Growth-promoting Effect of Arcopilus aureus XBAa1 on Tobacco and Its Colonization in Tobacco Root

doi:10.13560/j.cnki.biotech.bull.1985.2025-0732

Abstract

Abstract:

Objective This work is to investigate the growth-promoting effects of Arcopilus aureus strain XBAa1 on tobacco and the colonization dynamics in roots. Method By reconstruction of the tobacco-XBAa1 fungal symbiosis, its endophytic performance and plant growth promotion was evaluated. Based on the PEG-mediated protoplast transformation method, GFP gene carried by the P3641 plasmid was integrated into A. aureus XBAa1 genome, the transformants stably expressing GFP were obtained. Transformants were used to infect tobacco roots. The infection sites and colonization levels of strain XBAa1 within tobacco roots were detected using laser confocal microscopy (LCM) and quantitative absolute real-time PCR assays. Result A. aureus XBAa1 endophytically colonized in tobacco roots, which significantly promoted tobacco growth. The biomasses of the shoots and roots increased remarkably, with the chlorophyll a+b contents in the leaves enhanced by 63.17% compared with CK. It regulated the changes in hormones of tobacco, with cytokinin content increased by 143.65% and abscisic acid content decreased by 46.56% relative to CK, respectively. Meanwhile, it reduced the degree of oxidative damage in tobacco, and the malondialdehyde content decreased by 38.02% compared with CK. The optimal cell wall lysis conditions for protoplast preparation of XBAa1 were enzymatic hydrolysis with 12.5 mg/mL megalyase solution treated for 3 h. For the obtained XBAa1-Y3 transformant, laser confocal microscopy observations revealed that it colonized stably in the intercellular spaces of tobacco root tissue cells. Under the condition of sterile substrate in the greenhouse, the colonization biomasses of XBAa1-GFP strain in tobacco roots showed a “first increasing and then decreasing” trend within 30 d post-inoculation (dpi), reaching a peak of 1.23×10⁷ copies/mL at 15 dpi; at 30 dpi, the colonization density decreased to 8.12×10⁶ copies/mL. Conclusion A. aureus XBAa1 had stable colonization in tobacco roots and remarkable growth-promoting effects on host plants, indicating considerable application potential in agriculture. The successfully obtained GFP-tagged XBAa1 transformant verified that protoplasts are feasible materials for subsequent genetic transformation, establishing a critical basis for an in-depth clarifying the migration traits of XBAa1 in tobacco plants and its biocontrol functions.

Key words: Arcopilus aureus, endophytic fungi, protoplast, endogenous hormones, colonization

GENG Ming-yan, FENG Hui, HE Lei, ZHANG Guo-zheng, WANG Jing. Growth-promoting Effect of Arcopilus aureus XBAa1 on Tobacco and Its Colonization in Tobacco Root[J]. Biotechnology Bulletin, 2026, 42(4): 321-331.

Figures/Tables 8

Fig. 1 Re-isolation of strain XBAa1 in tobacco rootA, C: Sterile water. B, D: Strain XBAa1

Fig. 2 Promoting effects of XBAa1 on early growth of tobacco under tissue culture and sterile substrate conditionsA, B: Tobacco-XBAa1 symbiosis in tissue culture. C, D, E, F: Above ground and root of the tobacco-XBAa1 symbiosis in the greenhouse. G: Root weight. H: Shoot weight (* P<0.05, **P<0.01, ***P<0.001, the same below)

Fig. 4 Sensitivity of wild type Arcopilus aureus XBAa1 to Neo

Fig. 5 Release of strain XBAa1 protoplastsA: Top release. B: Lateral release. C: In-situ release

Fig. 6 PCR detection of strain XBAa1-GFP and observation in tobacco root colonization by confocal microscope (40×)

Fig. 7 Colony characteristics of tagged strain XBAa1-GFP and wild type strain XBAa1A: Wild type XBAa1. B: XBAa1-GFP strain. C: Growth rates of strains

Fig. 8 Observation of colonization of XBAa1 in tobacco root by confocal microscopy

Fig. 9 Colonization dynamics of strain XBAa1 in tobacco roots

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