Evaluation of the efficient propagation of Rhizophagus intraradices and its inoculation effects on rice

Abstract

Arbuscular mycorrhizal fungi (AMF) are a key group of fungi closely associated with agricultural production within soil microbial communities. However, large-scale propagation of AMF inoculum faces various challenges, limiting our ability to obtain and utilize these inocula on a broad scale. To address this, we designed a monolayer mesh cultivation system employing a hydroponic approach for propagating arbuscular mycorrhizal fungi, specifically Rhizophagus intraradices. We conducted a comparative analysis of quality and inoculation efficiency between the water culture inoculum (w-Ri) and traditional soil-based inoculum (s-Ri). Our findings revealed the following. (i) The propagation cycle of w-Ri inoculum is 35 days and only 23% of the 150-day cycle required for s-Ri inoculum. (ii) The spore density, viability, and purity of w-Ri inoculum are 5.25 times, 1.09 times, and 1.26 times higher, respectively, than those of s-Ri inoculum. (iii) The w-Ri inoculants demonstrate effects on enhancing rice biomass, root morphology, and photosynthesis that are consistent with those of the s-Ri inoculants, while requiring only 10% of the application rate of the s-Ri inoculants. These results provide crucial theoretical references for establishing a pure and efficient arbuscular mycorrhizal fungus propagation system and its promotion and application.IMPORTANCEThe development of a monolayer mesh hydroponic cultivation system for propagating Rhizophagus intraradices offers a significant advancement in overcoming the challenges of large-scale AMF inoculum production, which is critical for enhancing agricultural sustainability. The comparative analysis of water culture-based (w-Ri) and traditional soil-based (s-Ri) inoculum demonstrates the superior efficiency of the w-Ri system in terms of propagation speed, spore density, and inoculum quality, highlighting its potential for large-scale application in farming practices. The findings that w-Ri inoculants are equally effective in promoting plant growth while requiring only a fraction of the application rate of s-Ri inoculants underscore the potential for reducing both cost and environmental impact in agricultural inoculation practices.

Keywords: arbuscular mycorrhizal fungi; chlorophyll fluorescence parameters; inoculation effects; root morphology.

Fig 1

Colonization status at different time points during the propagation of Ri inoculum and the number of rhizosphere spores. (a) The colonization rates of two different Ri inoculum at various time points; (b) number of spores of two Ri inoculum at different times; (c) the spores of s-Ri inoculum; (d) the spores of w-Ri inoculum. The results are the means ± standard deviations of 5 values; there were no significant differences observed among the treatment groups containing the same letters, and * represents differences between different Ri inoculums at the same time. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 2

Yield of AMF propagules and the production capacity of the equipment obtained from the two production methods. (a) Fungal propagule production for both modalities; (b) the production capacity of the equipment for both modalities; (c) the spores of w-Ri inoculum; (d) the spores of s-Ri inoculum. The results are the means ± standard deviations of 5 values, and * represents differences between different Ri inoculums at the same time. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 3

Viability of AMF fungal spores, fungal purity, and fungal storage stability obtained from the two production methods. (a) Spore viability of the two Ri inoculums; (b) fungal purity of the two Ri inoculums; (c) storage stability of the two Ri inoculums. The results are the means ± standard deviations of 5 values; there were no significant differences observed among the treatment groups containing the same letters, and * represents differences between different Ri inoculums at the same time. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 4

Biomass and mycorrhizal colonization rates and colonization rates of rice seedlings under a microscope. (a) Colonization in group S (400×); (b) colonization in group W (200×); (c) mycelial structure in group S (100×); (d) mycelial structure in group W (400×); (e) colonization rate of mycorrhizae in rice seedlings at different colonization times; (f) rice plant height; (g) aboveground dry weight of rice; (h) root length of rice; and (i) root dry weight of rice. The results are the means ± standard deviations of 5 values; there were no significant differences observed among the treatment groups containing the same letters, and * represents differences between different Ri inoculums at the same time. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 5

Root morphology of rice after inoculation with different Ri inoculums. (a) Total root length; (b) root surface area; (c) root volume; (d) average root diameter; (e) photographs of rice root morphology. The results are the means ± standard deviations of 5 values; there were no significant differences observed among the treatment groups containing the same letters.

Fig 6

Photosynthetic gas exchange parameters, light response curves, and chlorophyll fluorescence parameters of rice in different treatment groups. (a) Net photosynthetic rate; (b) transpiration rate; (c) intercellular carbon dioxide concentration; (d) stomatal conductance; (e) light response curves of the S1 and W1 groups; (f) light response curves of the S2 and W2 groups. (g) Chlorophyll fluorescence parameters of the different treatments. (h) Changes in the photosynthetic parameters of the different treatments. A: net photosynthetic rate; E: transpiration rate; Ci: intercellular CO₂ concentration; GH₂O: stomatal conductance; Fv/Fm: maximum fluorescence efficiency; Fv/Fo: potential photochemical efficiency; Yield: PSII efficiency; ETR: apparent photosynthetic electron transfer rate; qL: relative component of quantum yield; qP: photochemical burst coefficient; qN: nonphotochemical burst coefficient. The results are the means ± standard deviations of 5 values; there were no significant differences observed among the treatment groups containing the same letters.

Fig 7

PCA and correlation analysis of various physiological growth parameters in rice. (a) PCA; (b) Mantel test. Rectangles are correlation heatmaps of various physiological parameters of rice seedlings, and the connecting lines of different colors indicate the differences in the significance levels of correlations between experimental conditions and physiological parameters. The blue line indicates a highly significant correlation (P < 0.01), the red line indicates a significant correlation (P < 0.05), and the gray line indicates no significant correlation (P > 0.05). *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 8

Soil propagation method and hydroponic propagation method for Ri inoculum assessment of inoculation effect.