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Isolation and identification of beneficial orchid mycorrhizal fungi in Bletilla striata (Thunb.) Rchb.f.(Orchidaceae).

ABSTRACT: Orchid mycorrhizal fungi (OMF) are essential for orchids growth. Bletilla striata (Thunb.) Rchb.f.(Orchidaceae) has high ornamental and medicinal value. Beneficial OMF isolation is crucial to improve the survival rate of B. striata tissue culture and transplanting. In this study, we isolated and identified the beneficial OMF in B. striata from the roots of sterilized wild B. striata seedlings by culturing in four different mediums. The germination states of B. striata seeds inoculated with diverse OMF were classified and calculated. Fresh and dry weight increments of B. striata seedlings inoculated with diverse OMF were recorded after 90 d of culturing on 1/2 MS medium. ITS sequences of beneficial fungi were amplified by PCR and taxonomically identified using BLAST against the GenBank nucleotide database. Ten kinds of OMF strains were isolated from B. striata and named R1 to R10. R6 significantly promoted B. striata seeds germination (p < .01). R3 and R6 significantly promoted both the fresh and dry weight increments of B. striata seedlings (p < .05). The ITS sequence of R6 was most similar to the sequence of Serendipita. R3 was identified as Schizothecium fimbriatum by 100% ITS identity. R6 and R3 were beneficial OMF in B. striata.

SUBMITTER: Xi G

PROVIDER: S-EPMC7671041 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:In nature, orchid plants depend completely on symbiotic fungi for their nutrition at the germination and the subsequent seedling (protocorm) stages. However, only limited quantitative methods for evaluating the orchid-fungus interactions at the protocorm stage are currently available, which greatly constrains our understanding of the symbiosis. Here, we aimed to improve and integrate quantitative evaluations of the growth and fungal colonization in the protocorms of a terrestrial orchid, Blettila striata, growing on a plate medium. RESULTS:We achieved both symbiotic and asymbiotic germinations for the terrestrial orchid B. striata. The protocorms produced by the two germination methods grew almost synchronously for the first three weeks. At week four, however, the length was significantly lower in the symbiotic protocorms. Interestingly, the dry weight of symbiotic protocorms did not significantly change during the growth period, which implies that there was only limited transfer of carbon compounds from the fungus to the protocorms in this relationship. Next, to evaluate the orchid-fungus interactions, we developed an ink-staining method to observe the hyphal coils in protocorms without preparing thin sections. Crushing the protocorm under the coverglass enables us to observe all hyphal coils in the protocorms with high resolution. For this observation, we established a criterion to categorize the stages of hyphal coils, depending on development and degradation. By counting the symbiotic cells within each stage, it was possible to quantitatively evaluate the orchid-fungus symbiosis. CONCLUSIONS:We describe a method for quantitative evaluation of orchid-fungus symbiosis by integrating the measurements of plant growth and fungal colonization. The current study revealed that although fungal colonization was observed in the symbiotic protocorms, the weight of the protocorm did not significantly increase, which is probably due to the incompatibility of the fungus in this symbiosis. These results suggest that fungal colonization and nutrition transfer can be differentially regulated in the symbiosis. The evaluation methods developed in this study can be used to study various quantitative aspects of the orchid-fungus symbiosis.

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Isolation and identification of beneficial orchid mycorrhizal fungi in Bletilla striata (Thunb.) Rchb.f.(Orchidaceae).

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