**Table 1.**

**161**

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

and mycorrhizal formation. In pure culture conditions, most of the studied fungi appeared to favor ammonium N. Some species, namely *Amanita caesarea*, *Cantharellus cibarius*, *Lactarius bicolor*, *Suillus variegatus* were not able to grow

However, many of the edible ECM fungi, namely *Amanita caesarea* [103, 105], *Cantharellus cibarius* [48], *Cortinarius variecolor* [67], *Paxillus involutus*, *Suillus variegatus* [113], *Tricholoma terreum* [123], and *Tuber sinense* [124] were able to grow on the media containing organic N (protein) as the sole nitrogen. Moreover, some fungi belonging to *Lactarius* genus had limited capacity to utilize protein N [113, 114]. *Hebeloma cylindrosporum* was able to experimentally utilize a wide range of amino acids and other simple (e.g. urea) or complex (e.g. proteins)

The studied forms of N often predominate soil solution and the culturing results might be assumed to hold true in nature. However, it is worth mentioning is that the optimal nitrogen in the mycelium culture does not necessarily reflect the nitrogen preference of the ECM fungus under natural conditions because environmental factors affect. This was shown with *H. cylindrosporum* growing in nature. Wild dikaryotic strains of *H. cylindrosporum* isolated from two different habitat types had

Cultivation of edible ECM mushrooms has been successful in cases of two truffles *Tuber melanosporum* Vittad. and *Tuber aestivum* Vittad. They are cultivated commercially around the world [125]. In addition, some success has been achieved with *Lactarius deliciosus* [126, 127] and *Boletus edulis* [128]. Regarding truffle

production, it has been suggested that most soils contain enough N to maintain both fungal and tree growth [125]. Similarly, *Lactarius deliciosus* was cultivated experimentally in forest soil, which was observed to meet the demands for fruitbody formation [126]. It has also been demonstrated that the nutritional properties of soil and the forestry history the natural development of ECM mushrooms in forest ecosystems [129]. A productive and diverse ECM mushroom community resembling natural communities developed when abandoned farmland in Mediterranean

In summary, productive ECM community can grow in natural soils. However,

**4.2 Nitrogen source requirements for** *Tricholoma matsutake* **mycelial culture** 

*Tricholoma matsutake* is among the most economically valuable mushrooms in the world. Its taxonomy, distribution, ecology, physiology, and cultivation has been studied widely [130]. Here, we summarize the key results linking matsutake

ecological characteristics and nutrient requirements focusing on nitrogen. Matsutake colonizes the roots of its host trees via an ECM association (**Figure 1a** and **b**). It develops an extraradical mycelium in the rhizosphere and in the surrounding soil area. This can be seen as a white rhizosphere area and it corresponds to the mycelium-soil aggregated zone, called a shiro [131] (**Figure 1c** and **d**). Matsutake shiro grows in the form of a concentric or horseshoe-like circle, depending on the rhizosphere conditions, around the host plant at the rate of approximately 10–15 cm per year [131, 132]. The production of matsutake mushrooms changes periodically. Based on our field observations, the part of mycorrhizal root tips is degraded prior to matsutake fruiting. The extraradical mycelium might grow towards new roots and colonizing. Such a hyphal growth strategy indicates that matsutake symbiosis may often need to be renewed and form new mycorrhizas to acquire nutrients (data not published). Among the mycorrhizal associations, such

the challenges faced in artificial cultivation has not been solved.

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

nitrate as the sole N source [48, 103, 113].

compounds [6, 59].

different N preferences [6].

dry area was forested with *Pinus* sp.

**and mycorrhizal synthesis**

 *Fungal growth, symbiosis and fruitbody formation observed using different nitrogen sources in edible ectomycorrhizal fungi in combination with the information of hydrophobicity, exploration type and* δ*15N of the fruitbodies.* *Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

and mycorrhizal formation. In pure culture conditions, most of the studied fungi appeared to favor ammonium N. Some species, namely *Amanita caesarea*, *Cantharellus cibarius*, *Lactarius bicolor*, *Suillus variegatus* were not able to grow nitrate as the sole N source [48, 103, 113].

However, many of the edible ECM fungi, namely *Amanita caesarea* [103, 105], *Cantharellus cibarius* [48], *Cortinarius variecolor* [67], *Paxillus involutus*, *Suillus variegatus* [113], *Tricholoma terreum* [123], and *Tuber sinense* [124] were able to grow on the media containing organic N (protein) as the sole nitrogen. Moreover, some fungi belonging to *Lactarius* genus had limited capacity to utilize protein N [113, 114]. *Hebeloma cylindrosporum* was able to experimentally utilize a wide range of amino acids and other simple (e.g. urea) or complex (e.g. proteins) compounds [6, 59].

The studied forms of N often predominate soil solution and the culturing results might be assumed to hold true in nature. However, it is worth mentioning is that the optimal nitrogen in the mycelium culture does not necessarily reflect the nitrogen preference of the ECM fungus under natural conditions because environmental factors affect. This was shown with *H. cylindrosporum* growing in nature. Wild dikaryotic strains of *H. cylindrosporum* isolated from two different habitat types had different N preferences [6].

Cultivation of edible ECM mushrooms has been successful in cases of two truffles *Tuber melanosporum* Vittad. and *Tuber aestivum* Vittad. They are cultivated commercially around the world [125]. In addition, some success has been achieved with *Lactarius deliciosus* [126, 127] and *Boletus edulis* [128]. Regarding truffle production, it has been suggested that most soils contain enough N to maintain both fungal and tree growth [125]. Similarly, *Lactarius deliciosus* was cultivated experimentally in forest soil, which was observed to meet the demands for fruitbody formation [126]. It has also been demonstrated that the nutritional properties of soil and the forestry history the natural development of ECM mushrooms in forest ecosystems [129]. A productive and diverse ECM mushroom community resembling natural communities developed when abandoned farmland in Mediterranean dry area was forested with *Pinus* sp.

In summary, productive ECM community can grow in natural soils. However, the challenges faced in artificial cultivation has not been solved.

#### **4.2 Nitrogen source requirements for** *Tricholoma matsutake* **mycelial culture and mycorrhizal synthesis**

*Tricholoma matsutake* is among the most economically valuable mushrooms in the world. Its taxonomy, distribution, ecology, physiology, and cultivation has been studied widely [130]. Here, we summarize the key results linking matsutake ecological characteristics and nutrient requirements focusing on nitrogen.

Matsutake colonizes the roots of its host trees via an ECM association (**Figure 1a** and **b**). It develops an extraradical mycelium in the rhizosphere and in the surrounding soil area. This can be seen as a white rhizosphere area and it corresponds to the mycelium-soil aggregated zone, called a shiro [131] (**Figure 1c** and **d**). Matsutake shiro grows in the form of a concentric or horseshoe-like circle, depending on the rhizosphere conditions, around the host plant at the rate of approximately 10–15 cm per year [131, 132]. The production of matsutake mushrooms changes periodically. Based on our field observations, the part of mycorrhizal root tips is degraded prior to matsutake fruiting. The extraradical mycelium might grow towards new roots and colonizing. Such a hyphal growth strategy indicates that matsutake symbiosis may often need to be renewed and form new mycorrhizas to acquire nutrients (data not published). Among the mycorrhizal associations, such

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

**160**

**ECM fungi**

*S. lutus* *S. variegatus* **Tricholoma** *T. imbricatum*

NH4

+ or NO3

orgN (gained

better grwoth in

iorgN)

*T. bakamatsutake*

*T. matsutake*

NH4

+ pluse orgN

orgN orgN (sustaining

symbiotic

relationship)

*T. terreum*

**Tuber** *T. sinense*

*\* NH4*

**Table 1.**

*type and* δ*15N of the fruitbodies.*

*+, ammonum nitrogen; NO3*

*−, nitrate nitrogen; orgN, organic nitrogen. Ho, hydrophobic; Hi, hydrophibic.*

*Fungal growth, symbiosis and fruitbody formation observed using different nitrogen sources in edible ectomycorrhizal fungi in combination with the information of hydrophobicity, exploration* 

orgN( gained

better growth)

orgN (gained

better growth)

NH4

+ or orgN

[118]

[119] [121]

[122]

[123]

Hi

[124]

Short

15.1 ± 0.6 (9)

[102]

16.8 ± 2.3 (15)

[120]

(poor on NO3

−

)

−

or

NH4

+ (poor on

orgN)

orgN

**Mycelium** 

**Mycorrhization**

**Fruitbody formation**

**Ref** [112]

[113] Ho

[117]

Medium-fringe

9.3 ± 0.6 (35)

[102]

**Hydrophobicity**

**Exploration** 

**δ15N (‰)** 

**Ref.**

**(Mean ± SD) (n)**

5.7 ± 1.1 (4) cap

[111]

**type**

**growth**

#### **Figure 1.**

*The ectomycorrhizal edible mushroom—*Tricholoma matsutake *(a) the root of* Pinus sylvestris *seedling is colonized by* T. matsutake *fungal mycelium, forms mycorrhizas; (b) the transverse section of ECM root showing the Hartig net (hn) development within the cortex; (c) matsutake mushrooms form in a conifer mixed forest in southern of Finland; (d) the matsutake shiro (arrow) after the mushrooms be harvested (photos were taken by Lu Min Vaario).*

a phenomenon does not seem to be rare. Hortal and colleagues [133] found that the plant had the ability to limit the root tip colonization of the least cooperative symbiont, and therefore, influence the outcome of ECM fungi competition. Such reduction in colonization did not result in a reduction in carbon allocation to the fungus providing the lowest amount of nitrogen.

It is worth noting that decayed mycorrhizal roots together with mycelium-soil aggregated zone might be important organic nutrient sources for matsutake. Recently, the natural abundance of isotopes data showed a very high δ15N value in *T. matsutake* fruitbodies, which were sampled from Finland and Japan [120]. Matsutake usually grow at B layer of mineral soil [131], such taxa obtain their N could explain for high δ 15N values (see review [102]). More importantly, the high δ 15N value in matsutake is an indicator of organic N uptake from soil because the great variation of 15N content observed among ECM taxa has been reported to be related to the differences in organic N utilization [111]. In addition, a literature study shows that mycorrhizal taxa with proteolytic activities generally show high δ 15N values [67]. Therefore, we conclude that matsutake has a greater proteolytic activity to digest chemically complex 15N-enriched organic matter in soil during matsutake fruitbody development.

In addition to proteases, matsutake produces organic matter degradation enzymes such as acid proteinase [134, 135] and β-glucosidase [136]. Relatively high enzyme activities, β-glucosidase and xylosidase, were detected from matsutake cultures in vitro and in shiro soil [137, 138]. The genome of *T. matsutake* encodes

**163**

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

two GH7 cellobiohydrolases [88], which is in agreement with its known facultative saprotrophic activity [136, 138]. However, no further evidence of any strong saprotrophic characteristics of matsutake was found. It could be speculated that these ECM fungi produce certain levels of carbohydrase, not to fully degrade organic matter to access C but N. Kawai and Abe [121] reported that dried beer yeast, corn steep liquor, casein hydrolysate, and polypeptone were good N sources for matsutake mycelium culture whereas nitrate was not. Dry beer yeast (Ebios, Asahi Beer Inc., Tokyo, Japan), as the sole N source, showed promising matsutake mycelium growth and as well mycorrhizal formation [139] (personal communication with Dr.

Several agar media such as MMN, MNC, Hamada containing both inorganic and organic N are widely used to culture the mycelium of *T. matsutake* [119, 140]. However, the question whether matsutake prefers organic nitrogen is worth of considering. Usually, more inorganic N than organic N is present in the soil top layer. Some studies suggested that increased N deposition could reduce fruitbody production [141]. Nohrstedt [142] reported a 30% decrease in sporocarp production by *Cantharellus cibarius* in a central Swedish pine forest after the application of 150 kg N ha−1 ammonium nitrate. The presence of nitrate ions has been shown to have negative effects on the development of some ECM fungi both in vitro and in soil [143, 144]. Removal of the litter layer has been considered an important method to improve the productivity of matsutake in many Asian countries [145]. It has also been shown that the removal of the upper organic soil layers of the forest floor can improve the sporocarp production of some other ECM fungi [146, 147]. The explanation might be that competition with other microbes diminishes. Litter and organic soil provide carbon and nutrients for microbes, especially for saprotrophic

Cultivation of ectomycorrhizal mushrooms is still facing many challenges. Although some species of ECM fungi can form the primordium of fruiting bodies on several media, they usually do not develop further into mature fruiting bodies. So far, the most successful efforts have been carried out with the mycorrhizal plants growing in soil. Soil nutrients and soil microbial communities together with climatic factors have shown to affect significantly the persistence of ectomycorrhizas in outplanted inoculated plants, and further, the successful fruiting. The observed suppression of many mycorrhizal mushrooms has been linked to indirect effects of air pollution, in particular to increases in nitrogen deposition accumulating into litter and humus [149, 150]. A thorough understanding of the ecological and environmental factors regulating the ECM fungal species is a prerequisite for

Ectomycorrhizal fungi colonize the roots of their host plants and improve plants' access to nutrients, especially nitrogen. In exchange, host plants deliver a significant portion of their photosynthesized carbon to the ECM fungi. However, we need more accurate understanding of the ECM fungi mediated C and N movement within forest ecosystems. ECM fungi may follow a similar pattern with the amount of C delivered being related to the amount of N sourced by the fungus [77, 151], although this is still controversial [133]. Production of ECM mushrooms do need a balanced nutrient either assimilating by ECM fungi or by other soil microbial.

It has been suggested that the growth of ECM fungi and the formation of mycorrhizas are promoted by certain mycorrhizosphere bacteria, termed 'mycorrhizal helper bacteria' [152]. Some mycorrhizal fungi-associated bacteria are also known to fix nitrogen [153, 154]. However, there is still no evidence that the fungus would

fungi that would compete with *T. matsutake* in the shiro [148].

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

A. Yamada).

**4.3 Research prospects**

their cultivation.

#### *Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

two GH7 cellobiohydrolases [88], which is in agreement with its known facultative saprotrophic activity [136, 138]. However, no further evidence of any strong saprotrophic characteristics of matsutake was found. It could be speculated that these ECM fungi produce certain levels of carbohydrase, not to fully degrade organic matter to access C but N. Kawai and Abe [121] reported that dried beer yeast, corn steep liquor, casein hydrolysate, and polypeptone were good N sources for matsutake mycelium culture whereas nitrate was not. Dry beer yeast (Ebios, Asahi Beer Inc., Tokyo, Japan), as the sole N source, showed promising matsutake mycelium growth and as well mycorrhizal formation [139] (personal communication with Dr. A. Yamada).

Several agar media such as MMN, MNC, Hamada containing both inorganic and organic N are widely used to culture the mycelium of *T. matsutake* [119, 140]. However, the question whether matsutake prefers organic nitrogen is worth of considering. Usually, more inorganic N than organic N is present in the soil top layer. Some studies suggested that increased N deposition could reduce fruitbody production [141]. Nohrstedt [142] reported a 30% decrease in sporocarp production by *Cantharellus cibarius* in a central Swedish pine forest after the application of 150 kg N ha−1 ammonium nitrate. The presence of nitrate ions has been shown to have negative effects on the development of some ECM fungi both in vitro and in soil [143, 144]. Removal of the litter layer has been considered an important method to improve the productivity of matsutake in many Asian countries [145]. It has also been shown that the removal of the upper organic soil layers of the forest floor can improve the sporocarp production of some other ECM fungi [146, 147]. The explanation might be that competition with other microbes diminishes. Litter and organic soil provide carbon and nutrients for microbes, especially for saprotrophic fungi that would compete with *T. matsutake* in the shiro [148].

#### **4.3 Research prospects**

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

a phenomenon does not seem to be rare. Hortal and colleagues [133] found that the plant had the ability to limit the root tip colonization of the least cooperative symbiont, and therefore, influence the outcome of ECM fungi competition. Such reduction in colonization did not result in a reduction in carbon allocation to the

*The ectomycorrhizal edible mushroom—*Tricholoma matsutake *(a) the root of* Pinus sylvestris *seedling is colonized by* T. matsutake *fungal mycelium, forms mycorrhizas; (b) the transverse section of ECM root showing the Hartig net (hn) development within the cortex; (c) matsutake mushrooms form in a conifer mixed forest in southern of Finland; (d) the matsutake shiro (arrow) after the mushrooms be harvested (photos were* 

It is worth noting that decayed mycorrhizal roots together with mycelium-soil

15N value in matsutake is an indicator of organic N uptake from soil because the great variation of 15N content observed among ECM taxa has been reported to be related to the differences in organic N utilization [111]. In addition, a literature study shows that mycorrhizal taxa with proteolytic activities generally show high

15N values [67]. Therefore, we conclude that matsutake has a greater proteolytic activity to digest chemically complex 15N-enriched organic matter in soil during

In addition to proteases, matsutake produces organic matter degradation enzymes such as acid proteinase [134, 135] and β-glucosidase [136]. Relatively high enzyme activities, β-glucosidase and xylosidase, were detected from matsutake cultures in vitro and in shiro soil [137, 138]. The genome of *T. matsutake* encodes

15N values (see review [102]). More importantly, the high

aggregated zone might be important organic nutrient sources for matsutake. Recently, the natural abundance of isotopes data showed a very high δ15N value in *T. matsutake* fruitbodies, which were sampled from Finland and Japan [120]. Matsutake usually grow at B layer of mineral soil [131], such taxa obtain their N

fungus providing the lowest amount of nitrogen.

could explain for high δ

matsutake fruitbody development.

**162**

δ

**Figure 1.**

*taken by Lu Min Vaario).*

δ

Cultivation of ectomycorrhizal mushrooms is still facing many challenges. Although some species of ECM fungi can form the primordium of fruiting bodies on several media, they usually do not develop further into mature fruiting bodies. So far, the most successful efforts have been carried out with the mycorrhizal plants growing in soil. Soil nutrients and soil microbial communities together with climatic factors have shown to affect significantly the persistence of ectomycorrhizas in outplanted inoculated plants, and further, the successful fruiting. The observed suppression of many mycorrhizal mushrooms has been linked to indirect effects of air pollution, in particular to increases in nitrogen deposition accumulating into litter and humus [149, 150]. A thorough understanding of the ecological and environmental factors regulating the ECM fungal species is a prerequisite for their cultivation.

Ectomycorrhizal fungi colonize the roots of their host plants and improve plants' access to nutrients, especially nitrogen. In exchange, host plants deliver a significant portion of their photosynthesized carbon to the ECM fungi. However, we need more accurate understanding of the ECM fungi mediated C and N movement within forest ecosystems. ECM fungi may follow a similar pattern with the amount of C delivered being related to the amount of N sourced by the fungus [77, 151], although this is still controversial [133]. Production of ECM mushrooms do need a balanced nutrient either assimilating by ECM fungi or by other soil microbial.

It has been suggested that the growth of ECM fungi and the formation of mycorrhizas are promoted by certain mycorrhizosphere bacteria, termed 'mycorrhizal helper bacteria' [152]. Some mycorrhizal fungi-associated bacteria are also known to fix nitrogen [153, 154]. However, there is still no evidence that the fungus would

directly benefit from its associated bacteria. Sporocarps of *Cantharellus cibarius* contain large amount of bacteria, in particular fluorescent *Pseudomonas* [155]. Some species of bacteria such as *Streptomyces* spp., *Paenibacillus* spp. and *Bacillales* spp. were isolated from the mycorrhizal root tips and fruitbodies of *T. matsutake* as well [156–158]. Otherwise, the information about mycorrhizas-associated bacteria and their effect on the nutrient uptake of ECM fungi is limited. These studies, however, hint that the production of ectomycorrhizal mushrooms may require teamwork to obtain enough nutrients from the environment.

In conclusion, ECM fungi play an important role in the nutrient cycle of forest ecosystems, especially on mediating C and N movement. A better understanding of the nitrogen status of the habitat of ECM fungi, nutrients movements within the ecosystems, as well as the ECM fungal hyphal structures should be the first step for cultivation of ECM edible mushrooms. The methodological advances in these areas in combination with forest management may allow the successful establishment of commercial plantations and production of edible ECM mushrooms in forests.

### **Author details**

Lu-Min Vaario1,2\* and Norihisa Matsushita2

1 Department of Forest Sciences, University of Helsinki, PO Box 27, FI-00014 Helsinki, Finland

2 Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo, Japan

\*Address all correspondence to: lu-min.vaario@helsinki.fi

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

**165**

559-565.

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

[10] Nadeem SM, Ahmad M, Zahir ZA, et al. The role of mycorrhizae and plant growth promoting rhizobacteria (PGPR) in improving crop productivity

under stressful environments. Biotechnol Adv 2014; 32: 429-448.

[11] Hall IR, Yun W, Amicucci A. Cultivation of edible ectomycorrhizal mushrooms. TRENDS Biotechnol 2003;

[12] Boa E. Non-wood forest products. Wild edible fungi a glob overview of their use and importance to people.

Mycorrhizal Symbiosis. Academic Press

Finlay RD, et al. Linking plants to rocks:

[15] Phillips RP, Brzostek E, Midgley MG. The mycorrhizal-associated nutrient economy: a new framework for predicting carbon–nutrient couplings in temperate forests. New Phytol 2013;

[13] Smith SE. Read. DJ 1997.

[14] Landeweert R, Hoffland E,

ectomycorrhizal fungi mobilize nutrients from minerals. Trends Ecol

[16] Brundrett MC. Mycorrhizal associations and other means of nutrition of vascular plants: understanding the global diversity of host plants by resolving conflicting information and developing reliable means of diagnosis. Plant Soil 2009;

[17] Rinaldi AC, Comandini O, Kuyper TW. Ectomycorrhizal fungal diversity: seperating the wheat from the chaff. Fungal Divers 2008; 33: 1-45.

[18] Tedersoo L, Mett M, Ishida TA, et al. Phylogenetic relationships among host plants explain differences in

Evol 2001; 16: 248-254.

21: 433-438.

2004 FAO.

London.

199: 41-51.

320: 37-77.

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

[1] Pilz D, Molina R. *Managing forest ecosystems to conserve fungus diversity and sustain wild mushroom harvests*. US Department of Agriculture, Forest Service, Pacific Northwest Research

[2] Führer E. Forest functions,

ecosystem stability and management. For Ecol Manage 2000; 132: 29-38.

[3] Mandzak JM, Moore JA. The role of nutrition in the health of inland western forests. J Sustain For 1994; 2: 191-210.

[4] O'Neill KP. *Soils as an indicator of forest health: a guide to the collection, analysis, and interpretation of soil indicator data in the Forest Inventory and Analysis program*. USDA Forest Service, North Central Research Station, 2005.

[5] Schimel JP, Bennett J. Nitrogen mineralization: challenges of a

Phytol 1988; 108: 425-431.

1995, pp. 1-11.

1997; 189: 303-319.

changing paradigm. Ecology 2004; 85:

[6] Abuarghub SM, Read DJ. The biology of mycorrhiza in the Ericaceae XI. The distribution of nitrogen in soil of a typical upland Callunetum with special reference to the 'free'amino acids. New

[7] Kelley KR, Stevenson FJ. Forms and nature of organic N in soil. In: Nitrogen Economy in Tropical Soils. Springer,

[8] Hartley J, Cairney JWG, Meharg AA. Do ectomycorrhizal fungi exhibit adaptive tolerance to potentially toxic metals in the environment? Plant Soil

[9] Bandou E, Lebailly F, Muller F, et al. The ectomycorrhizal fungus Scleroderma bermudense alleviates salt stress in seagrape (Coccoloba uvifera L.) seedlings. Mycorrhiza 2006; 16:

Station, 1996.

**References**

591-602.

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

#### **References**

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

obtain enough nutrients from the environment.

directly benefit from its associated bacteria. Sporocarps of *Cantharellus cibarius* contain large amount of bacteria, in particular fluorescent *Pseudomonas* [155]. Some species of bacteria such as *Streptomyces* spp., *Paenibacillus* spp. and *Bacillales* spp. were isolated from the mycorrhizal root tips and fruitbodies of *T. matsutake* as well [156–158]. Otherwise, the information about mycorrhizas-associated bacteria and their effect on the nutrient uptake of ECM fungi is limited. These studies, however, hint that the production of ectomycorrhizal mushrooms may require teamwork to

In conclusion, ECM fungi play an important role in the nutrient cycle of forest ecosystems, especially on mediating C and N movement. A better understanding of the nitrogen status of the habitat of ECM fungi, nutrients movements within the ecosystems, as well as the ECM fungal hyphal structures should be the first step for cultivation of ECM edible mushrooms. The methodological advances in these areas in combination with forest management may allow the successful establishment of commercial plantations and production of edible ECM mushrooms in forests.

**164**

**Author details**

Lu-Min Vaario1,2\* and Norihisa Matsushita2

PO Box 27, FI-00014 Helsinki, Finland

1-1-1 Yayoi, Bunkyo-ku, Tokyo, Japan

provided the original work is properly cited.

1 Department of Forest Sciences, University of Helsinki,

\*Address all correspondence to: lu-min.vaario@helsinki.fi

2 Graduate School of Agricultural and Life Sciences, The University of Tokyo,

© 2020 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

[1] Pilz D, Molina R. *Managing forest ecosystems to conserve fungus diversity and sustain wild mushroom harvests*. US Department of Agriculture, Forest Service, Pacific Northwest Research Station, 1996.

[2] Führer E. Forest functions, ecosystem stability and management. For Ecol Manage 2000; 132: 29-38.

[3] Mandzak JM, Moore JA. The role of nutrition in the health of inland western forests. J Sustain For 1994; 2: 191-210.

[4] O'Neill KP. *Soils as an indicator of forest health: a guide to the collection, analysis, and interpretation of soil indicator data in the Forest Inventory and Analysis program*. USDA Forest Service, North Central Research Station, 2005.

[5] Schimel JP, Bennett J. Nitrogen mineralization: challenges of a changing paradigm. Ecology 2004; 85: 591-602.

[6] Abuarghub SM, Read DJ. The biology of mycorrhiza in the Ericaceae XI. The distribution of nitrogen in soil of a typical upland Callunetum with special reference to the 'free'amino acids. New Phytol 1988; 108: 425-431.

[7] Kelley KR, Stevenson FJ. Forms and nature of organic N in soil. In: Nitrogen Economy in Tropical Soils. Springer, 1995, pp. 1-11.

[8] Hartley J, Cairney JWG, Meharg AA. Do ectomycorrhizal fungi exhibit adaptive tolerance to potentially toxic metals in the environment? Plant Soil 1997; 189: 303-319.

[9] Bandou E, Lebailly F, Muller F, et al. The ectomycorrhizal fungus Scleroderma bermudense alleviates salt stress in seagrape (Coccoloba uvifera L.) seedlings. Mycorrhiza 2006; 16: 559-565.

[10] Nadeem SM, Ahmad M, Zahir ZA, et al. The role of mycorrhizae and plant growth promoting rhizobacteria (PGPR) in improving crop productivity under stressful environments. Biotechnol Adv 2014; 32: 429-448.

[11] Hall IR, Yun W, Amicucci A. Cultivation of edible ectomycorrhizal mushrooms. TRENDS Biotechnol 2003; 21: 433-438.

[12] Boa E. Non-wood forest products. Wild edible fungi a glob overview of their use and importance to people. 2004 FAO.

[13] Smith SE. Read. DJ 1997. Mycorrhizal Symbiosis. Academic Press London.

[14] Landeweert R, Hoffland E, Finlay RD, et al. Linking plants to rocks: ectomycorrhizal fungi mobilize nutrients from minerals. Trends Ecol Evol 2001; 16: 248-254.

[15] Phillips RP, Brzostek E, Midgley MG. The mycorrhizal-associated nutrient economy: a new framework for predicting carbon–nutrient couplings in temperate forests. New Phytol 2013; 199: 41-51.

[16] Brundrett MC. Mycorrhizal associations and other means of nutrition of vascular plants: understanding the global diversity of host plants by resolving conflicting information and developing reliable means of diagnosis. Plant Soil 2009; 320: 37-77.

[17] Rinaldi AC, Comandini O, Kuyper TW. Ectomycorrhizal fungal diversity: seperating the wheat from the chaff. Fungal Divers 2008; 33: 1-45.

[18] Tedersoo L, Mett M, Ishida TA, et al. Phylogenetic relationships among host plants explain differences in

fungal species richness and community composition in ectomycorrhizal symbiosis. New Phytol 2013; 199: 822-831.

[19] Read DJ, Perez-Moreno J. Mycorrhizas and nutrient cycling in ecosystems–a journey towards relevance? New Phytol 2003; 157: 475-492.

[20] Orwin KH, Kirschbaum MUF, St John MG, et al. Organic nutrient uptake by mycorrhizal fungi enhances ecosystem carbon storage: a modelbased assessment. Ecol Lett 2011; 14: 493-502.

[21] Zak DR, Pellitier PT, Argiroff W, et al. Exploring the role of ectomycorrhizal fungi in soil carbon dynamics. New Phytol 2019; 223: 33-39.

[22] Toljander JF, Eberhardt U, Toljander YK, et al. Species composition of an ectomycorrhizal fungal community along a local nutrient gradient in a boreal forest. New Phytol 2006; 170: 873-884.

[23] Lin G, McCormack ML, Ma C, et al. Similar below-ground carbon cycling dynamics but contrasting modes of nitrogen cycling between arbuscular mycorrhizal and ectomycorrhizal forests. New Phytol 2017; 213: 1440-1451.

[24] Laliberté E, Lambers H, Burgess TI, et al. Phosphorus limitation, soil-borne pathogens and the coexistence of plant species in hyperdiverse forests and shrublands. New Phytol 2015; 206: 507-521.

[25] Egerton-Warburton LM, Johnson NC, Allen EB. Mycorrhizal community dynamics following nitrogen fertilization: a cross-site test in five grasslands. Ecol Monogr 2007; 77: 527-544.

[26] Steidinger BS, Crowther TW, Liang J, et al. Climatic controls of decomposition drive the global biogeography of forest-tree symbioses. Nature 2019; 569: 404-408.

[27] Smith ML, Bruhn JN, Anderson JB. The fungus Armillaria bulbosa is among the largest and oldest living organisms. Nature 1992; 356: 428-431.

[28] Agerer R. Exploration types of ectomycorrhizae. Mycorrhiza 2001; 11: 107-114.

[29] Unestam T, Sun Y-P. Extramatrical structures of hydrophobic and hydrophilic ectomycorrhizal fungi. Mycorrhiza 1995; 5: 301-311.

[30] Hobbie EA, Agerer R. Nitrogen isotopes in ectomycorrhizal sporocarps correspond to belowground exploration types. Plant Soil 2010; 327: 71-83.

[31] Finlay RD, Read DJ. The structure and function of the vegetative mycelium of ectomycorrhizal plants: II. The uptake and distribution of phosphorus by mycelial strands interconnecting host plants. New Phytol 1986; 103: 157-165.

[32] Burgess T, Dell B, Malajczuk N. Variation in mycorrhizal development and growth stimulation by 20 Pisolithus isolates inoculated on to Eucalyptus grandis W. Hill ex Maiden. New Phytol 1994; 127: 731-739.

[33] Burgess TI, Malajczuk N, Grove TS. The ability of 16 ectomycorrhizal fungi to increase growth and phosphorus uptake of Eucalyptus globulus Labill. and E. diversicolor F. Muell. Plant Soil 1993; 153: 155-164.

[34] Dell B, Malajczuk N, Bougher NL, et al. Development and function of Pisolithus and Scleroderma ectomycorrhizas formed in vivo with Allocasuarina, Casuarina and Eucalyptus. Mycorrhiza 1994; 5: 129-138.

**167**

209-219.

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

formed under widely differing climatic conditions. Geochim Cosmochim Acta

[45] Werdin-Pfisterer NR, Kielland K, Boone RD. Soil amino acid composition across a boreal forest successional sequence. Soil Biol Biochem 2009; 41:

[46] Vancampenhout K, Wouters K, De Vos B, et al. Differences in chemical composition of soil organic matter in natural ecosystems from different climatic regions–A pyrolysis–GC/ MS study. Soil Biol Biochem 2009; 41:

[47] Meyer FH. Distribution of ectomycorrhizae in native and manmade forests. Ectomycorrhizae their

[48] Rangel-Castro IJ, Danell E, Taylor AF. Use of different nitrogen sources by the edible ectomycorrhizal mushroom Cantharellus cibarius. Mycorrhiza 2002; 12: 131-137.

Ecol Physiol 1973; 79-105.

[49] Chalot M, Plassard C.

Metab plants 2011; 69-94.

155: 771-783.

Ectomycorrhiza and nitrogen provision to the host tree. Ecol Asp nitrogen

[50] Clemmensen KE, Sorensen PL, Michelsen A, et al. Site-dependent N uptake from N-form mixtures by arctic plants, soil microbes and ectomycorrhizal fungi. Oecologia 2008;

[51] Nygren CMR, Eberhardt U, Karlsson M, et al. Growth on nitrate and occurrence of nitrate reductaseencoding genes in a phylogenetically diverse range of ectomycorrhizal fungi.

New Phytol 2008; 180: 875-889.

[52] Turnbull MH, Goodall R,

Stewart GR. The impact of mycorrhizal colonization upon nitrogen source utilization and metabolism in seedlings of Eucalyptus grandis Hill ex Maiden

1977; 41: 1524-1526.

1210-1220.

568-579.

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

ectomycorrhizal fungi and the growth response of Pinus sylvestris L. New

[36] Islam MR, Tudryn G, Bucinell R, et al. Morphology and mechanics of fungal mycelium. Sci Rep 2017; 7: 1-12.

[37] Thomson BD, Grove TS, Malajczuk N, et al. The effectiveness of ectomycorrhizal

[38] Cairney JWG. Extramatrical mycelia of ectomycorrhizal fungi as moderators of carbon dynamics in forest soil. Soil Biol Biochem 2012; 47: 198-208.

[39] Vestgarden LS. Carbon and nitrogen turnover in the early stage of Scots pine (Pinus sylvestris L.) needle litter decomposition: effects of internal and external nitrogen. Soil Biol Biochem

[40] Wu T. Can ectomycorrhizal fungi circumvent the nitrogen mineralization for plant nutrition in temperate forest ecosystems? Soil Biol Biochem 2011; 43:

[41] Alexander IJ. The significance of ectomycorrhizas in the nitrogen cycle. In'Nitrogen as an Ecological Factor'. (Eds J. A. Lee, S. McNeill and IH Rorison.).1983; pp. 69-94.

[42] Schnitzer M, Schulten H-R. New ideas on the chemical make-up of soil humic and fulvic acids. Futur Prospect

[43] Jones DL, Kielland K. Soil amino acid turnover dominates the nitrogen flux in permafrost-dominated taiga forest soils. Soil Biol Biochem 2002; 34:

[44] Sowden FJ, Chen Y, Schnitzer M. The nitrogen distribution in soils

soil Chem 1998; 55: 153-177.

2001; 33: 465-474.

1109-1117.

fungi in increasing the growth of Eucalyptus globulus Labill. in relation to root colonization and hyphal development in soil. New Phytol 1994; 126: 517-524.

extramatrical mycelium of

Phytol 1992; 120: 127-135.

[35] Colpaert J V, Van Assche JA, Luijtens K. The growth of the

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

extramatrical mycelium of ectomycorrhizal fungi and the growth response of Pinus sylvestris L. New Phytol 1992; 120: 127-135.

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

decomposition drive the global

Nature 2019; 569: 404-408.

Nature 1992; 356: 428-431.

107-114.

biogeography of forest-tree symbioses.

[27] Smith ML, Bruhn JN, Anderson JB. The fungus Armillaria bulbosa is among the largest and oldest living organisms.

[28] Agerer R. Exploration types of ectomycorrhizae. Mycorrhiza 2001; 11:

[29] Unestam T, Sun Y-P. Extramatrical

[30] Hobbie EA, Agerer R. Nitrogen isotopes in ectomycorrhizal sporocarps correspond to belowground exploration types. Plant Soil 2010; 327: 71-83.

[31] Finlay RD, Read DJ. The structure and function of the vegetative mycelium of ectomycorrhizal plants: II. The uptake and distribution of phosphorus by mycelial strands interconnecting host plants. New Phytol 1986; 103: 157-165.

[32] Burgess T, Dell B, Malajczuk N. Variation in mycorrhizal development and growth stimulation by 20 Pisolithus isolates inoculated on to Eucalyptus grandis W. Hill ex Maiden. New Phytol

[33] Burgess TI, Malajczuk N, Grove TS. The ability of 16 ectomycorrhizal fungi to increase growth and phosphorus uptake of Eucalyptus globulus Labill. and E. diversicolor F. Muell. Plant Soil

Bougher NL, et al. Development and function of Pisolithus and Scleroderma ectomycorrhizas formed in vivo with Allocasuarina, Casuarina and Eucalyptus. Mycorrhiza 1994; 5: 129-138.

[35] Colpaert J V, Van Assche JA, Luijtens K. The growth of the

1994; 127: 731-739.

1993; 153: 155-164.

[34] Dell B, Malajczuk N,

structures of hydrophobic and hydrophilic ectomycorrhizal fungi. Mycorrhiza 1995; 5: 301-311.

fungal species richness and community composition in ectomycorrhizal symbiosis. New Phytol 2013; 199:

[19] Read DJ, Perez-Moreno J. Mycorrhizas and nutrient cycling in ecosystems–a journey towards relevance? New Phytol 2003; 157:

[20] Orwin KH, Kirschbaum MUF, St John MG, et al. Organic nutrient uptake by mycorrhizal fungi enhances ecosystem carbon storage: a modelbased assessment. Ecol Lett 2011; 14:

[21] Zak DR, Pellitier PT, Argiroff W, et al. Exploring the role of ectomycorrhizal fungi in soil carbon dynamics. New

Toljander YK, et al. Species composition

[23] Lin G, McCormack ML, Ma C, et al. Similar below-ground carbon cycling dynamics but contrasting modes of nitrogen cycling between arbuscular mycorrhizal and ectomycorrhizal forests.

[24] Laliberté E, Lambers H, Burgess TI, et al. Phosphorus limitation, soil-borne pathogens and the coexistence of plant species in hyperdiverse forests and shrublands. New Phytol 2015; 206:

nitrogen fertilization: a cross-site test in five grasslands. Ecol Monogr 2007; 77:

[26] Steidinger BS, Crowther TW, Liang J, et al. Climatic controls of

New Phytol 2017; 213: 1440-1451.

[25] Egerton-Warburton LM, Johnson NC, Allen EB. Mycorrhizal community dynamics following

Phytol 2019; 223: 33-39.

2006; 170: 873-884.

[22] Toljander JF, Eberhardt U,

of an ectomycorrhizal fungal community along a local nutrient gradient in a boreal forest. New Phytol

822-831.

475-492.

493-502.

**166**

507-521.

527-544.

[36] Islam MR, Tudryn G, Bucinell R, et al. Morphology and mechanics of fungal mycelium. Sci Rep 2017; 7: 1-12.

[37] Thomson BD, Grove TS, Malajczuk N, et al. The effectiveness of ectomycorrhizal fungi in increasing the growth of Eucalyptus globulus Labill. in relation to root colonization and hyphal development in soil. New Phytol 1994; 126: 517-524.

[38] Cairney JWG. Extramatrical mycelia of ectomycorrhizal fungi as moderators of carbon dynamics in forest soil. Soil Biol Biochem 2012; 47: 198-208.

[39] Vestgarden LS. Carbon and nitrogen turnover in the early stage of Scots pine (Pinus sylvestris L.) needle litter decomposition: effects of internal and external nitrogen. Soil Biol Biochem 2001; 33: 465-474.

[40] Wu T. Can ectomycorrhizal fungi circumvent the nitrogen mineralization for plant nutrition in temperate forest ecosystems? Soil Biol Biochem 2011; 43: 1109-1117.

[41] Alexander IJ. The significance of ectomycorrhizas in the nitrogen cycle. In'Nitrogen as an Ecological Factor'. (Eds J. A. Lee, S. McNeill and IH Rorison.).1983; pp. 69-94.

[42] Schnitzer M, Schulten H-R. New ideas on the chemical make-up of soil humic and fulvic acids. Futur Prospect soil Chem 1998; 55: 153-177.

[43] Jones DL, Kielland K. Soil amino acid turnover dominates the nitrogen flux in permafrost-dominated taiga forest soils. Soil Biol Biochem 2002; 34: 209-219.

[44] Sowden FJ, Chen Y, Schnitzer M. The nitrogen distribution in soils

formed under widely differing climatic conditions. Geochim Cosmochim Acta 1977; 41: 1524-1526.

[45] Werdin-Pfisterer NR, Kielland K, Boone RD. Soil amino acid composition across a boreal forest successional sequence. Soil Biol Biochem 2009; 41: 1210-1220.

[46] Vancampenhout K, Wouters K, De Vos B, et al. Differences in chemical composition of soil organic matter in natural ecosystems from different climatic regions–A pyrolysis–GC/ MS study. Soil Biol Biochem 2009; 41: 568-579.

[47] Meyer FH. Distribution of ectomycorrhizae in native and manmade forests. Ectomycorrhizae their Ecol Physiol 1973; 79-105.

[48] Rangel-Castro IJ, Danell E, Taylor AF. Use of different nitrogen sources by the edible ectomycorrhizal mushroom Cantharellus cibarius. Mycorrhiza 2002; 12: 131-137.

[49] Chalot M, Plassard C. Ectomycorrhiza and nitrogen provision to the host tree. Ecol Asp nitrogen Metab plants 2011; 69-94.

[50] Clemmensen KE, Sorensen PL, Michelsen A, et al. Site-dependent N uptake from N-form mixtures by arctic plants, soil microbes and ectomycorrhizal fungi. Oecologia 2008; 155: 771-783.

[51] Nygren CMR, Eberhardt U, Karlsson M, et al. Growth on nitrate and occurrence of nitrate reductaseencoding genes in a phylogenetically diverse range of ectomycorrhizal fungi. New Phytol 2008; 180: 875-889.

[52] Turnbull MH, Goodall R, Stewart GR. The impact of mycorrhizal colonization upon nitrogen source utilization and metabolism in seedlings of Eucalyptus grandis Hill ex Maiden

and Eucalyptus maculata Hook. Plant Cell Environ 1995; 18: 1386-1394.

[53] Hobbie EA, Colpaert J V, White MW, et al. Nitrogen form, availability, and mycorrhizal colonization affect biomass and nitrogen isotope patterns in Pinus sylvestris. Plant Soil 2008; 310: 121.

[54] Read DJ, Leake JR, Perez-Moreno J. Mycorrhizal fungi as drivers of ecosystem processes in heathland and boreal forest biomes. Can J Bot 2004; 82: 1243-1263.

[55] Melin E, Nilsson H. Transfer of labelled nitrogen from glutamic acid to pine seedlings through the mycelium of Boletus variegatus (Sw.) Fr. Nature 1953; 171: 134.

[56] Chalot M, Brun A. Physiology of organic nitrogen acquisition by ectomycorrhizal fungi and ectomycorrhizas. FEMS Microbiol Rev 1998; 22: 21-44.

[57] Plassard C, Bonafos B, Touraine B. Differential effects of mineral and organic N sources, and of ectomycorrhizal infection by Hebeloma cylindrosporum, on growth and N utilization in Pinus pinaster. Plant Cell Environ 2000; 23: 1195-1205.

[58] Taylor AFS, Fransson PM, Högberg P, et al. Species level patterns in 13C and 15N abundance of ectomycorrhizal and saprotrophic fungal sporocarps. New Phytol 2003; 159: 757-774.

[59] Guidot A, Verner M-C, Debaud J-C, et al. Intraspecific variation in use of different organic nitrogen sources by the ectomycorrhizal fungus Hebeloma cylindrosporum. Mycorrhiza 2005; 15: 167-177.

[60] Abuzinadah RA, Read DJ. The role of proteins in the nitrogen nutrition of ectomycorrhizal plants: I. Utilization of peptides and proteins by ectomycorrhizal fungi. New Phytol 1986; 103: 481-493.

[61] Courty P, Pritsch K, Schloter M, et al. Activity profiling of ectomycorrhiza communities in two forest soils using multiple enzymatic tests. New Phytol 2005; 167: 309-319.

[62] Lilleskov EA, Hobbie EA, Horton TR. Conservation of ectomycorrhizal fungi: exploring the linkages between functional and taxonomic responses to anthropogenic N deposition. fungal Ecol 2011; 4: 174-183.

[63] Peay KG, Kennedy PG, Bruns TD. Rethinking ectomycorrhizal succession: are root density and hyphal exploration types drivers of spatial and temporal zonation? fungal Ecol 2011; 4: 233-240.

[64] Dawson TE, Mambelli S, Plamboeck AH, et al. Stable isotopes in plant ecology. Annu Rev Ecol Syst 2002; 33: 507-559.

[65] Hobbie EA, Hobbie JE. Natural abundance of 15 N in nitrogen-limited forests and tundra can estimate nitrogen cycling through mycorrhizal fungi: a review. Ecosystems 2008; 11: 815-830.

[66] Trudell SA, Rygiewicz PT, Edmonds RL. Patterns of nitrogen and carbon stable isotope ratios in macrofungi, plants and soils in two old-growth conifer forests. New Phytol 2004; 164: 317-335.

[67] Lilleskov EA, Hobbie EA, Fahey TJ. Ectomycorrhizal fungal taxa differing in response to nitrogen deposition also differ in pure culture organic nitrogen use and natural abundance of nitrogen isotopes. New Phytol 2002; 154: 219-231.

[68] Hobbie EA, Sánchez FS, Rygiewicz PT. Controls of isotopic patterns in saprotrophic and ectomycorrhizal fungi. Soil Biol Biochem 2012; 48: 60-68.

**169**

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

2010-2022.

1047-1058.

ectomycorrhizal fungus. ISME J 2013; 7:

[77] Casieri L, Lahmidi NA, Doidy J, et al. Biotrophic transportome in mutualistic plant–fungal interactions.

Mycorrhiza 2013; 23: 597-625.

[79] Becquer A, Guerrero-

[78] Nehls U, Plassard C. Nitrogen and phosphate metabolism in

Galán C, Eibensteiner JL, et al. The ectomycorrhizal contribution to tree nutrition. In: Advances in Botanical Research. Elsevier, 2019, pp. 77-126.

[80] Javelle A, Rodrı́guez-Pastrana B-R, Jacob C, et al. Molecular characterization of two ammonium transporters from the ectomycorrhizal fungus Hebeloma cylindrosporum. FEBS Lett 2001; 505: 393-398.

[81] Montanini B, Moretto N, Soragni E, et al. A high-affinity ammonium transporter from the mycorrhizal ascomycete Tuber borchii. Fungal Genet Biol 2002; 36: 22-34.

[82] Willmann A, Weiß M, Nehls U. Ectomycorrhiza-mediated repression of the high-affinity ammonium importer gene AmAMT2 in Amanita muscaria.

[83] Kemppainen MJ, Pardo AG. pHg/ pSILBAγ vector system for efficient gene silencing in homobasidiomycetes: optimization of ihpRNA–triggering in the mycorrhizal fungus Laccaria bicolor. Microb Biotechnol 2010; 3: 178-200.

[84] Jargeat P, Rekangalt D, Verner M-C, et al. Characterisation and expression analysis of a nitrate transporter and nitrite reductase genes, two members of a gene cluster for nitrate assimilation from the symbiotic basidiomycete Hebeloma cylindrosporum. Curr Genet

Curr Genet 2007; 51: 71.

2003; 43: 199-205.

ectomycorrhizas. New Phytol 2018; 220:

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

[69] Kranabetter JM. Ectomycorrhizal fungi and the nitrogen economy of conifers—implications for genecology and climate change mitigation. Botany

[70] Pardo LH, Fenn ME, Goodale CL, et al. Effects of nitrogen deposition and empirical nitrogen critical loads for ecoregions of the United States. Ecol

2014; 92: 417-423.

Appl 2011; 21: 3049-3082.

[71] Morrison EW, Frey SD,

et al. Nitrogen addition alters

[73] Zhu H, Dancik BP,

[75] Javelle A, Morel M,

14-23.

227-234.

343-349.

47: 411-430.

Sadowsky JJ, et al. Chronic nitrogen additions fundamentally restructure the soil fungal community in a temperate forest. fungal Ecol 2016; 23: 48-57.

[72] Corrales A, Turner BL, Tedersoo L,

ectomycorrhizal fungal communities and soil enzyme activities in a tropical montane forest. fungal Ecol 2017; 27:

Higginbotham KO. Regulation of extracellular proteinase production in an ectomycorrhizal fungus Hebeloma crustuliniforme. Mycologia 1994; 86:

[74] Nehls UWE, Kleber R, Wiese J, et al. Isolation and characterization of a general amino acid permease from the ectomycorrhizal fungus Amanita muscaria. New Phytol 1999; 144:

Rodríguez-Pastrana B, et al. Molecular

characterization, function and regulation of ammonium transporters (Amt) and ammonium-metabolizing enzymes (GS, NADP-GDH) in the ectomycorrhizal fungus Hebeloma cylindrosporum. Mol Microbiol 2003;

[76] Rineau F, Shah F, Smits MM, et al. Carbon availability triggers the decomposition of plant litter and assimilation of nitrogen by an *Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

[69] Kranabetter JM. Ectomycorrhizal fungi and the nitrogen economy of conifers—implications for genecology and climate change mitigation. Botany 2014; 92: 417-423.

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

by ectomycorrhizal fungi. New Phytol

[61] Courty P, Pritsch K, Schloter M,

ectomycorrhiza communities in two forest soils using multiple enzymatic tests. New Phytol 2005; 167: 309-319.

[63] Peay KG, Kennedy PG, Bruns TD. Rethinking ectomycorrhizal succession: are root density and hyphal exploration types drivers of spatial and temporal zonation? fungal Ecol 2011; 4: 233-240.

Plamboeck AH, et al. Stable isotopes in plant ecology. Annu Rev Ecol Syst 2002;

[65] Hobbie EA, Hobbie JE. Natural abundance of 15 N in nitrogen-limited forests and tundra can estimate nitrogen cycling through mycorrhizal fungi: a review. Ecosystems 2008; 11: 815-830.

[66] Trudell SA, Rygiewicz PT, Edmonds RL. Patterns of nitrogen and carbon stable isotope ratios in macrofungi, plants and soils in two old-growth conifer forests. New Phytol

[68] Hobbie EA, Sánchez FS, Rygiewicz PT. Controls of isotopic patterns in saprotrophic and ectomycorrhizal fungi. Soil Biol Biochem 2012; 48: 60-68.

[67] Lilleskov EA, Hobbie EA, Fahey TJ. Ectomycorrhizal fungal taxa differing in response to nitrogen deposition also differ in pure culture organic nitrogen use and natural abundance of nitrogen isotopes. New Phytol 2002; 154: 219-231.

2004; 164: 317-335.

[64] Dawson TE, Mambelli S,

1986; 103: 481-493.

174-183.

33: 507-559.

et al. Activity profiling of

[62] Lilleskov EA, Hobbie EA, Horton TR. Conservation of ectomycorrhizal fungi: exploring the linkages between functional and taxonomic responses to anthropogenic N deposition. fungal Ecol 2011; 4:

and Eucalyptus maculata Hook. Plant Cell Environ 1995; 18: 1386-1394.

[53] Hobbie EA, Colpaert J V, White MW, et al. Nitrogen form, availability, and mycorrhizal colonization affect biomass and nitrogen isotope patterns in Pinus sylvestris. Plant Soil 2008; 310: 121.

Moreno J. Mycorrhizal fungi as drivers of ecosystem processes in heathland and boreal forest biomes. Can J Bot 2004; 82:

[55] Melin E, Nilsson H. Transfer of labelled nitrogen from glutamic acid to pine seedlings through the mycelium of Boletus variegatus (Sw.) Fr. Nature

[56] Chalot M, Brun A. Physiology of organic nitrogen acquisition by ectomycorrhizal fungi and

ectomycorrhizas. FEMS Microbiol Rev

[57] Plassard C, Bonafos B, Touraine B. Differential effects of mineral and

ectomycorrhizal infection by Hebeloma cylindrosporum, on growth and N utilization in Pinus pinaster. Plant Cell

Högberg P, et al. Species level patterns

[59] Guidot A, Verner M-C, Debaud J-C, et al. Intraspecific variation in use of different organic nitrogen sources by the ectomycorrhizal fungus Hebeloma cylindrosporum. Mycorrhiza 2005; 15:

[60] Abuzinadah RA, Read DJ. The role of proteins in the nitrogen nutrition of ectomycorrhizal plants: I. Utilization of peptides and proteins

[54] Read DJ, Leake JR, Perez-

1243-1263.

1953; 171: 134.

1998; 22: 21-44.

159: 757-774.

167-177.

organic N sources, and of

Environ 2000; 23: 1195-1205.

[58] Taylor AFS, Fransson PM,

in 13C and 15N abundance of ectomycorrhizal and saprotrophic fungal sporocarps. New Phytol 2003;

**168**

[70] Pardo LH, Fenn ME, Goodale CL, et al. Effects of nitrogen deposition and empirical nitrogen critical loads for ecoregions of the United States. Ecol Appl 2011; 21: 3049-3082.

[71] Morrison EW, Frey SD, Sadowsky JJ, et al. Chronic nitrogen additions fundamentally restructure the soil fungal community in a temperate forest. fungal Ecol 2016; 23: 48-57.

[72] Corrales A, Turner BL, Tedersoo L, et al. Nitrogen addition alters ectomycorrhizal fungal communities and soil enzyme activities in a tropical montane forest. fungal Ecol 2017; 27: 14-23.

[73] Zhu H, Dancik BP, Higginbotham KO. Regulation of extracellular proteinase production in an ectomycorrhizal fungus Hebeloma crustuliniforme. Mycologia 1994; 86: 227-234.

[74] Nehls UWE, Kleber R, Wiese J, et al. Isolation and characterization of a general amino acid permease from the ectomycorrhizal fungus Amanita muscaria. New Phytol 1999; 144: 343-349.

[75] Javelle A, Morel M, Rodríguez-Pastrana B, et al. Molecular characterization, function and regulation of ammonium transporters (Amt) and ammonium-metabolizing enzymes (GS, NADP-GDH) in the ectomycorrhizal fungus Hebeloma cylindrosporum. Mol Microbiol 2003; 47: 411-430.

[76] Rineau F, Shah F, Smits MM, et al. Carbon availability triggers the decomposition of plant litter and assimilation of nitrogen by an ectomycorrhizal fungus. ISME J 2013; 7: 2010-2022.

[77] Casieri L, Lahmidi NA, Doidy J, et al. Biotrophic transportome in mutualistic plant–fungal interactions. Mycorrhiza 2013; 23: 597-625.

[78] Nehls U, Plassard C. Nitrogen and phosphate metabolism in ectomycorrhizas. New Phytol 2018; 220: 1047-1058.

[79] Becquer A, Guerrero-Galán C, Eibensteiner JL, et al. The ectomycorrhizal contribution to tree nutrition. In: Advances in Botanical Research. Elsevier, 2019, pp. 77-126.

[80] Javelle A, Rodrı́guez-Pastrana B-R, Jacob C, et al. Molecular characterization of two ammonium transporters from the ectomycorrhizal fungus Hebeloma cylindrosporum. FEBS Lett 2001; 505: 393-398.

[81] Montanini B, Moretto N, Soragni E, et al. A high-affinity ammonium transporter from the mycorrhizal ascomycete Tuber borchii. Fungal Genet Biol 2002; 36: 22-34.

[82] Willmann A, Weiß M, Nehls U. Ectomycorrhiza-mediated repression of the high-affinity ammonium importer gene AmAMT2 in Amanita muscaria. Curr Genet 2007; 51: 71.

[83] Kemppainen MJ, Pardo AG. pHg/ pSILBAγ vector system for efficient gene silencing in homobasidiomycetes: optimization of ihpRNA–triggering in the mycorrhizal fungus Laccaria bicolor. Microb Biotechnol 2010; 3: 178-200.

[84] Jargeat P, Rekangalt D, Verner M-C, et al. Characterisation and expression analysis of a nitrate transporter and nitrite reductase genes, two members of a gene cluster for nitrate assimilation from the symbiotic basidiomycete Hebeloma cylindrosporum. Curr Genet 2003; 43: 199-205.

[85] Kohler A, Kuo A, Nagy LG, et al. Convergent losses of decay mechanisms and rapid turnover of symbiosis genes in mycorrhizal mutualists. Nat Genet 2015; 47: 410-415.

[86] Shah F, Nicolás C, Bentzer J, et al. Ectomycorrhizal fungi decompose soil organic matter using oxidative mechanisms adapted from saprotrophic ancestors. New Phytol 2016; 209: 1705-1719.

[87] Lindahl BD, Taylor AFS. Occurrence of N-acetylhexosaminidase-encoding genes in ectomycorrhizal basidiomycetes. New Phytol 2004; 164: 193-199.

[88] Miyauchi S, Kiss E, Kuo A, et al. Large-scale genome sequencing of mycorrhizal fungi provides insights into the early evolution of symbiotic traits. Nat Commun 2020; 11: 1-17.

[89] Talbot JM, Treseder KK. Controls over mycorrhizal uptake of organic nitrogen. Pedobiologia (Jena) 2010; 53: 169-179.

[90] Näsholm T, Högberg P, Franklin O, et al. Are ectomycorrhizal fungi alleviating or aggravating nitrogen limitation of tree growth in boreal forests? New Phytol 2013; 198: 214-221.

[91] Nicolás C, Martin-Bertelsen T, Floudas D, et al. The soil organic matter decomposition mechanisms in ectomycorrhizal fungi are tuned for liberating soil organic nitrogen. ISME J 2019; 13: 977-988.

[92] Lindahl BD, Ihrmark K, Boberg J, et al. Spatial separation of litter decomposition and mycorrhizal nitrogen uptake in a boreal forest. New Phytol 2007; 173: 611-620.

[93] Talbot JM, Allison SD, Treseder KK. Decomposers in disguise: mycorrhizal fungi as regulators of soil C dynamics in ecosystems under global change. Funct Ecol 2008; 22: 955-963.

[94] Sinsabaugh RL. Phenol oxidase, peroxidase and organic matter dynamics of soil. Soil Biol Biochem 2010; 42: 391-404.

[95] Hall IR, Zambonelli A. Laying the foundations. In: Edible ectomycorrhizal mushrooms. Springer, 2012, pp. 3-16.

[96] Poitou N, Mamoun M, Ducamp M, et al. After Boletus granulatus, Lactarius deliciosus fructification is obtained in the field from inoculated plants. PHM Rev Hortic 1984; 244: 65-68.

[97] Guerin-Laguette A, Cummings N, Butler RC, et al. Lactarius deliciosus and Pinus radiata in New Zealand: towards the development of innovative gourmet mushroom orchards. Mycorrhiza 2014; 24: 511-523.

[98] Wang R, Guerin-Laguette A, Yu F-Q. Optimum media for hyphal growth and mycorrhizal synthesis of two Lactarius species. Mycosystema 2020; 39: 1346-1355.

[99] Tan ZM, Danell E, Shen AR, et al. Successful cultivation of Lactarius hatsutake—an evaluation with molecular methods. Acta Edulis Fungi 2008; 15: 85-88.

[100] Visnovsky SB, Guerin-Laguette A, Wang Y, et al. Traceability of marketable Japanese shoro in New Zealand: using multiplex PCR to exploit phylogeographic variation among taxa in the Rhizopogon subgenus Roseoli. Appl Environ Microbiol 2010; 76: 294-302.

[101] Yamanaka K. Commercial cultivation of Lyophyllum shimeji. GAMU GmbH, Institut für Pilzforschung, 2008; pp. 197-202.

[102] Hobbie EA, Högberg P. Nitrogen isotopes link mycorrhizal fungi and plants to nitrogen dynamics. New Phytol 2012; 196: 367-382.

**171**

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

[112] Itoo ZA, Reshi ZA. Effect of different nitrogen and carbon sources and concentrations on the mycelial growth of ectomycorrhizal fungi under in-vitro conditions. Scand J For Res

[113] Finlay RD, Frostegård Å, Sonnerfeldt A. Utilization of organic and inorganic nitrogen sources by ectomycorrhizal fungi in pure culture and in symbiosis with Pinus contorta Dougl. ex Loud. New Phytol 1992; 120:

[114] Yamanaka T. Utilization of inorganic and organic nitrogen in pure cultures by saprotrophic and ectomycorrhizal fungi producing sporophores on urea-treated forest floor.

Mycol Res 1999; 103: 811-816.

[116] Sun Q, Li J, Finlay RD, et al. Oxalotrophic bacterial assemblages in the ectomycorrhizosphere of forest trees and their effects on oxalate degradation

and carbon fixation potential

Oxalotrophic bacterial assemblages in the ectomycorrhizosphere of forest trees and their effects on oxala. Chem Geol

[117] Lazarević J, Stojičić D, Keča N. Effects of temperature, pH and carbon and nitrogen sources on growth of in vitro cultures of ectomycorrhizal isolates from Pinus heldreichii forest.

[118] Terashima Y. Carbon and nitrogen

[119] Hamada M. Physiology and ecology of Armillaria matsutake. Bot Mag 1950;

utilization and acid production by mycelia of the ectomycorrhizal fungusTricholoma bakamatsutake in vitro. Mycoscience 1999; 40: 51.

1994; 35: 147-151.

2019; 514: 54-64.

For Syst 2016; 25: 3.

63: 40-41.

[115] Ohta A. Production of fruit-bodies of a mycorrhizal fungus, Lyophyllum shimeji, in pure culture. Mycoscience

2014; 29: 619-628.

105-115.

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

[103] Daza A, Manjón JL, Camacho M, et al. Effect of carbon and nitrogen sources, pH and temperature on in vitro culture of several isolates of Amanita caesarea (Scop.: Fr.) Pers. Mycorrhiza

[104] Tedersoo L, Naadel T, Bahram M, et al. Enzymatic activities and stable isotope patterns of ectomycorrhizal fungi in relation to phylogeny and exploration types in an afrotropical rain forest. New Phytol 2012; 195: 832-843.

Fukuda M, et al. In vitro mycorrhization

Tajiri A. Fruit body formation ofBoletus reticulatus in pure culture. Mycoscience

[107] Ohta A, Fujiwara N. Fruit-body production of an ectomycorrhizal fungus in genus Boletus in pure culture.

[108] Ohta A. Fruit-body production of two ectomycorrhizal fungi in the genus Hebeloma in pure culture. Mycoscience

Mycoscience 2003; 44: 295-300.

and acclimatization of Amanita caesareoides and its relatives on Pinus densiflora. Mycorrhiza 2013; 23:

[106] Yamanaka K, Namba K,

2006; 16: 133-136.

[105] Endo N, Gisusi S,

303-315.

2000; 41: 189.

1998; 39: 15-19.

431-434.

[109] Peksen A, Kibar B,

Yakupoglu G. Favourable culture conditions for mycelial growth of Hydnum repandum, a medicinal mushroom. African J Tradit Complement Altern Med 2013; 10:

[110] Davis EE, Jong SC. Basidiocarp formation by Laccaria laccata in agar culture. Mycologia 1976; 68: 211-214.

[111] Taylor AFS, Högbom L, Högberg M, et al. Natural 15N abundance in fruit bodies of ectomycorrhizal fungi from boreal forests. New Phytol 1997; 136: 713-720. *Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

[103] Daza A, Manjón JL, Camacho M, et al. Effect of carbon and nitrogen sources, pH and temperature on in vitro culture of several isolates of Amanita caesarea (Scop.: Fr.) Pers. Mycorrhiza 2006; 16: 133-136.

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

[94] Sinsabaugh RL. Phenol oxidase, peroxidase and organic matter dynamics of soil. Soil Biol Biochem 2010; 42:

[95] Hall IR, Zambonelli A. Laying the foundations. In: Edible ectomycorrhizal mushrooms. Springer, 2012, pp. 3-16.

[97] Guerin-Laguette A, Cummings N, Butler RC, et al. Lactarius deliciosus and Pinus radiata in New Zealand: towards the development of innovative gourmet mushroom orchards. Mycorrhiza 2014;

[98] Wang R, Guerin-Laguette A, Yu F-Q. Optimum media for hyphal growth and mycorrhizal synthesis of two Lactarius species. Mycosystema 2020;

[99] Tan ZM, Danell E, Shen AR, et al. Successful cultivation of Lactarius hatsutake—an evaluation with

molecular methods. Acta Edulis Fungi

[100] Visnovsky SB, Guerin-Laguette A, Wang Y, et al. Traceability of marketable

phylogeographic variation among taxa in the Rhizopogon subgenus Roseoli. Appl Environ Microbiol 2010; 76:

Japanese shoro in New Zealand: using multiplex PCR to exploit

[101] Yamanaka K. Commercial cultivation of Lyophyllum shimeji.

[102] Hobbie EA, Högberg P. Nitrogen isotopes link mycorrhizal fungi and plants to nitrogen dynamics. New

GAMU GmbH, Institut für Pilzforschung, 2008; pp. 197-202.

Phytol 2012; 196: 367-382.

[96] Poitou N, Mamoun M, Ducamp M, et al. After Boletus granulatus, Lactarius deliciosus fructification is obtained in the field from inoculated plants. PHM Rev Hortic

1984; 244: 65-68.

24: 511-523.

39: 1346-1355.

2008; 15: 85-88.

294-302.

391-404.

[85] Kohler A, Kuo A, Nagy LG, et al. Convergent losses of decay mechanisms and rapid turnover of symbiosis genes in mycorrhizal mutualists. Nat Genet

[86] Shah F, Nicolás C, Bentzer J, et al. Ectomycorrhizal fungi decompose soil organic matter using oxidative mechanisms adapted from saprotrophic ancestors. New Phytol 2016; 209:

[87] Lindahl BD, Taylor AFS. Occurrence of N-acetylhexosaminidase-encoding genes in ectomycorrhizal basidiomycetes.

New Phytol 2004; 164: 193-199.

Nat Commun 2020; 11: 1-17.

[90] Näsholm T, Högberg P,

2019; 13: 977-988.

[92] Lindahl BD, Ihrmark K,

Phytol 2007; 173: 611-620.

Ecol 2008; 22: 955-963.

Boberg J, et al. Spatial separation of litter decomposition and mycorrhizal nitrogen uptake in a boreal forest. New

[93] Talbot JM, Allison SD, Treseder KK. Decomposers in disguise: mycorrhizal fungi as regulators of soil C dynamics in ecosystems under global change. Funct

[88] Miyauchi S, Kiss E, Kuo A, et al. Large-scale genome sequencing of mycorrhizal fungi provides insights into the early evolution of symbiotic traits.

[89] Talbot JM, Treseder KK. Controls over mycorrhizal uptake of organic nitrogen. Pedobiologia (Jena) 2010; 53:

Franklin O, et al. Are ectomycorrhizal fungi alleviating or aggravating nitrogen limitation of tree growth in boreal forests? New Phytol 2013; 198: 214-221.

[91] Nicolás C, Martin-Bertelsen T, Floudas D, et al. The soil organic matter decomposition mechanisms in ectomycorrhizal fungi are tuned for liberating soil organic nitrogen. ISME J

2015; 47: 410-415.

1705-1719.

169-179.

**170**

[104] Tedersoo L, Naadel T, Bahram M, et al. Enzymatic activities and stable isotope patterns of ectomycorrhizal fungi in relation to phylogeny and exploration types in an afrotropical rain forest. New Phytol 2012; 195: 832-843.

[105] Endo N, Gisusi S, Fukuda M, et al. In vitro mycorrhization and acclimatization of Amanita caesareoides and its relatives on Pinus densiflora. Mycorrhiza 2013; 23: 303-315.

[106] Yamanaka K, Namba K, Tajiri A. Fruit body formation ofBoletus reticulatus in pure culture. Mycoscience 2000; 41: 189.

[107] Ohta A, Fujiwara N. Fruit-body production of an ectomycorrhizal fungus in genus Boletus in pure culture. Mycoscience 2003; 44: 295-300.

[108] Ohta A. Fruit-body production of two ectomycorrhizal fungi in the genus Hebeloma in pure culture. Mycoscience 1998; 39: 15-19.

[109] Peksen A, Kibar B, Yakupoglu G. Favourable culture conditions for mycelial growth of Hydnum repandum, a medicinal mushroom. African J Tradit Complement Altern Med 2013; 10: 431-434.

[110] Davis EE, Jong SC. Basidiocarp formation by Laccaria laccata in agar culture. Mycologia 1976; 68: 211-214.

[111] Taylor AFS, Högbom L, Högberg M, et al. Natural 15N abundance in fruit bodies of ectomycorrhizal fungi from boreal forests. New Phytol 1997; 136: 713-720. [112] Itoo ZA, Reshi ZA. Effect of different nitrogen and carbon sources and concentrations on the mycelial growth of ectomycorrhizal fungi under in-vitro conditions. Scand J For Res 2014; 29: 619-628.

[113] Finlay RD, Frostegård Å, Sonnerfeldt A. Utilization of organic and inorganic nitrogen sources by ectomycorrhizal fungi in pure culture and in symbiosis with Pinus contorta Dougl. ex Loud. New Phytol 1992; 120: 105-115.

[114] Yamanaka T. Utilization of inorganic and organic nitrogen in pure cultures by saprotrophic and ectomycorrhizal fungi producing sporophores on urea-treated forest floor. Mycol Res 1999; 103: 811-816.

[115] Ohta A. Production of fruit-bodies of a mycorrhizal fungus, Lyophyllum shimeji, in pure culture. Mycoscience 1994; 35: 147-151.

[116] Sun Q, Li J, Finlay RD, et al. Oxalotrophic bacterial assemblages in the ectomycorrhizosphere of forest trees and their effects on oxalate degradation and carbon fixation potential Oxalotrophic bacterial assemblages in the ectomycorrhizosphere of forest trees and their effects on oxala. Chem Geol 2019; 514: 54-64.

[117] Lazarević J, Stojičić D, Keča N. Effects of temperature, pH and carbon and nitrogen sources on growth of in vitro cultures of ectomycorrhizal isolates from Pinus heldreichii forest. For Syst 2016; 25: 3.

[118] Terashima Y. Carbon and nitrogen utilization and acid production by mycelia of the ectomycorrhizal fungusTricholoma bakamatsutake in vitro. Mycoscience 1999; 40: 51.

[119] Hamada M. Physiology and ecology of Armillaria matsutake. Bot Mag 1950; 63: 40-41.

[120] Vaario LM, Sah SP, Norisada M, et al. Tricholoma matsutake may take more nitrogen in the organic form than other ectomycorrhizal fungi for its sporocarp development: the isotopic evidence. Mycorrhiza 2019; 29: 51-59.

[121] Kawai M, Abe S. Studies on the artificial reproduction of Tricholoma matsutake (S. ito et Imai) Sing. i. effects of carbon and nitrogen sources in media on the vegetative growth of Tricholoma matsutake. Transactions. 1976.

[122] Yamada A, Maeda K, Kobayashi H, et al. Ectomycorrhizal symbiosis in vitro between Tricholoma matsutake and Pinus densiflora seedlings that resembles naturally occurring 'shiro'. Mycorrhiza 2006; 16: 111-116.

[123] Kibar B, Peksen A. Nutritional and environmental requirements for vegetative growth of edible ectomycorrhizal mushroom Tricholoma terreum. Agriculture 2011; 98: 409-414.

[124] Liu R-S, Li D-S, Li H-M, et al. Response surface modeling the significance of nitrogen source on the cell growth and Tuber polysaccharides production by submerged cultivation of Chinese truffle Tuber sinense. Process Biochem 2008; 43: 868-876.

[125] Bonet JA, Oliach D, Fischer C, et al. Cultivation methods of the black truffle, the most profitable mediterranean non-wood forest product; a state of the art review. In EFI proceedings 2009; 57: 57-71.

[126] Guerin-Laguette A, Plassard C, Mousain D. Effects of experimental conditions on mycorrhizal relationships between Pinus sylvestris and Lactarius deliciosus and unprecedented fruitbody formation of the Saffron milk cap under controlled soilless conditions. Can J Microbiol 2000; 46: 790-799.

[127] Parladé J, Pera J, Luque J. Evaluation of mycelial inocula of edible Lactarius species for the production

of Pinus pinaster and P. sylvestris mycorrhizal seedlings under greenhouse conditions. Mycorrhiza 2004; 14: 171-175.

[128] Águeda B, Parladé J, Fernández-Toirán LM, et al. Mycorrhizal synthesis between Boletus edulis species complex and rockroses (Cistus sp.). Mycorrhiza 2008; 18: 443-449.

[129] Oria-de-Rueda JA, Hernández-Rodríguez M, Martín-Pinto P, et al. Could artificial reforestations provide as much production and diversity of fungal species as natural forest stands in marginal Mediterranean areas? For Ecol Manage 2010; 260: 171-180.

[130] Yamanaka T, Yamada A, Furukawa H. Advances in the cultivation of the highly-prized ectomycorrhizal mushroom Tricholoma matsutake. Mycoscience 2020; 61: 49-57.

[131] Ogawa M. Microbial ecology of mycorrhizal fungus, Tricholoma matsutake Sing. in pine forest 1. Fungal colony (' Shiro') of T. matsutake. Bull Prod Res Inst 1975; 272: 79-121.

[132] Narimatsu M, Koiwa T, Sakamoto Y, et al. Estimation of novel colony establishment and persistence of the ectomycorrhizal basidiomycete Tricholoma matsutake in a Pinus densiflora forest. fungal Ecol 2016; 24: 35-43.

[133] Hortal S, Plett KL, Plett JM, et al. Role of plant–fungal nutrient trading and host control in determining the competitive success of ectomycorrhizal fungi. ISME J 2017; 11: 2666-2676.

[134] Terashita T, Kono M. Carboxyl proteinases from Tricholoma matsutake and its related species. Mem Fac Agric Kinki Univ 1989; 22:5-12.

**173**

New York. 1983.

*Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi…*

187-199.

61-73.

403-407.

1063-1073.

Ecol 2003; 45: 49-57.

1994; 128: 197-210.

supply of nitrate. Plant Soil 1965; 22:

[145] Takeshi I, Ogawa M. Cultivating method of the mycorrhizal fungus, Tricholoma matsutake (Ito et Imai) Sing. (II) increasing number of Shiro (fungal colony) of T. matsutake by thinning the understory vegetation. J

Ectomycorrhizal sporocarp occurrence as affected by manipulation of litter and humus layers in Scots pine stands of different age. Appl Soil Ecol 1996; 4:

[147] Gundersen P, Callesen I, De Vries W. Nitrate leaching in forest ecosystems is related to forest floor CN ratios. Environ Pollut 1998; 102:

[148] Vaario LM, Kiikkilä O,

Soil Ecol 2013; 66: 56-60.

[149] Yun W, Hall IR. Edible

[150] Smit E, Veenman C, Baar J. Molecular analysis of ectomycorrhizal basidiomycete communities in a Pinus sylvestris L. stand reveals long-term increased diversity after removal of litter and humus layers. FEMS Microbiol

Hamberg L. The influences of litter cover and understorey vegetation on fruitbody formation of Tricholoma matsutake in southern Finland. Appl

ectomycorrhizal mushrooms: challenges and achievements. Can J Bot 2004; 82:

[151] Bogar L, Peay K, Kornfeld A, et al. Plant-mediated partner discrimination in ectomycorrhizal mutualisms. Mycorrhiza 2019; 29: 97-111.

[152] Garbaye J. Tansley review no. 76 helper bacteria: a new dimension to the mycorrhizal symbiosis. New Phytol

Jpn For Soc 1979; 61: 163-173.

[146] Baar J, Ter Braak CJF.

*DOI: http://dx.doi.org/10.5772/intechopen.95399*

hydrolytic enzymes by mycorrhizal mushrooms. Mycoscience 1995; 36:

[136] Kusuda M, Ueda M, Konishi Y, et al. Detection of β-glucosidase as saprotrophic ability from an

ectomycorrhizal mushroom, Tricholoma matsutake. Mycoscience 2006; 47:

[137] Vaario L-M, Guerin-Laguette A, Matsushita N, et al. Saprobic potential of Tricholoma matsutake: growth over pine bark treated with surfactants.

Mycorrhiza 2002; 12: 1-5.

[138] Vaario L-M, Heinonsalo J, Spetz P, et al. The ectomycorrhizal fungus Tricholoma matsutake is a facultative saprotroph in vitro. Mycorrhiza 2012; 22: 409-418.

[139] Endo N, Dokmai P, Suwannasai N, et al. Ectomycorrhization of Tricholoma matsutake with Abies veitchii and Tsuga diversifolia in the subalpine forests of Japan. Mycoscience 2015; 56: 402-412.

[140] Yamada A, Ogura T, Degawa Y, et al. Isolation of Tricholoma matsutake and T. bakamatsutake cultures from field-collected ectomycorrhizas. Mycoscience 2001; 42: 43-50.

[141] Van Der Eerden L, De Vries W, Van Dobben H. Effects of ammonia deposition on forests in the Netherlands.

Atmos Environ 1998; 32: 525-532.

[142] Nohrstedt HO. Fruit-body production and 137Cs-activity of Cantharellus cibarius after nitrogen and potassium fertilization. Forestry Research Inst. of Sweden, 1994.

[144] Richards BN. Mycorrhiza

[143] Harley JL, Smith SE. Mycorrhizal Symbiosis Academic Press. London/

development of loblolly pine seedlings in relation to soil reaction and the

221-225.

184-189.

[135] Terashita T, Kono M, Yoshikawa K, et al. Productivity of *Conservation of Edible Ectomycorrhizal Mushrooms: Understanding of the ECM Fungi… DOI: http://dx.doi.org/10.5772/intechopen.95399*

hydrolytic enzymes by mycorrhizal mushrooms. Mycoscience 1995; 36: 221-225.

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

of Pinus pinaster and P. sylvestris mycorrhizal seedlings under greenhouse conditions. Mycorrhiza 2004; 14:

Fernández-Toirán LM, et al. Mycorrhizal synthesis between Boletus edulis species complex and rockroses (Cistus sp.). Mycorrhiza 2008; 18: 443-449.

Martín-Pinto P, et al. Could artificial reforestations provide as much production and diversity of fungal species as natural forest stands in marginal Mediterranean areas? For Ecol

[128] Águeda B, Parladé J,

[129] Oria-de-Rueda JA, Hernández-Rodríguez M,

Manage 2010; 260: 171-180.

[130] Yamanaka T, Yamada A, Furukawa H. Advances in the cultivation of the highly-prized

ectomycorrhizal mushroom Tricholoma matsutake. Mycoscience 2020; 61: 49-57.

[131] Ogawa M. Microbial ecology of mycorrhizal fungus, Tricholoma matsutake Sing. in pine forest 1. Fungal colony (' Shiro') of T. matsutake. Bull Prod Res Inst 1975; 272: 79-121.

[132] Narimatsu M, Koiwa T,

35-43.

Sakamoto Y, et al. Estimation of novel colony establishment and persistence of the ectomycorrhizal basidiomycete Tricholoma matsutake in a Pinus densiflora forest. fungal Ecol 2016; 24:

[133] Hortal S, Plett KL, Plett JM, et al. Role of plant–fungal nutrient trading and host control in determining the competitive success of ectomycorrhizal fungi. ISME J 2017; 11: 2666-2676.

[134] Terashita T, Kono M. Carboxyl proteinases from Tricholoma matsutake and its related species. Mem Fac Agric

Yoshikawa K, et al. Productivity of

Kinki Univ 1989; 22:5-12.

[135] Terashita T, Kono M,

171-175.

[120] Vaario LM, Sah SP, Norisada M, et al. Tricholoma matsutake may take more nitrogen in the organic form than other ectomycorrhizal fungi for its sporocarp development: the isotopic evidence.

[121] Kawai M, Abe S. Studies on the artificial reproduction of Tricholoma matsutake (S. ito et Imai) Sing. i. effects of carbon and nitrogen sources in media on the vegetative growth of Tricholoma

[122] Yamada A, Maeda K, Kobayashi H, et al. Ectomycorrhizal symbiosis in vitro between Tricholoma matsutake and Pinus densiflora seedlings that resembles naturally occurring 'shiro'. Mycorrhiza 2006; 16: 111-116.

[123] Kibar B, Peksen A. Nutritional and environmental requirements for vegetative growth of edible

[124] Liu R-S, Li D-S, Li H-M, et al. Response surface modeling the significance of nitrogen source on the cell growth and Tuber polysaccharides production by submerged cultivation of Chinese truffle Tuber sinense. Process

Biochem 2008; 43: 868-876.

[125] Bonet JA, Oliach D, Fischer C, et al. Cultivation methods of the black truffle, the most profitable mediterranean non-wood forest product; a state of the art review. In EFI proceedings 2009; 57: 57-71.

[126] Guerin-Laguette A, Plassard C, Mousain D. Effects of experimental conditions on mycorrhizal relationships between Pinus sylvestris and Lactarius deliciosus and unprecedented fruitbody formation of the Saffron milk cap under controlled soilless conditions. Can J Microbiol 2000; 46: 790-799.

[127] Parladé J, Pera J, Luque J.

Evaluation of mycelial inocula of edible Lactarius species for the production

ectomycorrhizal mushroom Tricholoma terreum. Agriculture 2011; 98: 409-414.

Mycorrhiza 2019; 29: 51-59.

matsutake. Transactions. 1976.

**172**

[136] Kusuda M, Ueda M, Konishi Y, et al. Detection of β-glucosidase as saprotrophic ability from an ectomycorrhizal mushroom, Tricholoma matsutake. Mycoscience 2006; 47: 184-189.

[137] Vaario L-M, Guerin-Laguette A, Matsushita N, et al. Saprobic potential of Tricholoma matsutake: growth over pine bark treated with surfactants. Mycorrhiza 2002; 12: 1-5.

[138] Vaario L-M, Heinonsalo J, Spetz P, et al. The ectomycorrhizal fungus Tricholoma matsutake is a facultative saprotroph in vitro. Mycorrhiza 2012; 22: 409-418.

[139] Endo N, Dokmai P, Suwannasai N, et al. Ectomycorrhization of Tricholoma matsutake with Abies veitchii and Tsuga diversifolia in the subalpine forests of Japan. Mycoscience 2015; 56: 402-412.

[140] Yamada A, Ogura T, Degawa Y, et al. Isolation of Tricholoma matsutake and T. bakamatsutake cultures from field-collected ectomycorrhizas. Mycoscience 2001; 42: 43-50.

[141] Van Der Eerden L, De Vries W, Van Dobben H. Effects of ammonia deposition on forests in the Netherlands. Atmos Environ 1998; 32: 525-532.

[142] Nohrstedt HO. Fruit-body production and 137Cs-activity of Cantharellus cibarius after nitrogen and potassium fertilization. Forestry Research Inst. of Sweden, 1994.

[143] Harley JL, Smith SE. Mycorrhizal Symbiosis Academic Press. London/ New York. 1983.

[144] Richards BN. Mycorrhiza development of loblolly pine seedlings in relation to soil reaction and the

supply of nitrate. Plant Soil 1965; 22: 187-199.

[145] Takeshi I, Ogawa M. Cultivating method of the mycorrhizal fungus, Tricholoma matsutake (Ito et Imai) Sing. (II) increasing number of Shiro (fungal colony) of T. matsutake by thinning the understory vegetation. J Jpn For Soc 1979; 61: 163-173.

[146] Baar J, Ter Braak CJF. Ectomycorrhizal sporocarp occurrence as affected by manipulation of litter and humus layers in Scots pine stands of different age. Appl Soil Ecol 1996; 4: 61-73.

[147] Gundersen P, Callesen I, De Vries W. Nitrate leaching in forest ecosystems is related to forest floor CN ratios. Environ Pollut 1998; 102: 403-407.

[148] Vaario LM, Kiikkilä O, Hamberg L. The influences of litter cover and understorey vegetation on fruitbody formation of Tricholoma matsutake in southern Finland. Appl Soil Ecol 2013; 66: 56-60.

[149] Yun W, Hall IR. Edible ectomycorrhizal mushrooms: challenges and achievements. Can J Bot 2004; 82: 1063-1073.

[150] Smit E, Veenman C, Baar J. Molecular analysis of ectomycorrhizal basidiomycete communities in a Pinus sylvestris L. stand reveals long-term increased diversity after removal of litter and humus layers. FEMS Microbiol Ecol 2003; 45: 49-57.

[151] Bogar L, Peay K, Kornfeld A, et al. Plant-mediated partner discrimination in ectomycorrhizal mutualisms. Mycorrhiza 2019; 29: 97-111.

[152] Garbaye J. Tansley review no. 76 helper bacteria: a new dimension to the mycorrhizal symbiosis. New Phytol 1994; 128: 197-210.

[153] Paul LR, Chapman BK, Chanway CP. Nitrogen fixation associated with Suillus tomentosus tuberculate ectomycorrhizae on Pinus contorta var. latifolia. Ann Bot 2007; 99: 1101-1109.

[154] Barbieri E, Ceccaroli P, Saltarelli R, et al. New evidence for nitrogen fixation within the Italian white truffle Tuber magnatum. Fungal Biol 2010; 114: 936-942.

[155] Danell E, Alström S, Ternström A. Pseudomonas fluorescens in association with fruit bodies of the ectomycorrhizal mushroom Cantharellus cibarius. Mycol Res 1993; 97: 1148-1152.

[156] Oh S-Y, Lim YW. Root-associated bacteria influencing mycelial growth of Tricholoma matsutake (pine mushroom). J Microbiol 2018; 56: 399-407.

[157] Vaario L-M, Asamizu S, Sarjala T, et al. Bioactive properties of streptomyces may affect the dominance of Tricholoma matsutake in shiro. Symbiosis 2020; 81: 1-13.

[158] Oh S-Y, Kim M, Eimes JA, et al. Effect of fruiting body bacteria on the growth of Tricholoma matsutake and its related molds. PLoS One 2018; 13: e0190948.

**175**

**Chapter 11**

**Abstract**

nitrogen use efficiency

plants in the form of nitrate (NO3

contamination of groundwater [11].

**1. Introduction**

Promotion of Nitrogen

Growth-Promoting Rhizobacteria

Nitrogen fertilizers are one of the highest expenses in agricultural systems and usually a limitation to the productions of many agricultural crops worldwide. The intensive use of this element in modern agriculture represents a potential environmental threat, one of the many tools for the sustainable use of this resource without losing productivity is the use of plant growth-promoting rhizobacteria, especially nitrogen-fixing bacteria. However, in considering the competitiveness of the market, studies are still needed to determine the most efficient way to use this resource and if the nitrogen mineral fertilization is indeed substitutable. As a result, this study aims to deepen the scientific knowledge of the plant-microbe interactions by addressing their main characteristics and functionalities for plant growth and development and efficiency in the use of nitrogen. For this we reviewed relevant

*Gabriel Monteiro, Glauco Nogueira, Cândido Neto,* 

information from scientific works that address these issues.

**Keywords:** biochemistry, nitrogen-fixation, growth, nitrogen fertilizers,

−

Nitrogen (N) is a key component of most proteins, secondary metabolites and signaling molecules [1]. It is one of the most important macronutrients for plant development and usually one of the most limiting factor to plant production [2]. The use of N-fertilizers has produced a significant increase in food production in recent decades [3], and its consumption has grown from 11.3 Tg N year−1 in 1961 to 107.6 Tg N year−1 in 2013 [4]. However, less than 50% of the added N is effectively absorbed by most cultivated plants [5, 6], and even N effectively converted to biomass, eventually returns to the environment [7]. In the soil, N is available to

), ammonium (NH4

(usually amino acids), being the NO3, the most abundant [8]. In its ionic form, NO3

has a negative charge and high water solubility, being susceptible to leaching and runoff [9]. It can also be volatilized by denitrifying microorganisms [10], and lost to the atmosphere in the form of nitrous oxide (N2O, a greenhouse gas 296-fold more potent than a unit of CO2). Leaching of N causes eutrophication of water bodies and

+

) and organic compounds

−

Assimilation by Plant

*Vitor Nascimento and Joze Freitas*

#### **Chapter 11**

*Nitrogen in Agriculture - Physiological, Agricultural and Ecological Aspects*

[153] Paul LR, Chapman BK, Chanway CP. Nitrogen fixation associated with Suillus tomentosus tuberculate ectomycorrhizae on Pinus contorta var. latifolia. Ann Bot 2007; 99:

[154] Barbieri E, Ceccaroli P, Saltarelli R, et al. New evidence for nitrogen fixation within the Italian white truffle Tuber magnatum. Fungal Biol 2010; 114:

[155] Danell E, Alström S, Ternström A. Pseudomonas fluorescens in association with fruit bodies of the ectomycorrhizal mushroom Cantharellus cibarius. Mycol

[156] Oh S-Y, Lim YW. Root-associated bacteria influencing mycelial growth of Tricholoma matsutake (pine mushroom). J Microbiol 2018; 56:

Sarjala T, et al. Bioactive properties of streptomyces may affect the dominance of Tricholoma matsutake in shiro.

[158] Oh S-Y, Kim M, Eimes JA, et al. Effect of fruiting body bacteria on the growth of Tricholoma matsutake and its related molds. PLoS One 2018; 13:

Res 1993; 97: 1148-1152.

[157] Vaario L-M, Asamizu S,

Symbiosis 2020; 81: 1-13.

1101-1109.

936-942.

399-407.

e0190948.

**174**
