rhizobacteria on aerobic rice in Brazil. Biological Control, 58(2), 160–166. https://doi.org/10.1016/j.biocontrol.2011.04.006
Gnanamanickam, S. S. (2009). An overview of progress in biological control. In Biological Control of Rice Diseases (pp. 43–51). Springer.
Higa, T. (2000). What is EM technology. EM World Journal, 1, 1–6.
Islam, T., Haque, M. A., Barai, H. R., Istiaq, A., & Kim, J. J. (2024). Antibiotic resistance in plant pathogenic bacteria: Recent data and environmental impact of unchecked use and the potential of biocontrol agents as an eco-friendly alternative. Plants, 13(8), 1135. https://doi.org/10.3390/plants13081135
Kawaharada, Y., Kelly, S., Nielsen, M. W., Hjuler, C. T., Gysel, K., Muszyński, A., & Stougaard, J. (2015). Receptor-mediated exopolysaccharide perception controls bacterial infection. Nature, 523, 308–312. https://doi.org/10.1038/nature14611
Kumar, A., Gul, M. Z., Zeeshan, A., Bimolata, W., Qureshi, I. A., & Ghazi, I. A. (2013). Differential antioxidative responses of three different rice genotypes during bacterial blight infection. Australian Journal of Crop Science, 7(12), 1893–1900.
Li, J., Wei, J., Shao, X., Yan, X., & Liu, K. (2024). Effective microorganisms input efficiently improves the vegetation and microbial community of degraded alpine grassland. Frontiers in Microbiology, 14, 1330149. https://doi.org/10.3389/fmicb.2023.1330149
Li, M., Shafique, M. S., Zhou, H., Wang, J., Liu, Y., Wang, C., Wang, Z., & Ji, Z. (2025). A new bacterial blight resistance gene Xa50(t) in the Xa4 locus confers resistance against Xanthomonas oryzae pv. oryzae in rice. Frontiers in Plant Science, 16, 1657476. https://doi.org/10.3389/fpls.2025.1657476
McManus, P. S., Stockwell, V. O., Sundin, G. W., & Jones, A. L. (2002). Antibiotic use in plant agriculture. Annual Review of Phytopathology, 40(1), 443–465. https://doi.org/10.1146/annurev.phyto.40.120301.093927
Moghadam, H. R. T. (2015). Humic acid as an ecological pathway to protect corn plants against oxidative stress. Biological Forum – An International Journal, 7(1), 46–53.
Nino-Liu, D. O., Ronald, P. C., & Bogdanove, A. J. (2006). Xanthomonas oryzae pathovars: Model pathogens of a model crop. Molecular Plant Pathology, 7(5), 303–324. https://doi.org/10.1111/j.1364-3703.2006.00344.x
Qiu, C., Bao, Y., Lu, D., Yan, M., Li, G., Liu, K., Wei, S., Wu, M., & Li, Z. (2025). The synergistic effects of humic acid, chitosan and Bacillus subtilis on tomato growth and against plant diseases. Frontiers in Microbiology, 16, 1574765. https://doi.org/10.3389/fmicb.2025.1574765
Rathna Priya, T., Eliazer Nelson, A. R. L., Ravichandran, K., & Antony, U. (2019). Nutritional and functional properties of coloured rice varieties of South India: A review. Journal of Ethnic Foods, 6, 11. https://doi.org/10.1186/s42779-019-0017-3
Ritbamrung, O., Inthima, P., Ratanasut, K., Buddhachat, K., Chansongkram, W., Jumpathong, J., Sujipuli, K., & Boonsrangsom, T. (2025). Evaluating Xanthomonas oryzae pv. oryzae (Xoo) infection dynamics in rice for distribution routes and environmental reservoirs by molecular approaches. Scientific Reports, 15, 1408. https://doi.org/10.1038/s41598-025-85422-3
Saikia, K., & Bora, L. C. (2021). Exploring actinomycetes and endophytes of rice ecosystem for induction of disease resistance against bacterial blight of rice. European Journal of Plant Pathology, 159, 67–79. https://doi.org/10.1007/s10658-020-02145-9
Sakthivel, N., Mortensen, C. N., & Mathur, S. B. (2001). Detection of Xanthomonas oryzae pv. oryzae in artificially inoculated and naturally infected rice seeds and plants by molecular techniques. Applied Microbiology and Biotechnology, 56, 435–441. https://doi.org/10.1007/s002530100641
Sheteiwy, M. S., Dong, Q., An, J., Song, W., Guan, Y., He, F., & Hu, J. (2017). Regulation of ZnO nanoparticles-induced physiological and molecular changes by seed priming with humic acid in Oryza sativa seedlings. Plant Growth Regulation, 83(1), 27–41. https://doi.org/10.1007/s10725-017-0295-0
Shivalingaiah, U. S., & Umesha, S. (2013). Pseudomonas fluorescens inhibits the Xanthomonas oryzae pv. oryzae, the bacterial leaf blight pathogen in rice. Canadian Journal of Plant Protection, 1(5), 119–123.
Szymanek, M., Dziwulska-Hunek, A., & Tanas, W. (2020). The influence of effective microorganisms on the soil properties and yield of selected crops. Agriculture, 10(9), 400. https://doi.org/10.3390/agriculture10090400
Teja, B. S., Jamwal, G., Gupta, V., Verma, M., Sharma, A., Sharma, A., & Pandit, V. (2025). Biological control of bacterial leaf blight (BLB) in rice–A sustainable approach. Heliyon, 11, e41769. https://doi.org/10.1016/j.heliyon.2025.e41769
Tyagi, A., Tamang, T. L., Kashtoh, H., Mir, R. A., Mir, Z. A., Manzoor, S., et al. (2024). A review on biocontrol agents as sustainable approach for crop disease management: Applications, production, and future perspectives. Horticulturae, 10, 805. https://doi.org/10.3390/horticulturae10080805
Udayashankar, A. C., Nayaka, S. C., Reddy, M. S., & Srinivas, C. (2011). Plant growth-promoting rhizobacteria mediate induced systemic resistance in rice against bacterial leaf blight caused by Xanthomonas oryzae pv. oryzae. Biological Control, 59(2), 114–122. https://doi.org/10.1016/j.biocontrol.2011.05.010
Wang, R., Ji, Z., Kou, Y., Yang, X., Huang, W., Sun, Z., et al. (2025). Occurrence and integrated control of major rice diseases in China. New Plant Protection, 2, e70004.
Yasmin, S., Hafeez, F. Y., Mirza, M. S., Rasul, M., Arshad, H. M. I., Zubair, M., & Iqbal, M. (2017). Biocontrol of bacterial leaf blight of rice and profiling of secondary metabolites produced by rhizospheric Pseudomonas aeruginosa BRp3. Frontiers in Microbiology, 8, 1895. https://doi.org/10.3389/fmicb.2017.01895
Zhu, D., Wei, H., Guo, B., Dai, Q., Wei, C., Gao, H., & Huo, Z. (2017). The effects of chilling stress after anthesis on the physicochemical properties of rice (Oryza sativa L) starch. Food Chemistry, 237, 936–941. https://doi.org/10.1016/j.foodchem.2017.06.039
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