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Article

Differences in Soil Microflora between the Two Chinese Geographical Indication Products of “Tricholoma matsutake Shangri-la” and “T. matsutake Nanhua”

by
Chunxin Yao
1,†,
Ping Yu
1,†,
Jisheng Yang
2,
Jiaxun Liu
3,
Zhengquan Zi
4,
Defen Li
5,
Mingtai Liang
3 and
Guoting Tian
1,*
1
Biotechnology and Genetic Germplasm Resources Research Institute, Yunnan Academy of Agricultural Sciences, Kunming 650205, China
2
The Green Food Development Center of Diqing Tibetan Autonomous Prefecture, Diqing 674499, China
3
Horticultural Research Institute, Yunnan Academy of Agricultural Sciences, Kunming 650205, China
4
Nanhua County Forestry and Grassland Bureau, Chuxiong 675200, China
5
Fungus Industry Technology Research Institute of Chuxiong Yi Autonomous Prefecture, Chuxiong 675200, China
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
Agronomy 2024, 14(4), 792; https://doi.org/10.3390/agronomy14040792
Submission received: 15 March 2024 / Revised: 1 April 2024 / Accepted: 9 April 2024 / Published: 11 April 2024
(This article belongs to the Special Issue Edible and Medicinal Fungi in Sustainable Agricultural Production: Technology and Applications)
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Versions Notes

Abstract

:
Tricholoma matsutake is a kind of ectomycorrhizal fungus. In addition to its vital influence on symbiotic plants, the impact of the soil microbial community on the growth and development of T. matsutake has been garnering attention. To clarify the differences in soil microflora between “T. matsutake Shangri-la” and “T. matsutake Nanhua”, and the effects of soil bacteria and fungi on the growth and development of T. matsutake, this study took the soil of “T. matsutake Shangri-la” and “T. matsutake Nanhua” at different developmental stages. A total of 7694 bacterial OTUs and 2170 fungal OTUs were obtained through microbial omics based on amplicon sequencing. The results indicate that the α diversity and composition of the soil microorganisms in the T. matsutake Shangri-la shiro were higher than those of the T. matsutake Nanhua. It is affected by species, geographical locations, and the growth period of matsutake. Matsutake mycelia also recruit certain types of bacteria and fungi in the stage of fruiting body development. Both bacteria and fungi positively and negatively regulate the fruiting body development of matsutake mushrooms. This study will provide a basis for the semi-artificial cultivation of matsutake.

1. Introduction

T. matsutake (S. Ito and S. Imai) Singer is a fungus belonging to the Tricholoma family and the Tricholoma genus, also known as pine fungus and peeling fungus [1]. It was listed in the “Redlist of China’s Biodiversity–Macrofungi” [2,3] as a precious and protected wild edible fungus that is widely distributed in Yunnan, China, with a high economic value [4,5]. The geographical indication protection products “T. matsutake Shangri-la” and “T. matsutake Nanhua” have been approved by the General Administration of Quality Supervision, Inspection, and Quarantine of China as two international business cards with high plateau characteristics. These fungi are also famous for their bright appearance, strong body, thick mushroom meat, tender meat, and rich taste. As raw food, they are crisp, tender, fragrant, and sweet. When cooked, they have the characteristics of toughness, lubrication, and refreshment. Furthermore, they are high in potassium and low in sodium, rich in a variety of essential amino acids for the human body, and rich in nutritional value, which are well-known facts at home and abroad [6,7].
T. matsutake Shangri-la is primarily distributed in the Diqing Prefecture, while T. matsutake Nanhua is distributed in the Chuxiong Prefecture. Both Matsutake form a unique symbiotic relationship with pine, fir, oak, and other tree species, which are widely distributed and abundant in the aforementioned territory [8,9]. Recently, due to the influence of various factors, it has been challenging to realize sustainable development and utilization of resources. T. matsutake, as a symbiotic mycorrhizal fungus with plants, still has unclear growth mechanisms under natural conditions. It grows slowly in pure culture and has harsh requirements for environmental and biological factors. Thus, artificial cultivation has not been achieved [10,11,12]. At present, the research on production mainly focuses on the conservation of the original habitats, mycelium fermentation, ectomycorrhizal synthesis, etc. [13,14,15,16,17]. Moreover, the technology of matsutake conservation and propagation remains in the aspects of forest land management and protection, and there is little research on micro-ecosystems, which makes it difficult to achieve breakthroughs in artificial intervention technology [18,19].
Matsutake mycelia form mycorrhiza underground with host plant roots and form mycelium aggregates with surrounding soil particles, called “shiro” [20]. Microorganisms in the shiro play an essential role in the growth and development of matsutake mushrooms [21]. Previous studies have also shown that compared with non-shiro (the bulk soil which does not have matsutake fruiting bodies), shiro have lower fungal diversity [22,23,24,25] due to the influence of matsutake-dominant species. Additionally, there is no difference in bacterial diversity [26]. Physical and chemical properties, including pH, organic matter, and N, P, and K element contents also indirectly affect the structure of the microbial community in the shiro [27,28,29].
In this study, we collected the shiro soil in the fruit body development stage, the non-shiro soil away from its co-growing trees, and the old shiro soil of the harvested matsutake. High-throughput sequencing technology was used to analyze the changes in soil microbial flora structures, evaluate the diversity of soil microflora, and analyze its relationships with soil physicochemical chemistry. It has a certain significance in guiding the artificial interventions of matsutake conservation and propagation.

2. Materials and Methods

2.1. Collection of Soil Samples

Jidi Village, Jiantang Town, Shangri-la City, Diqing Prefecture (99°65′39″ E, 28°02′87″ N) is located at an altitude of 3355.2 ± 99.6 m, with an average annual temperature of 5.0 °C and an average yearly rainfall of 1100.1 mm. The vegetation type is a mixed forest of Yunnan pine, alpine pine, and gray-backed oak. Malonghe Village, Wujie Town, Nanhua County, Chuxiong Prefecture (100°99′11″ E, 25°16′45″ N) is located at 2532.1 ± 124.1 m, with an average annual temperature of 17.1 °C and an average yearly rainfall of 1124.6 mm. The vegetation is mainly composed of tree species such as Pinaceae and Fagaceae.
In the two matsutake conservation areas, 8 to 9 wild matsutake shiro (A) were selected from each site and dispersed at the stage of the fruiting body development. The non-shiro soil (B) was taken as the control at a distance of 2 times from the straight lines of the co-growing tree. Additionally, the old mushroom shiro marked with matsutake harvested one year ago was labeled as the mushroom shiro (C) during the mycelium incubation period. Three replicates were set at each group of sampling sites (Table 1). According to the scattered sampling method, the surface humus and gravel were removed. The soil within a depth of 3 cm and surrounding 10 cm was collected with a sterilized spatula, and 200 g of mixed soil samples were composed and placed in sampling sealed bags. All samples were kept fresh at 4 °C and brought back to the laboratory for further processing within 24 h.

2.2. Soil Physical and Chemical Property Testing

The samples were sent to Yunnan Tongchuan Agricultural Analysis and Testing Technology Co., Ltd. (Kunming, China) for soil physical and chemical properties. The pH value was determined using the potentiometric method, organic matter was determined using the potassium dichromate volumetric method, the available nitrogen was determined with the alkali diffusion method, the available phosphorus was determined via the extraction of 0.5 mol·L−1 of NaHCO3 and the molybdenum anticolorimetric method, and the available potassium was determined via the extraction of 1 mol·L−1 of NH4OAc and flame spectrophotometry. The available zinc, manganese, and copper values were determined using the diethylenetriamine pentaacetic acid (DTPA) extraction method [30].

2.3. Microbial Diversity Sequencing

The Mag Pure Soil DNA LQ Kit (Magen Biotech, Guangzhou, China) was used to extract total genomic DNA from soil samples, diluted with sterile water to 1 ng/μL. It was then stored at −80 °C. The PCR amplification of the soil microbial DNA was performed using specific primers with barcodes. The primers were 343F-798R for the 16S V4-V5 region [31]; the primers in the ITS region were 1737F-2043R [32]. PCR reaction system and program (30 μL): 2 × Gflex PCR Buffer 15 μL, Tks Gflex DNA Polymerase (1.25 U/μL) 0.6 μL, Primer (5 μM) 1 μL, gDNA 1 μL (50 ng); H2O was replenished to 30 μL. Reaction procedure: pre-denaturation at 94 °C for 5 min, and 26 cycles (including 94 °C for 30 s, 56 °C for 30 s, and 72 °C for 20 s) at 72 °C for 5 min. PCR products were detected with electrophoresis and purified using Qubit dsDNA Assay Kit (Life Technologies, Carlsbad, CA, USA) magnetic beads. Following purification, a second round of PCR amplification was performed, followed by further detection and purification. Furthermore, Qubit quantification was performed on the PCR products. According to the concentration of PCR products, equal amounts of samples were mixed and sent to the Illumina MiSeq high-throughput sequencing platform of the Shanghai Majorbio Bio-pharm Technology Co., Ltd. (Shanghai, China) for sequencing.

2.4. Data Statistics and Analysis

SPSS 19.0 statistical software was used to analyze and process the test data. A t-test was used for comparison between groups; p < 0.05 meant a significant difference, and p < 0.01 meant an extremely significant difference. Bioinformatic analysis: Illumina MiSeq data was in the fastq format, Cutadapt software was used to cut raw data, and quality control analysis such as filtering, noise reduction, and splicing was performed on DADA 2 [33]. ASVs’ abundance of data and representative sequences based on sequencing were obtained. QIIME 2 software [34] was used to compare and annotate the database, the Silva (version 138) database was used for 16S, and the Unite database was used for ITS. The statistical analysis of microbial community structure distribution, flora Alpha diversity analysis, Beta diversity analysis, and microbial multivariate statistical analysis were also carried out. Furthermore, we used FAPROTAX 1.1 [35] and FUNGuild 1.0 [36] databases to predict the function of soil bacteria and fungi, respectively. At the genus level, the sparse correlation of component data (SparCC) network analysis was performed for microorganisms with a relative abundance greater than 0.1% [37]. Moreover, we used the Gephi 0.9.6 version to visualize the network [38].

3. Results

3.1. Comparison of Soil Physical and Chemical Properties

The results demonstrated that both matsutake shiro soil and non-shiro soil were classified as loamy clay (LC), with moderate silt (37~44%) and weakly acidic pH (4.4~5.5). The results of soil physicochemical analysis and comparison are shown in Table 2. As shown, the pH value of the T. matsutake Nanhua habitat increases with the growth and development of matsutake, while the T. matsutake Shangri-la habitat saw the opposite effect. Notably, the contents of soil organic matter and nitrogen in the matsutake habitat first increased and then decreased with the growth and development of matsutake. Potassium increased with the growth and development of matsutake and reached the highest at the fruiting body development stage. It is worth noting that the content of organic matter in T. matsutake Shangri-la shiro soil was significantly lower than that of T. matsutake Nanhua shiro soil, while the contents of Mn and Ca were significantly higher than those of T. matsutake Nanhua shiro soil. The high contents of potassium (K), manganese (Mn), calcium (Ca), and other ash elements in the soil indicate that the nutrient elements are rich and easy to neutralize the acids generated during the decomposition of organic matter, which is more conducive to the transformation of organic matter. This is consistent with the relatively high pH of T. matsutake Shangri-la shiro soil and the rich diversity of soil microorganisms.

3.2. Analysis of Soil Microbial Diversity

To determine the difference between the microbiota of “T. matsutake Shangri-la” and “T. matsutake Nanhua”, a Chinese geographical protection indication, DNA was extracted from the non-shiro soil and shiro soil of T. matsutake Shangri-la and T. matsutake Nanhua at the mycelium incubation stage and fruit body development stage, respectively. Moreover, the microbial community characteristics were determined using the HiSeq high-throughput sequencing method. The original sequences amplified by bacteria and fungi were 1,565,970 and 1,274,676, respectively. Following quality control and chimeric filtering, a total of 1,468,586 bacterial sequences and 1,197,161 fungal sequences were obtained. Through database comparison, a total of 7694 bacterial OTUs and 2170 fungal OTUs were identified, with a sequence similarity of 97%.
The Shannon and Chao diversity indices were used to analyze the α diversity of microbial communities in different soil samples (Figure 1). The results of the bacterial colony demonstrated that α diversity had a significant difference in the soil of Matsutake shiro at the fruiting body development stage, compared with the control group and the mycelium incubation stage. There was also no significant difference in the α diversity of soil bacteria in the T. matsutake Shangri-la shiro in different periods (Figure 1A,B). However, there were significant differences in the bacteria of matsutake in different geographical locations during the same period, indicating that the richness and diversity of matsutake in Shangri-la was much higher than that in Nanhua. Importantly, the results of fungi showed that there was no significant difference between the fungi in different regions at different developmental stages of matsutake (Figure 1C,D). The results also showed that the fungus was relatively stable and little affected by time and space. In both T. matsutake Shangri-la and T. matsutake Nanhua shiro, the soil microbial diversity of matsutake shiro was higher in the fruiting body development stage than in the incubation stage of mycelium, indicating that the soil microbial diversity of matsutake shiro with matsutake as the dominant species was elevated. Notably, Matsutake will recruit certain types of bacteria and fungi in the stage of mycelium to fruiting body.
To clarify the bacterial and fungal community composition differences in the various growth stages of T. matsutake Shangri-la and T. matsutake Nanhua, the β diversity of 18 samples was analyzed based on unweighted_UniFrac distance. A PCoA analysis (ANOSIM, R= 0.9366, p = 0.0010) showed that the microorganisms of T. matsutake Shangri-la and T. matsutake Nanhua were separated (Figure 1E,F). Moreover, there were differences in different periods, indicating that the variety, geographical location, and growth period of Matsutake significantly affected the composition of bacterial and fungal communities in shiro.

3.3. Soil Microbial Community Structure

Of the top 10 phyla, Proteobacteria, Acidobacteriota, Actinobacteriota, and Chloroflexi were the main bacteria in different geographical locations and different development stages. The relative content of Chloroflexi was higher in the non-shiro soil. It decreased with the appearance of mycelia and fruiting bodies, while the contents of Actinobacteriota and Bacteroidota were higher than those of non-shiro in the fruition body development stage (Figure 2A). At the genus level, Mycobacterium, Acidipila, and Acidothermus have low contents in the non-shiro soil (Figure 2C). In contrast, the content of matsutake mycelia and fruiting bodies is higher in the soil of matsutake shiro. Furthermore, the shiro in T. matsutake Nanhua is more evident than that in T. matsutake Shangri-la.
Regarding fungal taxonomic composition, Basidiomycota and Ascomycota accounted for more than 95% of each control group and shiro group. They were also in the absolute dominant phylum. Mucoromycota was low in non-shiro soils and high in shiro soils. Chytridiomycota and Glomeromycota are the opposite, with high values in non-shiro soils and low values in shiro soils (Figure 2B). At the genus level, Tricholoma was the dominant species in the growing stage of matsutake in Nanhua and Shangri-la, accounting for 30–50% (Figure 2D). Notably, these results indicated that the matsutake fruiting body development stage inhibited the growth of other fungi and was dominant in soil fungal colonies. Moreover, Cladophialophora, Penicillium, and Devriesia accounted for a higher proportion in the fruit body development stage than in the non-shiro and mycelium incubation stage. Compared with non-shiro, the number of Mortierella in T. matsutake Shangri-la shiro was significantly reduced.

3.4. Functional Predictive Analysis

FAPROTAX was used to predict the assumed functional differences of soil bacterial communities in different development stages of Shangri La Matsutake and T. matsutake Nanhua. Compared with T. matsutake Nanhua soil, the functions of nitrogen cycling and anaerobic photoautotrophy in the matsutake soil of Shangri-la were more prominent. Compared to non-shiro soil, the fruit body development stage soil dissimilatory_arsenale_reduction, arsenate_detoxification, plastic_degradation, plant_pathogen, aromatic_compound_Degradation, fermentation, and nitrate_reduction ureolysis functions are more prominent (Figure 3A). Following the Kruskal–Wallis rank sum test, FDR multiple test correction, Tukey–Kramer 0.95, and Post hoc test, nitrogen_fixation was significantly different among groups. Compared with non-shiro soil, shiro soil had a significantly lower function during the development of the fruit body.
FUNGuild performs functional classifications of fungal communities [39]. The results showed that the Ectomycorrhizal-Fungal Parasite had an absolute advantage during the development of matsutake fruits. The function of Undefined Saprotroph and Plant Pathogen was the lowest in the mycelium incubation period, with the non-shiro as the control. Moreover, the development process of matsutake showed a trend of decreasing first and then increasing. The functions of Ectomycorrhizal and Ericoid Mycorrhizal were the highest at the incubation stage of mycelium and showed a trend of first increasing and then decreasing. Furthermore, plant Saprotroph-Wood Saprotroph gradually increased, and Ectomycorrhizal-Orchid Mycorrhizal-Root Associated Biotroph gradually reduced (Figure 3B).

3.5. Features of Co-Occurrence Network

We constructed a co-occurring network of microorganisms with a relative abundance of OTUs greater than 0.1% at the genus level at different growth and development stages of matsutake in different locations. Then, we screened for microorganisms directly related to Tricholoma (Figure 4). We observed that in both Shangri-la and Nanhua, the microbial community number in the growth stage of matsutake fruitum was greater than that in the non-shiro, with non-shiro greater than that in the incubation stage of mycelium. Our results showed that soil microorganisms decreased first and then increased with the development of matsutake. The average degree, number of nodes, and edges of fruiting bodies in the development stage samples were higher than those of other samples. Notably, this indicates that the relationship between members of the network is the largest, and the interaction between the members is the closest during the development of the fruit body. In addition, the T. matsutake was directly negatively regulated by Jatrophihabitans and Conexibacter and positively and negatively regulated by other bacteria and fungi in the T. matsutake Nanhua fruiting body development stage shiro. Furthermore, Flavisohibacterhe, Reyranella, and Oidiodendron negatively regulated the T. matsutake. Thus, it is positively regulated by the bacteria Burkholderia-Caballeronia-Paraburkholderia, TM7a, and the fungus Russula in the T. matsutake Shangri-la fruiting body development stage shiro.

4. Discussion

As an ectomycorrhizal fungus symbiosis with higher plants, the formation of matsutake fruiting bodies is affected by host species, climatic conditions, soil properties, microorganisms, and other factors [29,40,41,42]. As such, high-throughput sequencing technology has been used to study the effects of matsutake soil microorganisms on the growth and development of matsutake under different forest ecologies [24,43,44,45]. However, relatively few studies have been conducted, and most studies of microbial diversity in ponds have focused on the composition of fungal communities [23,43,46]. There are also few reports on the effects of the bacterial community and the combination of the bacterial community and fungal communities on the growth and development of matsutake at different development stages [29].
Due to the scarcity of wild resources, the artificial or semi-artificial cultivation of wild edible fungi is imminent. At present, ectomycorrhizal fungi are relatively mature in truffles, which can be cultivated semi-artificially. At present, ectomycorrhizal fungi are relatively mature in Tuber indicum, which can be cultivated semi-artificially. At the moment, outplanted pine seedlings synthesizing T. matsutake ectomycorrhizae in vitro in a soil volume of 1 L were found to sustain mycorrhizal status and shiro structures for 2 y. However, the shiro did not expand in the area, and extended pine roots were colonized by native ectomycorrhizal fungi such as suilloids. The direct inoculation of cultured T. matsutake mycelia to non-ectomycorrhizal roots of adult pine trees in situ led to successful ectomycorrhization but mycorrhizal status did not persist beyond 1 y. At present, matsutake has not been cultivated artificially or semi-artificially. T. matsutake has unique mechanisms associated with shiro structures that promote their survival and growth in situ. In this study, T. matsutake Shangri-la and T. matsutake Nanhua fungi soil and non-matsutake soil with high plateau characteristics in Yunnan were used. By implementing 16S and ITS high-throughput sequencing analysis, the compositions and functional differences of bacterial and fungal communities in the shiro of two Chinese geographical indication products were studied. Additionally, the bacteria and fungi related to the growth and development of matsutake were screened, as well as their interactions. Subsequently, the relevant microorganisms will be isolated and cultured and the shiro will be treated to enhance the vitality of ectomycorrhizal matsutake and promote the shiro to produce matsutake fruit bodies. These results will provide a basis for the semi-artificial cultivation of matsutake from the perspective of microbial interaction, so as to promote the stable and sustainable development of the wild matsutake industry.
Our results demonstrated that the α diversity of T. matsutake Shangri-la was higher than that of T. matsutake Nanhua, no matter whether bacteria or fungi, or shiro or non-shiro. Notably, bacteria were more prominent. This indicates that soil microbial diversity in different regions is correlated with climate factors. Shangri-la belongs to the temperate and cold temperate monsoon climate, while Nanhua belongs to the subtropical monsoon climate. Previous studies have also shown that the soil microbial community diversity in temperate climates is higher than that in subtropical climates [47]. The microbial analysis of matsutake soil in Shangri-la showed that the fungal diversity in matsutake-dominant soil samples was lower than that in non-shiro samples, indicating that matsutake mycelia inhibited the growth of other fungi and occupied a dominant position in the soil fungal community. Importantly, this is consistent with previous investigations [22,26,43]. However, the opposite was true in T. matsutake Nanhua soil microbial analysis. Compared with the mycelium incubation stage, the number of microorganisms (bacteria and fungi) increased in the fruiting body development stage. Therefore, it is of general significance to study the growth of the fruiting body regarding the incubation period of mycelia. Moreover, the β diversity analysis showed that there were significant differences between the groups of matsutake, whether bacteria or fungi, during different growth and development periods, especially between the groups of Matsutake Nanhua. This is consistent with previous studies. Thus, these results indicate that the significant difference in microorganisms between matsutake shiro and non-shiro was not affected by geographical location or forest ecology [46].
The soil nutrition of T. matsutake Shangri-la shiro is relatively poor, and the pH value is between 5.3 and 5.6. Additionally, the soil of T. matsutake Nanhua shiro is relatively dry, loose, and more acidic (pH 4.3–4.5). Notably, the soil pH value is the main factor affecting the uniformity of the underground mycorrhizal community. According to the many years of experience of matsutake conservation area management personnel, matsutake fruiting bodies will grow out of old shiro after the mycelium incubation period, and no fruiting bodies will emerge from non-shiro. The composition of soil microbial communities is determined by many biological and abiotic factors, of which nitrogen is a crucial element controlling species composition, diversity, and productivity in many terrestrial ecosystems [48]. In this study, there was no significant difference in nitrogen content in the soil of matsutake at different stages of development in different places. Still, there were relatively more bacteria with nitrogen cycling functions in the soil of Shangri-la. Furthermore, nitrogen fixation in T. matsutake Nanhua soil and T. matsutake Shangri-la soil is significantly different among groups, and the function of nitrogen fixation in matsutake soil during fruiting body development is significantly reduced compared with non-matsutake soil.
As early as 1978, Ogawa discovered that the mycorrhiza formed by matsutake and symbiotic plants can produce antibacterial substances that inhibit the growth of specific root pathogens. The underground mycelia and mycorrhiza in the matsutake shiro can also release some substances to inhibit the growth of bacteria and actinomycetes in the area during the annual expansion process to reduce host root infection by pathogens. Interestingly, our prediction results of bacterial and fungal function showed that plant pathogen-related microorganisms were relatively low in the matsutake mycelium incubation stage but relatively high in the fruiting body development stage and higher than in non-shiro. This indicates that matsutake interacts with symbiotic plants during the mycelium incubation period to help plants fight pathogens and reduce plant energy consumption. Once the interaction is successful, matsutake pays more attention to self-growth and development and reduces the consumption of antibacterial substances to achieve a self-energy balance.
The results of microbial interaction with Tricholoma showed that Tricholoma growth and development were not only related to fungi but also to bacteria. In particular, T. matsutake Nanhua was directly negatively regulated by the bacteria Jatrophihabitans and Conexibacter and was occasionally regulated by other bacteria and fungi. Additionally, the T. matsutake Shangri-la may be directly affected by more microorganisms because of the higher diversity of microorganisms in the shiro. It was also negatively regulated by Oidiodendron and positively regulated by TM7. Furthermore, Oidiodendron is another common type of rhododendron mycorrhizal fungus [49]. Notably, TM7 bacteria suppress pathogenic bacteria, as observed in other studies [50].

5. Conclusions

The transformation of organic matter is made possible by ash elements, such as Mn and Ca, in the T. matsutake Shangri-la soil. As such, it regulates soil nutrition and microbial diversity. The species, geographical location, and growth period of matsutake significantly affected the composition of the microbial community in the shiro. Certain types of bacteria and fungi were also recruited from matsutake mycelia to fruiting bodies in the shiro. Furthermore, different bacteria and fungi positively and negatively regulated the fruiting body development of matsutake in different geographical locations.

Author Contributions

G.T. designed and supervised the experiment and was responsible for funding. C.Y. and P.Y. carried out the experiments and the manuscript. J.Y. and J.L. completed the data analysis. Z.Z., D.L. and M.L. reviewed and edited the manuscript. All authors have read and agreed to the published version of the manuscript.

Funding

This research was funded by the Yunnan Major Science and Technology Special Program (202102AE090051) and Yunnan Academician Expert Workstation (202305AF150164).

Data Availability Statement

The authors confirm that the data supporting the findings of this study are available within the article.

Conflicts of Interest

The authors declare no conflicts of interest.

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Agronomy 14 00792 g001aAgronomy 14 00792 g001b
Figure 1. α diversity and β diversity of microbial communities in T. matsutake shiro at different development stages. Alpha diversity in bacterial (A,B) and fungal (C,D) communities. All inter−group difference analyses are based on the Kruskal−Wallis rank sum test. The principal coordinate analysis of β diversity based on unweighted_UniFrac distance shows the community structure of bacteria (E) and fungi (F) at different development stages. Group difference analyses are based on ANOSIM. * 0.01 < p ≤ 0.05, ** 0.001 < p ≤ 0.01, *** p ≤ 0.001.
Figure 1. α diversity and β diversity of microbial communities in T. matsutake shiro at different development stages. Alpha diversity in bacterial (A,B) and fungal (C,D) communities. All inter−group difference analyses are based on the Kruskal−Wallis rank sum test. The principal coordinate analysis of β diversity based on unweighted_UniFrac distance shows the community structure of bacteria (E) and fungi (F) at different development stages. Group difference analyses are based on ANOSIM. * 0.01 < p ≤ 0.05, ** 0.001 < p ≤ 0.01, *** p ≤ 0.001.
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Figure 2. Relative abundance in the phylum (A,B) and genus (C,D) level of bacterial (A–C) and fungal (B–D) taxa under different developmental stages.
Figure 2. Relative abundance in the phylum (A,B) and genus (C,D) level of bacterial (A–C) and fungal (B–D) taxa under different developmental stages.
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Figure 3. Functional prediction of soil bacterial (A) and fungal (B) communities at different growth and development stages of matsutake.
Figure 3. Functional prediction of soil bacterial (A) and fungal (B) communities at different growth and development stages of matsutake.
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Figure 4. Co-occurrence network of bacterial and fungal communities and tricholoma-associated network (G,H) in the soil at different growth and development stages of matsutake. (A) Shiro in the fruiting body development stage of T. matsutake Nanhua; (B) non-shiro of T. matsutake Nanhua; (C) shiro in the mycelium incubation period of T. matsutake Nanhua; (D) shiro in the fruiting body development stage of T. matsutake Shangri-la; (E) non-shiro of T. matsutake Shangri-la; (F) shiro in the mycelium incubation period of T. matsutake Shangri-la; (G) interacting microorganism with Tricholoma during fruiting body development stage of T. matsutake Nanhua; and (H) interacting microorganism with Tricholoma during fruiting body development stage of T. matsutake Shangri-la. Green line, negative regulation; Red line, positive regulation.
Figure 4. Co-occurrence network of bacterial and fungal communities and tricholoma-associated network (G,H) in the soil at different growth and development stages of matsutake. (A) Shiro in the fruiting body development stage of T. matsutake Nanhua; (B) non-shiro of T. matsutake Nanhua; (C) shiro in the mycelium incubation period of T. matsutake Nanhua; (D) shiro in the fruiting body development stage of T. matsutake Shangri-la; (E) non-shiro of T. matsutake Shangri-la; (F) shiro in the mycelium incubation period of T. matsutake Shangri-la; (G) interacting microorganism with Tricholoma during fruiting body development stage of T. matsutake Nanhua; and (H) interacting microorganism with Tricholoma during fruiting body development stage of T. matsutake Shangri-la. Green line, negative regulation; Red line, positive regulation.
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Table 1. Soil sampling information related to T. matsutake.
Table 1. Soil sampling information related to T. matsutake.
Shangri-la CityNanhua
Shiro in the development stage of fruiting bodiesXANhA
Non-shiroXBNhB
Shiro in the hyphal incubation periodXCNhC
Table 2. Soil physicochemical analysis.
Table 2. Soil physicochemical analysis.
HabitatsNhANhBNhCXAXBXC
Soil Physicochemical
Properties
pH4.474.374.415.305.535.47
OM (g/kg)104.8393.62106.4438.1120.7240.24
N (g/kg)2.372.382.661.361.041.39
P (mg/kg)8.4019.2010.9012.221.917.36
K (mg/kg)515.00205.00207.30490.10175.7210.30
moisture (%)28.5031.1031.2023.8017.5019.70
Zn (g/kg)2.411.842.091.601.211.27
Mn (g/kg)9.643.767.6391.0490.5289.49
Cu (mg/kg)0.350.170.210.320.360.32
Mg (mg/kg)12495104162169149
Ca (mg/kg)543.00632.00576.00846.00772.00798.00
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Yao, C.; Yu, P.; Yang, J.; Liu, J.; Zi, Z.; Li, D.; Liang, M.; Tian, G. Differences in Soil Microflora between the Two Chinese Geographical Indication Products of “Tricholoma matsutake Shangri-la” and “T. matsutake Nanhua”. Agronomy 2024, 14, 792. https://doi.org/10.3390/agronomy14040792

AMA Style

Yao C, Yu P, Yang J, Liu J, Zi Z, Li D, Liang M, Tian G. Differences in Soil Microflora between the Two Chinese Geographical Indication Products of “Tricholoma matsutake Shangri-la” and “T. matsutake Nanhua”. Agronomy. 2024; 14(4):792. https://doi.org/10.3390/agronomy14040792

Chicago/Turabian Style

Yao, Chunxin, Ping Yu, Jisheng Yang, Jiaxun Liu, Zhengquan Zi, Defen Li, Mingtai Liang, and Guoting Tian. 2024. "Differences in Soil Microflora between the Two Chinese Geographical Indication Products of “Tricholoma matsutake Shangri-la” and “T. matsutake Nanhua”" Agronomy 14, no. 4: 792. https://doi.org/10.3390/agronomy14040792

APA Style

Yao, C., Yu, P., Yang, J., Liu, J., Zi, Z., Li, D., Liang, M., & Tian, G. (2024). Differences in Soil Microflora between the Two Chinese Geographical Indication Products of “Tricholoma matsutake Shangri-la” and “T. matsutake Nanhua”. Agronomy, 14(4), 792. https://doi.org/10.3390/agronomy14040792

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