Assessment of strain-dependent variation of phytostimulatory activity in Streptomyces ambofaciens-based bioformulations for agricultural applications

O. V. Andriushchenko; I. V. Strashnova; T. V. Ivanytsia; V. O. Ivanytsia; M. B. Galkin

doi:10.15421/0225206

O. V. Andriushchenko Odesa National I. I. Mechnikov University
I. V. Strashnova Odesa National I. I. Mechnikov University
T. V. Ivanytsia Odesa National I. I. Mechnikov University
V. O. Ivanytsia Odesa National I. I. Mechnikov University
M. B. Galkin Odesa National I. I. Mechnikov University

Keywords: plant growth promotion, barley, mono-strain inoculants, multi-species inoculants, biofertilizers.

Abstract

The biotechnological potential of Streptomyces ambofaciens is highly strain-dependent, yet practical evaluations of how di f ferent strains influence the effects of microbial complexes used for agricultural bioformulations remain unstudied. In this invest i gation, we evaluated two microbial complexes differing only in their Streptomyces component: complex A1 containing S. ambofaciens ONU 1016, and complex A3 containing S. ambofaciens ONU 561, both combined with Bacillus subtilis ONU 1125 and Trichoderma harzianum LBX-181. Barley grains were inoculated with microbial suspensions across a concentration range to reveal how Streptomyces strain selection affects complex-mediated plant growth responses. Our findings showed that S. ambofaciens strains acted differently when applied alone versus in microbial complexes, reshaping the plant growth responses. S. ambofaciens ONU 1016 and ONU 561, as single inoculants, increased the fresh weight of the barley seedlings by 9.0 – 51.4% when applied as 75% suspensions. This gain in weight was the result of ability of the studied strains to increase leaf area and height of the barley seedlings at this concentration. PCA showed that the effects of S. ambofaciens ONU 1016 and ONU 561 on plant growth, similar at 10 – 50% concentrations (distance in PCA space 0.1 – 0.5), began to diverge when 75% suspensions were applied: ONU 1016 acted evenly across height, leaf area, and roots, whereas ONU 561 shifted away from root promotion, with the distances between them in PCA space 1.7 at 75% and 2.1 at 100%. When included into microbial complexes with B. subtilis ONU 1125 and T. harzianum LBX-181, differences in phytostimulatory effects became more pronounced. The A1 complex consistently promoted both aboveground biomass and roots at all concentrations, although increases in fresh weights were signi f icant only at 10% and 25%. The A3 complex increased fresh weight only at 10% due to improved leaf growth, but at higher concentrations it only promoted root development while suppressing aboveground biomass growth. PCA confirmed that the A1 and A3 were far apart, with distances of 4.0 – 5.4 across concentrations, reflecting their fundamentally different orientations. Thus, the inclusion of either strain determines whether multi-microbial mixtures support balanced plant growth (A1) or only root-focused responses (A3). The selection of an appropriate S. ambofaciens strain is crucial for enhancing the performance of micr o bial fertilizers.

References

Andriuschenko, O. V., Strashnova, I. V., Ivanytsia, T. V., Rakytska, S. I., & Galkin, M. B. (2024). Antagonistic activity of Black Sea actinobacteria against phytopathogenic microorganisms. Microbiology and Biotechnology, 62(3), 59–81.

Ankati, S., Srinivas, V., Pratyusha, S., & Gopalakrishnan, S. (2021). Streptomyces consortia-mediated plant defense against Fusarium wilt and plant growth-promotion in chickpea. Microbial Pathogens, 157, 104961.

de Andrade, da Silva, M. S. R., de Carvalho, L. A. L., Santos, C. H. B., Frezarin, E. T., da Silva, C. G. N., Pinheiro, D. G., Zonta, E., Baba-lola, O. O., & Rogobelo, E. C. (2025). Effect of co-inoculation with plant growth-promoting bacteria on the microbiome of soybean roots. Frontiers Sustainable Food Systems, 9, 1505001.

de Souza, L. F., Oliveira, H. G., Pellegrinetti, T. A., Mendes, L. W., Bonatelli, M. L., Dumaresq, A. S. R., Sinatti, V. V., Pinheiro, J. B., Azevedo, J. L., & Quecine, M. C. (2025). Co-inoculation with Bacillus thuringiensis RZ2MS9 and rhizobia improves the soybean development and modulates soil functional diversity. FEMS Microbiology Ecology, 101(2), fiaf013.

Dev, A. S. R., Harish, S., Karthikeyan, G., Nivedha, M., & Sangeetha, C. (2024). Consortia of Streptomyces spp. triggers defense/PAMP genes during the interaction of Groundnut bud necrosis orthotospovirus in tomato. 3 Biotech, 9, 196.

Fondi, M., Pinatel, E., Talà, A., Damiano, F., Consolandi, C., Mattorre, B., Fico, D., Testini, M., De Benedetto, G.E., Siculella, L., De Bellis, G., Alifano, P., & Peano, C. (2017). Time-resolved transcriptomics and constraint-based modeling identify system-level metabolic features and overexpression targets to increase spiramycin production in Streptomyces ambofaciens. Frontiers in Microbiology, 8, 835.

Gates, A. D., French, A. M., Demetros, A. A., Kelley, B. R., & Lebeis, S. L. (2023). A Streptomyces consortium contributes distinct microbial interactions during Arabidopsis thaliana microbiome assembly. Phytobiomes, 7(4), 515–525.

Ghorbani-Nasrabadi, R., Greiner, R., Alikhani, H.A., Hamedi, J. (2012). Identification and determination of extracellular phytate-degrading activity in actinomycetes. World Journal of Microbiology and Biotechnology, 28(7), 2601–2608.

Haring, F., Rostia, E., Syam’un, E., & Ginting, N.M. (2019). Effect of Trichoderma sp. and Streptomyces sp. on the growth and production of true seed shallots (TSS). Earth and Environmental Science, 343, 012020.

He, D., Gao, C., Zhao, S., Chen, H., Li, P., Yang, X., Li, D., Zhao, T., Jiang, H., & Liu, C. (2024). Antibacterial, herbicidal, and plant growth-promoting properties of Streptomyces sp. STD57 from the rhizosphere of Adenophora stricta. Microorganisms, 12(11), 2245.

He, Y., Guo, W., Peng, J., Guo, J., Ma, J., Wang, X., Zhang, C., Jia, N., Wang, E., Hu, D., & Wang, Z. (2022). Volatile organic compounds of Streptomyces sp. TOR3209 stimulated tobacco growth by up-regulating the expression of genes related to plant growth and development. Frontiers in Microbiology, 13, 891245.

Heng, J. L. S., Shah, U., Rahman, N. A., Shaari, K., & Hamzah, H. (2015). Streptomyces ambofaciens S2 – a potential biological control agent for Colletotrichum gleosporioides the causal agent for anthracnose in red chilli fruits. Journal of Plant Pathology and Microbiology, 6, S1-006.

Jin, S., & Alberti, F. (2025). Advances in the discovery and study of Trichoderma natural products for biological control applications. Natural Products Reports, 42, 1367–1386.

Kabir, A. H., Thapa, A., Hasan, M. R., & Parvej, M. R. (2024). Local signal from Trichoderma afroharzianum T22 induces host transcriptome and endophytic microbiome leading to growth promotion in sorghum. Journal of Experimental Botany, 75(22), 7107–7126.

Kirubakaran, R., Shameem, N., Saranya, E., Meenambigai, K., Dhanasekar, R., Parray, J. A., Yadav, N., Singh, S., Rustagi, S., Puri, P., Sharma, B., Negi, R., & Yadav, A. N. (2025). Streptomyces as endomicrobiome: Potential bioinoculants for agricultural sustainability. Journal of Applied Biology and Biotechnology, 13(4), 115.

Kuang, A., Fu, X., Liu, Z., Chen, Q., Jin, R., & Mao, H. (2024). Biocontrol effect of the complex inoculants of Trichoderma and Bacillus amyloliquefaciens on chrysanthemum white rust. Biocatalysis and Agricultural Biotechnology, 56, 103010.

Kulik, T., Staniszewska, P., Wisniewski, P., Treder, Z., Przybylski, M., Wronska, E., Mazdziarz, M., Krawczyk,K., Bilska, K., Paukszto, L., & Olszewski, J. (2025). In-depth comparison of commercial Trichoderma-based products: Integrative approaches to quantitative analysis, taxonomy and efficacy. Frontiers in Microbiology, 16, 1646394.

Liu, X., Mei, S., & Salles, J. F. (2023). Inoculated microbial consortia perform better than single strains in living soil: A meta-analysis. Applied Soil Ecology, 190, 105011.

Liu, Y., Jia, B., Ren, Y., Xun, W., Stefanic, P., Yang, T., Miao, Y., Zhang, N., Yao, Y., Zhang, R., Xu, Z., Shen, Q., & Mandic-Mulec, I. (2025). Bacterial social interactions in synthetic Bacillus consortia enhance plant growth. IMeta, 4(4), e70053.

Matselyukh, B. P., Golembiovska, S. L., & Bambura, O. I. (2020). Screening of soil Streptomycetes – producers of antibiotics against phytopathogenic bacteria. Mikrobiologichnyi Zhurnal, 82(5), 36–40.

Nguyen, H. C., Karray, F., Lautru, S., Gagnat, J., Lebrihi, A., Ho Huynh, T. D., & Pernodet, J. (2010). Glycosylation steps during spiramycin biosynthesis in Streptomyces ambofaciens: Involvement of three glycosyltransferases and their interplay with two auxiliary proteins. Antimicrobial Agents and Chemotherapy, 54(7), 1602–1609.

Nonthakaew, N., Panbangred, W., & Intra, B. (2022). Plant growth-promoting properties of Streptomyces spp. isolates and their impact on mung bean plantlets’ rhizosphere microbiome. Frontiers in Microbiology, 13, 967415.

Orouji, E., Fathi Ghare Baba, M., Sadeghi, A.,Gharanjik, S., & Koobaz, P. (2023). Specific Streptomyces strain enhances the growth, defensive mechanism, and fruit quality of cucumber by minimizing its fertilizer consumption. BMC Plant Biology, 23, 246.

Özdemir Koçak, F. (2019). Identification of Streptomyces strains isolated from Humulus lupulus rhizosphere and determination of plant growth promotion potential of selected strains. Turkish Journal of Biology, 43(6), 391–403.

Potapenko, K., Lisiutin, G., Vasylieva, N., Strashnova, I., Franke, R., Petriv, N., Duduyemi, O. P., Baklan, K., Korotaieva, N., Gudzenko, T., Manns, M. P., Broenstrup, M., Lenzen, H., Vital, M., Ivanytsia, V., & Yevsa, T. (2025). Antimicrobial and anticancer activity of Streptomyces ambofaciens (Myt 8) and S. globisporus ONU 1019 (Myt 11) secondary metabolites isolated from the Odesa Bay, the Black Sea: An in vitro study. Biomedicine and Pharmacotherapy, 186, 117981.

Prigigallo, M. I., Staropoli, A., Vinale, F., & Bubici, G. (2023). Interactions between plant-beneficial microorganisms in a consortium: Streptomyces microflavus and Trichoderma harzianum. Microbial Biotechnology, 16, 2292–2312.

Qiao, R., Xu, M., Jiang, J., Song, Z., Wang, M., Yang, L., Guo, H., & Mao, Z. (2024). Plant growth promotion and biocontrol properties of a synthetic community in the control of apple disease. BMC Plant Biology, 24, 546.

Ramírez-Pool, J. A., Calderón-Pérez, B., Ruiz-Medrano, R., Ortiz-Castro, R., & Xoconostle-Cazares, B. (2024). Bacillus strains as effective biocontrol agents against phytopathogenic bacteria and promoters of plant growth. Microbial Ecology, 87(1), 76.

Reid, T. E., & Gifford, M. L. (2024). Trichoderma gets by with a little help from Streptomyces: Fungal-bacterial symbiosis in plant growth promotion. Journal of Experimental Botany, 75(22), 6893–6897.

Rigobelo, E.C., de Andrade, L. A., Santos, C. H. B., Frezarin, E. T., Sales, L. R., de Carvalho, L. A. L., Guariz Pinheiro, D., Nicodemo, D., Babalola, O. O., Verdi, M. C. Q., Mondin, M., & Desoignies, N. (2024). Effects of Trichoderma harzianum and Bacillus subtilis on the root and soil microbiomes of the soybean plant INTACTA RR2 PROTM. Frontiers in Plant Sciences, 15, 1403160.

Santoyo, G., Orozco-Mosqueda, M. C., Afridi, M. S., Mitra, D., Valencia-Can-tero, E., & Macias-Rodriguez, L. (2024). Trichoderma and Bacillus multifactorial allies for plant growth and health in saline soils: Recent advances and future challenges. Frontiers in Microbiology, 15, 1423980.

Schmidt, R., Koberl, M., Mostafa, A., Ramadan, E. M., Monschein, M., Jensen, K. B., Rudolf, B., & Berg, G. (2014). Effects of bacterial inoculants on the indigenous microbiome and secondary metabolites of chamomile plants. Frontiers in Microbiology, 5, 64.

Sharma, N., Mahawar, L., Mishra, A., & Albrectsen, B. R. (2025). Microbial contributions to plant growth and stress tolerance: Mechanisms for sustainable plant production. Plant Stress, 17, 100966.

Shtenikov, M. D., Ostapchuk, A. M., & Ivanytsia, V. O. (2018). Antagonistic activity of endospore forming bacteria of deep water the Black Sea sediments. Miсrobiology and Biotechnology, 43(3), 82–89.

Staropoli, A., Vassetti, A., Salvatore, M. M., Andolfi, A., Prigigallo, M. I., Bubici, G., Scagliola, M., Salerno, P., & Vinale, F. (2021). Improvement of nutraceutical value of parsley leaves (Petroselinum crispum) upon field applications of beneficial microorganisms. Horticulturae, 7(9), 281.

Sun, T., Liu, H., Wang, N., Huang, M., Banerjee, S., Jousset, A., Xu, Y., Shen, Q., Wang, S., Wang, X., & Wei, Z. (2025). Interactions with native microbial keystone taxa enhance the biocontrol efficiency of Streptomyces. Microbiome, 13, 126.

Vuolo, F., Novello, G., Bona, E., Gorrasi, S., & Gamalero, E. (2022). Impact of plant-beneficial bacterial inocula on the resident bacteriome: Current knowledge and future perspectives. Microorganisms, 10(12), 2462.