Plant growth-promoting rhizobacteria and Trichoderma shift common vetch (Vicia sativa) physiology and phyllosphere bacteria toward antagonism against anthracnose caused by Colletotrichum spinaciae

Abstract

Background: Plant phyllosphere microbes are important for the host plant’s protection. Plant growth-promoting rhizobacteria (PGPR) and Trichoderma are common biocontrol agents (BCAs) for disease management. Pathogens and BCAs can change the rhizosphere microbial composition; however, the effect of PGPR or Trichoderma on plant phyllosphere microbes, particularly for mesocosms involving the interaction between pathogens and BCAs, is not well known.

Methods: High-throughput sequencing was used to identify the phyllosphere bacterial community of common vetch interacting with Colletotrichum spinaciae, two PGPRs (Bacillus subtilis and Bacillus licheniformis), and Trichoderma longibrachiatum. We evaluated anthracnose severity, phyllosphere bacteria diversity and composition, and the relationship between the activities of plant defense enzymes and hormonal molecules in plants treated with individual and combined inoculations of PGPRs, Trichoderma, and C. spinaciae.

Results: PGPR or Trichoderma alone reduced disease severity. Trichoderma reduced the salicylic acid content, PGPR increased the catalase activity in plants, and co-inoculation of PGPR and Trichoderma decreased the salicylic acid content. Inoculation of PGPR and Trichoderma individually or in combination changed the disease-associated phyllosphere bacteria, and this effect was related to plant defense enzymes and hormonal molecules.

Conclusions: We suggest that the plant defense response induced by PGPR and Trichoderma results in the enrichment of a fraction of favorable chloroplastic bacteria, which facilitates plant defense against diseases.

Keywords

anthracnose / biocontrol / PGPR / phyllosphere bacteria / physiology / Trichoderma

References

[1] Abadi, V. A. J. M., Sepehri, M., Rahmani, H. A., Zarei, M., Ronaghi, A., Taghavi, S. M., & Shamshiripour, M. (2020). Role of dominant phyllosphere bacteria with plant growth-promoting characteristics on growth and nutrition of maize (Zea mays L.). Journal of Soil Science and Plant Nutrition, 20(4), 2348–2363.

[2] Andrews, J. H., & Harris, R. F. (2000). The ecology and biogeography of microorganisms on plant surfaces. Annual Review of Phytopathology, 38, 145–180.

[3] Arif, I., Batool, M., & Schenk, P. M. (2020). Plant microbiome engineering: Expected benefits for improved crop growth and resilience. Trends in Biotechnology, 38(12), 1385–1396.

[4] Avis, T. J., Gravel, V., Antoun, H., & Tweddell, R. J. (2008). Multifaceted beneficial effects of rhizosphere microorganisms on plant health and productivity. Soil Biology & Biochemistry, 40(7), 1733–1740.

[5] Boss, B. L., Wanees, A. E., Zaslow, S. J., Normile, T. G., & Izquierdo, J. A. (2022). Comparative genomics of the plant-growth promoting bacterium Sphingobium sp. strain AEW4 isolated from the rhizosphere of the beachgrass Ammophila breviligulata. BMC Genomics, 23(1), 508.

[6] Caporaso, J. G., Lauber, C. L., Walters, W. A., Berg-Lyons, D., Huntley, J., Fierer, N., Owens, S. M., Betley, J., Fraser, L., Bauer, M., Gormley, N., Gilbert, J. A., Smith, G., & Knight, R. (2012). Ultra-high-throughput microbial community analysis on the illumina HiSeq and MiSeq platforms. The ISME Journal, 6(8), 1621–1624.

[7] Carrión, V. J., Perez-Jaramillo, J., Cordovez, V., Tracanna, V., de Hollander, M., Ruiz-Buck, D., Mendes, L. W., van Ijcken, W. F. J., Gomez-Exposito, R., Elsayed, S. S., Mohanraju, P., Arifah, A., van der Oost, J., Paulson, J. N., Mendes, R., van Wezel, G. P., Medema, M. H., & Raaijmakers, J. M. (2019). Pathogen-induced activation of disease-suppressive functions in the endophytic root microbiome. Science, 366(6465), 606–612.

[8] Carvalhais, L. C., Dennis, P. G., Badri, D. V., Kidd, B. N., Vivanco, J. M., & Schenk, P. M. (2015). Linking jasmonic acid signaling, root exudates, and rhizosphere microbiomes. Molecular Plant-Microbe Interactions, 28(9), 1049–1058.

[9] Chandanie, W. A., Kubota, M., & Hyakumachi, M. (2006). Interactions between plant growth promoting fungi and arbuscular mycorrhizal fungus Glomus mosseae and induction of systemic resistance to anthracnose disease in cucumber. Plant and Soil, 286(1–2), 209–217.

[10] Chen, L., Jiang, Y., Liang, C., Luo, Y., Xu, Q., Han, C., Zhao, Q., & Sun, B. (2019). Competitive interaction with keystone taxa induced negative priming under biochar amendments. Microbiome, 7, 77.

[11] Chen, X., Marszałkowska, M., & Reinhold-Hurek, B. (2020). Jasmonic acid, not salicyclic acid restricts endophytic root colonization of rice. Frontiers in Plant Science, 10, 1758.

[12] Cui, H. Y., Sun, W., Delgado-Baquerizo, M., Song, W. Z., Ma, J. Y., Wang, K. Y., & Ling, X. L. (2020). The effects of mowing and multi-level N fertilization on soil bacterial and fungal communities in a semiarid grassland are year-dependent. Soil Biology & Biochemistry, 151, 108040.

[13] Darre, M. J., Minior, D. N., Tatake, J. G., & Ressler, C. (1998). Nutritional evaluation of detoxified and raw common vetch seed (Vicia sativa L.) using diets of broilers. Journal of Agricultural & Food Chemistry, 46(11), 4675–4679.

[14] Delaney, T. P. (1997). Genetic dissection of acquired resistance to disease. Plant Physiology, 113(1), 5–12.

[15] Díaz-Cruz, G. A., & Cassone, B. J. (2022). Changes in the phyllosphere and rhizosphere microbial communities of soybean in the presence of pathogens. FEMS Microbiology Ecology, 98(3), fiac022.

[16] Dixon, P. (2003). VEGAN, a package of R functions for community ecology. Journal of Vegetation Science, 14(6), 927–930.

[17] Freeman, S., Minz, D., Kolesnik, I., Barbul, O., Zveibil, A., Maymon, M., Nitzani, Y., Kirshner, B., Rav-David, D., Bilu, A., Dag, A., Shafir, S., & Elad, Y. (2004). Trichoderma biocontrol of Colletotrichum acutatum and Botrytis cinerea and survival in strawberry. European Journal of Plant Pathology, 110(4), 361–370.

[18] Gao, P., Li, Y. D., Guo, Y. E., & Duan, T. Y. (2018). Co-inoculation of lucerne (Medicago sativa) with an AM fungus and a rhizobium reduces occurrence of spring black stem and leaf spot caused by Phoma medicaginis. Crop & Pasture Science, 69(9), 933–943.

[19] Guevara-Avendaño, E., Bejarano-Bolívar, A. A., Kiel-Martínez, A. L., Ramírez-Vázquez, M., Méndez-Bravo, A., von Wobeser, E. A., Sánchez-Rangel, D., Guerrero-Analco, J. A., Eskalen, A., & Reverchon, F. (2019). Avocado rhizobacteria emit volatile organic compounds with antifungal activity against Fusarium solani, Fusarium sp. associated with Kuroshio shot hole borer, and Colletotrichum gloeosporioides. Microbiological Research, 219, 74–83.

[20] Gutiérrez-Moreno, K., Ruocco, M., Monti, M. M., Vega, O. M., & Heil, M. (2021). Context-dependent effects of Trichoderma seed inoculation on anthracnose disease and seed yield of bean (Phaseolus vulgaris): Ambient conditions override cultivar-specific differences. Plants, 10(8), 1739.

[21] Harman, G. E., Howell, C. R., Viterbo, A., Chet, I., & Lorito, M. (2004). Trichoderma species: Opportunistic, avirulent plant symbionts. Nature Reviews Microbiology, 2(1), 43–56.

[22] Hirano, S. S., & Upper, C. D. (2000). Bacteria in the leaf ecosystem with emphasis on Pseudomonas syringae: A pathogen, ice nucleus, and epiphyte. Microbiology and Molecular Biology Reviews, 64(3), 624–653.

[23] Huang, S., Sheng, P., & Zhang, H. (2012). Isolation and identification of cellulolytic bacteria from the gut of Holotrichia parallela larvae (Coleoptera: Scarabaeidae). International Journal of Molecular Sciences, 13(3), 2563–2577.

[24] Izquierdo-García, L. F., González-Almario, A., Cotes, A. M., & Moreno-Velandia, C. A. (2020). Trichoderma virens Gl006 and Bacillus velezensis Bs006: A compatible interaction controlling Fusarium wilt of cape gooseberry. Scientific Reports, 10(1), 6857.

[25] Jalmi, S. K., & Sinha, A. K. (2022). Ambiguities of PGPR-induced plant signaling and stress management. Frontiers in Microbiology, 13, 899563.

[26] Jetiyanon, K., & Kloepper, J. W. (2002). Mixtures of plant growth-promoting rhizobacteria for induction of systemic resistance against multiple plant diseases. Biological Control, 24(3), 285–291.

[27] Jing, X., Sanders, N. J., Shi, Y., Chu, H., Classen, A. T., Zhao, K., Chen, L., Shi, Y., Jiang, Y., & He, J. S. (2015). The links between ecosystem multifunctionality and above- and below-ground biodiversity are mediated by climate. Nature Communications, 6, 8159.

[28] Kim, D. R., Cho, G., Jeon, C. W., Weller, D. M., Thomashow, L. S., Paulitz, T. C., & Kwak, Y. S. (2019). A mutualistic interaction between Streptomyces bacteria, strawberry plants and pollinating bees. Nature Communications, 10, 4802.

[29] Kloepper, J. W., Leong, J., Teintze, M., & Schroth, M. N. (1980). Enhanced plant-growth by siderophores produced by plant growth-promoting rhizobacteria. Nature, 286(5776), 885–886.

[30] Kwak, M. J., Kong, H. G., Choi, K., Kwon, S. K., Song, J. Y., Lee, J., Lee, P. A., Choi, S. Y., Seo, M., Lee, H. J., Jung, E. J., Park, H., Roy, N., Kim, H., Lee, M. M., Rubin, E. M., Lee, S. W., & Kim, J. F. (2018). Rhizosphere microbiome structure alters to enable wilt resistance in tomato. Nature Biotechnology, 36(11), 1100–1109.

[31] Larkin, R. P., & Fravel, D. R. (1998). Efficacy of various fungal and bacterial biocontrol organisms for control of Fusarium wilt of tomato. Plant Disease, 82(9), 1022–1028.

[32] Lebeis, S. L., Paredes, S. H., Lundberg, D. S., Breakfield, N., Gehring, J., McDonald, M., Malfatti, S., Glavina del Rio, T., Jones, C. D., Tringe, S. G., & Dangl, J. L. (2015). Salicylic acid modulates colonization of the root microbiome by specific bacterial taxa. Science, 349(6250), 860–864.

[33] Li, P. D., Zhu, Z. R., Zhang, Y., Xu, J., Wang, H., Wang, Z., & Li, H. (2022). The phyllosphere microbiome shifts toward combating melanose pathogen. Microbiome, 10(1), 56.

[34] Li, Y., Nan, Z., Matthew, C., Wang, Y., & Duan, T. (2023). Arbuscular mycorrhizal fungus changes alfalfa (Medicago sativa) metabolites in response to leaf spot (Phoma medicaginis) infection, with subsequent effects on pea aphid (Acyrthosiphon pisum) behavior. New Phytologist, 239(1), 286–300.

[35] Liu, C. M., Yang, Z. F., He, P. F., Munir, S., Wu, Y. X., Ho, H. H., & He, Y. Q. (2018). Deciphering the bacterial and fungal communities in clubroot-affected cabbage rhizosphere treated with Bacillus Subtilis XF-1. Agriculture, Ecosystems & Environment, 256, 12–22.

[36] Liu, K., McInroy, J. A., Hu, C. H., & Kloepper, J. W. (2018). Mixtures of plant-growth-promoting rhizobacteria enhance biological control of multiple plant diseases and plant-growth promotion in the presence of pathogens. Plant Disease, 102(1), 67–72.

[37] Lugtenberg, B., & Kamilova, F. (2009). Plant-growth-promoting rhizobacteria. Annual Review of Microbiology, 63, 541–556.

[38] Manzar, N., Singh, Y., Kashyap, A. S., Sahu, P. K., Rajawat, M. V. S., Bhowmik, A., Sharma, P. K., & Saxena, A. K. (2021). Biocontrol potential of native Trichoderma spp. against anthracnose of great millet (Sorghum bicolour L.) from Tarai and hill regions of India. Biological Control, 152, 104474.

[39] Mathews, A. A., Basha, S. T., & Reddy, N. P. E. (2011). Management of post harvest disease of mango anthracnose. Journal of Pure and Applied Microbiology, 5(1), 97–104.

[40] Mawarda, P. C., Le Roux, X., Van Elsas, J. D., & Salles, J. F. (2020). Deliberate introduction of invisible invaders: A critical appraisal of the impact of microbial inoculants on soil microbial communities. Soil Biology & Biochemistry, 148, 107874.

[41] Mia, M. A. B., Shamsuddin, Z. H., Wahab, Z., & Marziah, M. (2010). Effect of plant growth promoting rhizobacterial (PGPR) inoculation on growth and nitrogen incorporation of tissue-cultured Musa plantlets under nitrogen-free hydroponics condition. Australian Journal of Crop Science, 4(2), 85–90.

[42] Mikić A., Mihailović V., Ćupina, B., Milić D., Katić S., Karagić Đ., Pataki, I., D’Ottavio, P., & Kraljević-Balalić M. (2014). Forage yield components and classification of common vetch (Vicia sativa L.) cultivars of diverse geographic origin. Grass and Forage Science, 69(2), 315–322.

[43] Mille-Lindblom, C., Fischer, H., & J. Tranvik, L. (2006). Antagonism between bacteria and fungi: Substrate competition and a possible tradeoff between fungal growth and tolerance towards bacteria. Oikos, 113(2), 233–242.

[44] Min, X., Liu, Z., Wang, Y., & Liu, W. (2020). Comparative transcriptomic analysis provides insights into the coordinated mechanisms of leaves and roots response to cold stress in common vetch. Industrial Crops and Products, 158, 112949.

[45] Nakayama, T., Homma, Y., Hashidoko, Y., Mizutani, J., & Tahara, S. (1999). Possible role of xanthobaccins produced by Stenotrophomonas sp strain SB-K88 in suppression of sugar beet damping-off disease. Applied and Environmental Microbiology, 65(10), 4334–4339.

[46] Niu, B., Wang, W., Yuan, Z., Sederoff, R. R., Sederoff, H., Chiang, V. L., & Borriss, R. (2020). Microbial interactions within multiple-strain biological control agents impact soil-borne plant disease. Frontiers in Microbiology, 11, 585404.

[47] Oleńska, E., Małek, W., Wójcik, M., Swiecicka, I., Thijs, S., & Vangronsveld, J. (2020). Beneficial features of plant growth-promoting rhizobacteria for improving plant growth and health in challenging conditions: A methodical review. Science of the Total Environment, 743, 140682.

[48] Penha, R. O., Vandenberghe, L. P. S., Faulds, C., Soccol, V. T., & Soccol, C. R. (2020). Bacillus lipopeptides as powerful pest control agents for a more sustainable and healthy agriculture: Recent studies and innovations. Planta, 251(3), 70.

[49] Rais, A., Jabeen, Z., Shair, F., Hafeez, F. Y., & Hassan, M. N. (2017). Bacillus spp., a bio-control agent enhances the activity of antioxidant defense enzymes in rice against Pyricularia oryzae. PLoS One, 12(11), e0187412.

[50] Ryder, M. H., Yan, Z. N., Terrace, T. E., Rovira, A. D., Tang, W. H., & Correll, R. L. (1999). Use of strains of Bacillus isolated in China to suppress take-all and Rhizoctonia root rot, and promote seedling growth of glasshouse-grown wheat in Australian soils. Soil Biology & Biochemistry, 31(1), 19–29.

[51] Sarma, B. K., Yadav, S. K., Singh, S., & Singh, H. B. (2015). Microbial consortium-mediated plant defense against phytopathogens: Readdressing for enhancing efficacy. Soil Biology & Biochemistry, 87, 25–33.

[52] Schlechter, R. O., Miebach, M., & Remus-Emsermann, M. N. P. (2019). Driving factors of epiphytic bacterial communities: A review. Journal of Advanced Research, 19(SI), 57–65.

[53] Segata, N., Izard, J., Waldron, L., Gevers, D., Miropolsky, L., Garrett, W. S., & Huttenhower, C. (2011). Metagenomic biomarker discovery and explanation. Genome Biology, 12(6), R60.

[54] Shi, F. M., Yao, L. L., Pei, B. L., Zhou, Q., Li, X. L., Li, Y., & Li, Y. Z. (2009). Cortical microtubule as a sensor and target of nitric oxide signal during the defence responses to Verticillium dahliaetoxins in Arabidopsis. Plant, Cell & Environment, 32(4), 428–438.

[55] Shu, Z., Zhang, X. S., Chen, J., Chen, G. Y., & Xu, D. Q. (2010). The simplification of chlorophyll content measurement. Plant Physiology Journal, 46(4), 399–402.

[56] Singh, U. B., Malviya, D., Wasiullah, Singh, S., Pradhan, J. K., Singh, B. P., Roy, M., Imram, M., Pathak, N., Baisyal, B. M., Rai, J. P., Sarma, B. K., Singh, R. K., Sharma, P. K., Kaur, S. D., Manna, M. C., Sharma, S. K., & Sharma, A. K. (2016). Bio-protective microbial agents from rhizosphere eco-systems trigger plant defense responses provide protection against sheath blight disease in rice (Oryza sativa L.). Microbiological Research, 192, 300–312.

[57] Sohrabi, R., Paasch, B. C., Liber, J. A., & He, S. Y. (2023). Phyllosphere microbiome. Annual Review of Plant Biology, 74, 539–568.

[58] Spadaro, D., & Droby, S. (2016). Development of biocontrol products for postharvest diseases of fruit: The importance of elucidating the mechanisms of action of yeast antagonists. Trends in Food Science & Technology, 47, 39–49.

[59] Vorholt, J. A. (2012). Microbial life in the phyllosphere. Nature Reviews Microbiology, 10(12), 828–840.

[60] Vorholt, J. A., Vogel, C., Carlström, C. I., & Müller, D. B. (2017). Establishing causality: Opportunities of synthetic communities for plant microbiome research. Cell Host & Microbe, 22(2), 142–155.

[61] Wang, Q., Duan, T. Y., & Nan, Z. B. (2019). First report of anthracnose caused by Colletotrichum spinaciae on Vicia sativa in China. Plant Disease, 103(8), 2138–2139.

[62] Wang, W., Fei, Y., Wang, Y., Song, B., Li, L., Zhang, W., Cheng, H., Zhang, X., Chen, S., & Zhou, J. M. (2023). SHOU4/4L link cell wall cellulose synthesis to pattern triggered immunity. New Phytologist, 238(4), 1620–1635.

[63] Wang, X. M., Yang, B., Ren, C. G., Wang, H. W., Wang, J. Y., & Dai, C. C. (2015). Involvement of abscisic acid and salicylic acid in signal cascade regulating bacterial endophyte-induced volatile oil biosynthesis in plantlets of Atractylodes lancea. Physiologia Plantarum, 153(1), 30–42.

[64] Wawrzynska, A., Christiansen, K. M., Lan, Y., Rodibaugh, N. L., & Innes, R. W. (2008). Powdery mildew resistance conferred by loss of the enhanced disease resistance1 protein kinase is suppressed by a missense mutation in keep on going, a regulator of abscisic acid signaling. Plant Physiology, 148(3), 1510–1522.

[65] Van Wees, S. C., Van der Ent, S., & Pieterse, C. M. (2008). Plant immune responses triggered by beneficial microbes. Current Opinion in Plant Biology, 11(4), 443–448.

[66] Xiong, W., Guo, S., Jousset, A., Zhao, Q. Y., Wu, H. S., Li, R., Kowalchuk, G. A., & Shen, Q. R. (2017). Bio-fertilizer application induces soil suppressiveness against Fusarium wilt disease by reshaping the soil microbiome. Soil Biology & Biochemistry, 114, 238–247.

[67] Xu, N., Zhao, Q., Zhang, Z., Zhang, Q., Wang, Y., Qin, G., Ke, M., Qiu, D., Peijnenburg, W. J. G. M., Lu, T., & Qian, H. (2022). Phyllosphere microorganisms: sources, drivers, and their interactions with plant hosts. Journal of Agricultural and Food Chemistry, 70(16), 4860–4870.

[68] Yin, C., Casa Vargas, J. M., Schlatter, D. C., Hagerty, C. H., Hulbert, S. H., & Paulitz, T. C. (2021). Rhizosphere community selection reveals bacteria associated with reduced root disease. Microbiome, 9(1), 86.

[69] Zarraonaindia, I., Owens, S. M., Weisenhorn, P., West, K., Hampton-Marcell, J., Lax, S., Bokulich, N. A., Mills, D. A., Martin, G., Taghavi, S., van der Lelie, D., & Gilbert, J. A. (2015). The soil microbiome influences grapevine-associated microbiota. mBio, 6(2), e02527–14.

[70] Zhang, Z., & Yuen, G. Y. (2000). The role of chitinase production by Stenotrophomonas maltophilia strain C3 in biological control of Bipolaris sorokiniana. Phytopathology, 90(4), 384–389.

RIGHTS & PERMISSIONS

2024 The Authors. Grassland Research published by John Wiley & Sons Australia, Ltd on behalf of Chinese Grassland Society and Lanzhou University.