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A qPCR assay that specifically quantifies Tricholoma matsutake biomass in natural soil

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Abstract

Tricholoma matsutake is an ectomycorrhizal basidiomycete that produces prized, yet uncultivable, “matsutake” mushrooms along densely developed mycelia, called “shiro,” in the rhizosphere of coniferous forests. Pinus densiflora is a major host of this fungus in Japan. Measuring T. matsutake biomass in soil allows us to determine the kinetics of fungal growth before and after fruiting, which is useful for analyzing the conditions of the shiro and its surrounding mycorrhizosphere, predicting fruiting timing, and managing forests to obtain better crop yields. Here, we document a novel method to quantify T. matsutake mycelia in soil by quantifying a single-copy DNA element that is uniquely conserved within T. matsutake but is absent from other fungal species, including close relatives and a wide range of ectomycorrhizal associates of P. densiflora. The targeted DNA region was amplified quantitatively in cultured mycelia that were mixed with other fungal species and soil, as well as in an in vitro co-culture system with P. densiflora seedlings. Using this method, we quantified T. matsutake mycelia not only from shiro in natural environments but also from the surrounding soil in which T. matsutake mycelia could not be observed by visual examination or distinguished by other means. It was demonstrated that the core of the shiro and its underlying area in the B horizon are predominantly composed of fungal mycelia. The fungal mass in the A or A0 horizon was much lower, although many white mycelia were observed at the A horizon. Additionally, the rhizospheric fungal biomass peaked during the fruiting season.

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Acknowledgments

This study was conducted under a “Technology Development for Optimizing Forest Resource Utilization” grant from the Ministry of Agriculture, Forestry, and Fishery of Japan. We also acknowledge financial support from the Forestry and Forest Products Research Institute (FFPRI, Tsukuba, Japan) and the FFPRI Encouragement Model in Support of Researchers with Family Responsibilities.

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Authors and Affiliations

  1. Department of Applied Microbiology and Mushroom Science, Forestry and Forest Products Research Institute, 1 Matsunosato, Tsukuba, Ibaraki, 305-8687, Japan

    Muneyoshi Yamaguchi, Hitoshi Neda, Tomoko Shimokawa & Hitoshi Murata

  2. Iwate Prefectural Forestry Technology Center, Kemuyama, Yahaba, Iwate, 028-3623, Japan

    Maki Narimatsu

  3. Kyoto Prefectural Forestry Technology Center, 1 Tsuchiya Honjo, Kyotanba, Kyoto, 629-1121, Japan

    Toru Fujita

  4. Nara Forest Research Institute, Takatori, Nara, 635-0133, Japan

    Masataka Kawai

  5. Ibaraki Prefectural Forestry Research Institute, Naka, Ibaraki, 311-0122, Japan

    Hisayasu Kobayashi

  6. Shiga Forest Research Center, Yasu, Shiga, 520-2321, Japan

    Akira Ohta

  7. Faculty of Agriculture, Shinshu University, Minami-minowa, Nagano, 399-4598, Japan

    Akiyoshi Yamada

  8. Graduate School of Agricultural and Life Sciences, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo, 113-8657, Japan

    Norihisa Matsushita

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  1. Muneyoshi Yamaguchi
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  2. Maki Narimatsu
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  3. Toru Fujita
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  4. Masataka Kawai
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  5. Hisayasu Kobayashi
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  6. Akira Ohta
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  7. Akiyoshi Yamada
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  8. Norihisa Matsushita
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  9. Hitoshi Neda
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  10. Tomoko Shimokawa
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  11. Hitoshi Murata
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Correspondence to Muneyoshi Yamaguchi.

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Funding

This study was funded by the Ministry of Agriculture, Forestry, and Fishery of Japan. This study was also funded by the Forestry and Forest Products Research Institute (FFPRI, Tsukuba, Japan) and the FFPRI Encouragement Model in Support of Researchers with Family Responsibilities.

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The authors declare that they have no conflict of interest.

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Yamaguchi, M., Narimatsu, M., Fujita, T. et al. A qPCR assay that specifically quantifies Tricholoma matsutake biomass in natural soil. Mycorrhiza 26, 847–861 (2016). https://doi.org/10.1007/s00572-016-0718-z

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  • DOI: https://doi.org/10.1007/s00572-016-0718-z

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