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Production of edible mushrooms in forests:

Trends in development of a mycosilviculture

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Appl Microbiol Biotechnol (2011) 89:971–979
DOI 10.1007/s00253-010-3022-4
MINI-REVIEW
Production of edible mushrooms in forests: trends
in development of a mycosilviculture
Jean-Michel Savoie & Michèle L. Largeteau
Received: 21 October 2010 / Revised: 17 November 2010 / Accepted: 17 November 2010 / Published online: 4 December 2010
# Springer-Verlag 2010
Abstract Developing the production of ectomycorrhizal
(ECM) mushrooms in forest has become a challenge. Only
a few ECM species are currently cultivable. Controlled
mycorrhization practices offer promising advance to pro-
duce currently uncultivable ECM mushrooms. The persis-
tence of the production of edible species, either cultivated
or wild, depends on both the tree and the ecological
environment (fungal communities, climate, soil, tree develop-
ment). Developing adapted forest management practices
appears to be means to improve production of edible ECM
mushrooms. This review summarises current knowledge on
the development of a science-based mycosilviculture for the
production of edible ECM mushrooms.
Keywords Controlled mycorrhization . Ectomycorrhizal
mushrooms . Forest management . Fungal succession
Introduction
Mushrooms were defined by Chang and Miles (1992) as
macrofungus with a distinctive fruiting body that is large
enough to be seen with the naked eye and to be picked up
by hand. According to this definition, mushrooms can be
either Basidiomycetes or Ascomycetes, aerial or under-
ground (Chang 1993). Mushrooms can be roughly divided
into various categories depending on their ecology or their
effects on human health. Saprophytic mushrooms play an
important role in the cycling of carbon and other elements
through the breakdown of lignocellulosic plant residues and
J.-M. Savoie (*) : M. L. Largeteau
INRA, UR1264, MycSA,
33883 Villenave d’Ornon, France
e-mail: savoie@bordeaux.inra.fr
animal dung, whereas mycorrhizal mushrooms are involved
in symbiotic associations with plant roots. Apart from
edible mushrooms, that might have been used by man as
food since his earliest beginnings, there are medicinal
mushrooms originally applied in daily medicinal practice in
China at least 5,000 years ago (Van Griensven 2008); and a
third category contains poisonous or suspected poisonous
mushrooms. There are more than 10,000 known species of
fleshy macrofungi, but only a handful are lethal (Kendrick
1985), and the properties of a large number of mushrooms
remain not defined (Chang 1993).
According to historical records of intentionally cultivated
mushrooms, the first one was Auricularia auricula around
600 A.D. in China (Chang 1993). Significant cultivation of
edible mushrooms started around 700 years ago in China
with the cultivation of Shii-take (Lentinula edodes) on
inoculated wood logs placed in forests. During the 18th
century, cultivation of the button mushroom (Agaricus
bisporus) spread all over Europe. It was based on flat
cultivation beds consisting of mixtures of weathered horse
manure placed in caves (Van Griensven 2008). According to
Chang (1993), about 80 of 2,000 species which are regarded
as potentially edible mushrooms have been experimentally
grown, 40 economically cultivated, around 20 commercially
cultivated, and 5 to 6 have reached an industrial scale in
many countries. Today, most of the mushrooms recognised
as cultivated are saprophytic species. Some of them can be
produced in forest on inoculated wood logs or other
substrates, but this outdoor culture is dependent on local
climatic conditions, and generally, they are cultivated indoor.
Besides, forest planning and management has paid little
or no attention to the harvesting of wild edible fungi for a
long time. Because of a decline in forest-based industries in
some countries, wild mushrooms are now considered as
new sources of income (Román and Boa 2006). A new
972
challenge is the development of a science-based production
of ectomycorrhizal (ECM) mushrooms. Of more than 140
ECM species regarded as edible, only a small percentage
has been cultivated on a commercial scale (Salerni and
Perini 2004). The management of forest stands for
mushroom production is called mycosilviculture and is of
increasing importance in Europe. There is still a need for
research and communication of results. The aim of the
present review is to contribute to a science-based develop-
ment of the mycosilviculture for the production of edible
ECM mushrooms.
Cultivation of ECM in forest?
The production of mushrooms in forests may result from
natural processes, and picking generates important incomes
for local populations. In a recent review, Wang and Hall
(2004) stated that over the past 100 years, harvests of many
mycorrhizal mushrooms have declined dramatically. This
decrease has been attributed to indirect effects of air
pollution (in particular to increases in nitrogen deposition,
possibly in combination with acidification) and to accumu-
lation of litter and humus (Arnolds 1991; Smit et al. 2003).
This decline has prompted interest in the development of
methods for cultivating mycorrhizal mushrooms. Cultiva-
tion of edible ECM mushrooms is difficult because of the
slow in vitro growth rate of the mycelium on nutrient
media, the absence of fruiting body formation without
mycorrhiza formation with a host plant, the difficulty for
sustaining a mycorrhizal association with a host plant, and
lack of information on fruiting mechanisms.
The prime condition for the development of an ECM
mushroom species in a natural environment is the presence
of an inoculum and its ability to be disseminated in
appropriate places. Arrival in an ecosystem can be
manipulated by intentional introduction of mycelium or
spores. However, the inoculum potential of the species,
which can be defined as the energy of growth available for
colonisation of a compatible tree root, is governed by
factors such as propagule density, mycelium age and
quality, ability to reach a target root from a distant point.
Inoculum potential can be favoured by introducing prop-
agules in soil close to tree roots. However, roots of
uninoculated trees at planting are colonised quickly by
indigenous mycorrhizal fungi, often without interest as food
product, and there is a great competition with the
introduced propagules. Consequently, it seems more effi-
cient to use artificial mycorrhization before tree planting in
forests.
Inoculation with specific mycorrhizal fungi began in
French truffle culture 200 years ago. Joseph Talon
Appl Microbiol Biotechnol (2011) 89:971–979
discovered that truffle “orchards” could be established on
certain sites by planting them close to oaks (Malençon
1938). That stimulated French and Italian scientists to try
various inoculation methods and to understand the life
cycle of truffles. Thanks to this research effort, more was
known about the ecology and management of truffles and
their host trees than of any other ECM association (Grente
and Delmas 1974). The recent sequencing of the Perigord
black truffle (Tuber melanosporum) genome will increase
this knowledge (Martin et al. 2010). Apart from these
hypogeous edible mushrooms for which various reviews
were written (Olivier 2000; Bonet et al. 2007), cultivation
of epigeous edible ECM mushrooms has been less
successful in terms of application. So far, only a few
species of truffles have been produced in commercial
quantities, although methods have been developed for the
large-scale cultivation of other species, such as Canthar-
ellus cibarius, Lyophyllum shimeji, Lactarius deliciosus,
and Suillus granulatus (Poitou et al. 1989; Otha 1994;
Danell and Camacho 1997). Experiments are in progress
for these species. Most significantly, many of the most
expensive mycorrhizal mushrooms, including Tuber mag-
natum and Tricholoma matsutake, have defied cultivation
(Wang and Hall 2004). The comparison of genomic traits
in two ECM fungi (Laccaria bicolor, T. melanosporum)
showed that genetic predispositions for symbiosis appar-
ently evolved in different ways in Ascomycetes and
Basidiomycetes (Martin et al. 2010), and it might be
difficult to use the knowledge and experience cumulated on
truffles for cultivation development of epigeous ECM
Basidiomycetes.
After a mycorrhizal association with a host plant, the
sustainability of the symbiosis could be improved by forest
management aimed at stimulating the development of the
edible mushroom. In Japan, traditional management of
forests that produce Matsutake or pine mushrooms
(T. matsutake) can include thinning of overstory and
understory vegetation and manipulation of the forest floor.
Even with a sustainable symbiosis, the production of
mushroom is not guaranteed. A major problem up to know
is that our current knowledge on the biological processes of
fruiting body initiation and development is limited. Envi-
ronmental conditions play a crucial role on the determina-
tion whether a fruiting body will be formed, but there is no
universal set of conditions that leads to fructification in all
fungi (Kües and Liu 2000). Observations of the natural
environment of the fungus may give valuable indication as
how to proceed in forest management.
The cultivation of epigeous edible ECM mushrooms is
dependent on both improvements in controlled mycorrhiza-
tion and adaptation of forest management to the ecological
requirements of the selected species.
Appl Microbiol Biotechnol (2011) 89:971–979
Controlled mycorrhization
There is a considerable body of literature on techniques
for producing tree seedlings infected with competitive
and efficient ECM fungi (Le Tacon et al. 1992) but
mainly for tree growth helpers that are not edible or have a
poor interest as food. Besides, few data show that
infection with efficient ECM fungi can be obtained after
inoculation of seedlings at the time of field planting or
direct inoculation in the field a few days to a few months
after planting (Duñabeitia et al. 2004). Duponnois and
Garbaye (1991) proposed techniques for controlled syn-
theses of ECM symbiosis that may be adapted to various
systems. They included different steps: fungal inoculum
preparation, seed treatment, choice and treatment of
substrates, choice of nutrient solutions, aseptic or non-
aseptic experimental systems, and bare-root nursery
techniques. In fact, most of the techniques that have been
developed for infecting plants with edible mycorrhizal
mushrooms used seedlings raised free of contaminating
ECM fungi before infecting them with the fungus of one's
choice. On another hand, inoculant production of ECM
fungi is a critical point that has recently been reviewed
(Rossi et al. 2007).
Following similar approaches, in vitro controlled
mycorrhization techniques were developed for several
epigeous edible mushrooms, but because controlled
mycorrhization techniques take many years to perfect,
they are jealously guarded by the companies that have
developed them. For instance, the establishment of
plantations with L. deliciosus or Suillus luteus—inoculat-
ed Pinus pinaster seedlings to obtain fruiting bodies—was
first attempted by Poitou et al. (1984). The evolution of
these plantations was investigated by Guinberteau et al.
(1989) who reported the production of sporocarps in the
stand 3 years after outplanting. Further attempts at
producing quality plants of Pinus halepensis, Pinus pinea,
and Pinus sylvestris mycorrhized with L. deliciosus and
Lactarius sanguifluus under nursery or greenhouse con-
ditions were reported (Torres and Honrubia 1994; Rincon
et al. 1999; Guerin-Laguette et al. 2000), but complements
with outplanting studies were not reported until the work
of Parladé et al. (2004). However, during this time,
commercial production of mycorrhized trees have been
developed. For instance, a French nursery developed
strains and mycorrhization techniques for the production
of Pinus trees mycorrhized with S. luteus, L. sanguifluus,
and L. deliciosus. From the early 2000s, dozens of
orchards were planted in the south of France and in Italy,
and they produce edible mushrooms with some yields of
80 kg/ha the sixth year after outplanting 2-year-old
seedlings (http://www.robinpepinieres.com). Wang and
973
Hall (2004) reported from the literature that a plantation
of P. pinaster in North Otage, NZ, produces 50 kg of L.
deliciosus per hectare.
Mycorrhiza helper bacteria
The formation of ECM symbiosis can be significantly
improved by selected soil and mycorhizosphere bacteria
(Bacillus, Burkolderia, Paenibacillus, Pseudomonas, Strep-
tomyces) named “mycorrhiza helper bacteria” (MHB), and
believed to be ubiquitous in their distribution (Garbaye
1994; Poole et al. 2001; Deveau et al. 2007). Different
mechanisms may operate for different bacteria to promote
mycorrhiza formation (Poole et al. 2001; Aspray et al.
2006). Direct effects of MHB on presymbiotic survival and
growth of various ECM (L. bicolor, Pisolithus albus,
Amanita muscaria) in the soil have been well documented
(Frey-Klett et al. 1997; Brulé et al. 2001; Founoune et al.
2002; Schrey et al. 2005). Changes in A. muscaria gene
expression upon interaction with Streptomyces sp. have
been observed (Schrey et al. 2005; Riedlinger et al. 2006).
Some L. bicolor pathways are mutually regulated by
different rhizobacteria, whilst others are specific to some
MHB (Deveau et al. 2007). MHB seem to be fungal-
specific, and not plant-specific (Garbaye and Duponnois
1992; Poole et al. 2001).
The major part of available documentation on MHB
concerns ECM used as model to understand mycorrhizal
fungi development or/and useful for tree development.
Little information concerns edible ECM. Danell and
Camacho (1997) obtained in vitro mycorrhizal association
between C. cibarius and Pinus seedlings. Fruiting bodies
developed on 16-month-old plants kept in greenhouse, and
great quantities of Pseudomonas were present in the
fruiting bodies, as observed in wild strains of C. cibarius.
That leaded the authors to wonder whether these bacteria
might contribute to the mycorrhizal association. However,
5 years later, Danell (2002) reported that experimental
plots of C. cibarius have failed to produce fruiting bodies,
even though mycorrhizas have spread onto new roots.
Mello et al. (2010) identified a Moraxella osloensis
population associated with productive sites of the white
truffle T. magnatum. This finding open the question as to
whether M. osloensis had mediated mechanisms that might
modify fruiting body production of this uncultivated
truffle species and if the bacteria could be used as MHB.
One example of MHB used successfully with an edible
ECM is Bacillus subtilis which significantly stimulated
the mycorrhizal formation of the edible fungus S.
granulatus on Pinus thunbergii seedling roots (Kataoka
et al. 2009).
974
Host plant cloning
Intraspecific variations in the ability of host plants to form
ectomycorrhizas have been well documented. The signifi-
cance of genetic variation in the host plant with genetically
uniform plant material was studied by Van der Heijden and
Kuyper (2001). In their experiment, the origin of Salix
repens clones (cuttings) had a large effect on the effective-
ness of mycorrhizal symbiosis with Hebeloma leucosanx
and Pisolithus tinctorius. Tonkin et al. (1989) observed
differences in clonal lines of Eucalyptus marginata for in
vitro development of P. tinctorius mycorrhizas. However,
to our knowledge, there is only one available report on the
use of cloned seedling for mycorrhizal association with an
edible fungus. In this study, higher percentages of T.
melanosporum mycorrhizas were obtained with cloned
Corylus avellana L. seedlings compared to uncloned plants.
Heterogeneity of the root volume was not completely
suppressed, but very extensive development unfavourable
to Tuber colonisation rarely occurred. Cloning the host
plant improved the quality of the controlled mycorrhization
(Mamoun and Olivier 1996). This kind of approach could
be also useful for the improvement of mycorrhization with
basidiomycetes.
Fungal succession management
As plants grow, changes in succession of the associated
fungi may occur. Competitors may belong either to early-
or to late-stage fungi by their occurrence in young and old
plantations, respectively. The succession of ectomycorrhi-
zas in growing plants is well documented. ECM fungi are
considered as indicators of succession stage of forests
(Frankland 1998). For instance, some ECM fungi, e.g.
Laccaria species, dominate in the early-stage of succession
and have broad host range, whereas more host-selective
ECM fungi, e.g., species of Suillus and Lactarius, are
generally considered as late-stage species (Frankland 1998).
Fungi were also found to be associated with certain parts of
the root system, i.e., the younger roots with early
succession species, and the older part of the root system
with late-stage species (Ford et al. 1980; Last et al. 1992).
Interestingly, the most abundantly fruiting species formed
less than 5% of the mycorrhizal roots examined in a
detailed study in a plantation of Sitka spruce (Taylor and
Alexander 1990). Optimisation of mushroom production in
forest comes down to answer the question: how to
manipulate this succession for having both a dominant
mycorrhization by the chosen fungal species and optimal
environment conditions for fruiting body development?
Investigation carried out in 2007 on natural truffières
located in woods (P. pinaster, P. pinea, and Quercus ilex)
Appl Microbiol Biotechnol (2011) 89:971–979
established between 1933 and 1938 revealed that Tuber
borchii formed 20% of ectomycorrhizas and was dominated
by large numbers of rare species and a few common species
(Thelephoracae, Inocybaceae, and Sebacinaceae). Tuber
dryophilum is a common, poorly flavoured truffle compet-
ing with T. borchii (Lotti et al. 2010). Competition between
cultivated Tuber spp. and truffles of less or no commercial
interest is a major problem during the first few years after
planting. Tuber aestivum and Tuber brumale Vittad. often
replace T. melanosporum in French, Italian, and Spanish
truffle orchards (Donnini et al. 2001; Sourzat et al. 2001;
García-Montero et al. 2008).
Early- and late-stage fungi may be equally efficient in
developing mycorrhiza with roots. However, they vary in
their nutrient demand (Dighton and Mason 1985). The late-
stage fungi, unlike early-stage ones, are unable to form
mycorrhizas while growing in first rotation in unsterile
forest soil, owing to their lower competitive ability
compared to other microorganisms. Among the 12 ECM
species tested by Ishida et al. (2008), all the second- and
third-stage fungi, except Hebeloma spp., exhibited lower
germination rates and infectivity of 1–2 month-old Salix
seedlings compared to first stage fungi. Components
released only from mature tree roots might be necessary
to late-stage ECM to form mycorrhizas. The effect of
mycelium and mycorrhizas of early-stage ECM might also
be considered as potential stimulators.
Poitou et al. (1989) investigated both the fructification of
ECM fungi previously inoculated to P. pinaster seedings in
nursery and the development of natural competitors. Seed-
lings inoculated with S. granulatus and L. deliciosus started
to produce fruiting bodies 2 and 3 years, respectively, after
outplanting in a sandy–clayey soil. Fruiting bodies also
developed 2 and 4 years after outplanting, respectively,
under P. pinaster uninoculated or inoculated with mycor-
rhizal fungi poorly adapted to the soil characteristics,
proving the ability of S. granulatus and L. deliciosus to
rapidly infect the surrounding expanding root sytems. S.
granulatus sporophores developed during spring (April–
June) and autumn (end of August–October), whilst L.
deliciosus showed a single fruiting period (end of October–
beginning of December). During the 7-year period ana-
lysed, the natural colonisers had no effect on L. deliciosus
and S. granulatus yields. The fruiting area moved away
from the stem every year. Correlations between the stem
circumference and the distance between sporophores and
stem separated two sets of ECM fungi. Similar correlations
were obtained with (1) S. granulatus, S. bovinus, Gomphi-
dius viscidus, and Hygrophorus hypothejus, and (2) L.
deliciosus, Rhizopogon luteolus, and Tricholoma pessunda-
tum. Correlations meant that S. granulatus and L. deliciosus
colonised different root sites, which was evident from trees
producing both fungi in autumn. The small distance
Appl Microbiol Biotechnol (2011) 89:971–979
between trees did not allow analysing the mushroom
production the following years. However, these observa-
tions showing that ECM fungal diversity increases with
stand age agree with the results of Twieg et al. (2007).
In pine forests, S. granulatus had the characteristic of
both early- and late-stage fungi. Rao et al. (1997) related
the absence of late-stage fungi from younger pine stands to
the low amount of pine litter and less carbohydrate than in
older stands. In P. sylvestris forest in the Pyrenees, France,
L. deliciosus proliferates across young, medium, and
mature forests (5–84 years) (Bonet et al. 2004). In Oregon,
USA, L. deliciosus fruiting bodies were found in Pseudot-
suga menzesii forest of 30–50 years, but absent in old forest
of 400+ years (Smith et al. 2002). On the parcel planted by
Poitou et al. (1989), L. deliciosus and S. granulatus
continue to produce fruiting bodies in autumn 20 years
later.
After outplanting, in addition to the presence of indige-
neous competitors, the physical–chemical and biotic charac-
teristics of the soil also conditioned the persistence and spread
of the cultivated fungus mycorrhizas, as Hortal et al. (2009)
observed for L. deliciosus.
ECM ecology and forest management
ECM communities contribute to a number of key ecosys-
tem functions in forest (for a review, see Courty et al.
2010). Forest practices can affect the occurrence, produc-
tivity, and reproduction of mushrooms. Understanding the
ecology of ECM and the effect of forest management
practices could be the means to improve natural mushroom
production in forest stands planted with edible ECM
inoculated trees. Forest management techniques, soil
inoculation with spores, and retaining some of the mature
fruiting bodies were used to maximise T. matsutake natural
production in Japan forests (Wang and Hall 2004). Bonet et
al. (2008, 2010) developed a predictive model that could
facilitate forest management decisions. In P. sylvestris
plantations of Central Pyrenees, Spain, the production of
edible and marketed mushrooms, including Lactarius spp.,
is related to the stand basal area, elevation, aspect, and
steepness of slope. Therefore, it is important, as notified by
the authors, to extent the network of mushroom plots to
cover different tree species, sites, stand ages, and densities.
A 20-year inventory of mushroom productivity in mixed
forest in south-western Switzerland reveals a clear temporal
relationship between thinning, tree growth reaction, and the
reaction of the associated fungal community. Thinning
favours strongly growing trees with a high photosynthetic
capacity which tend to produce more ECM fruiting bodies
than poorly growing trees. Many of the edible mushrooms
occurred exclusively after thinning: Amanita rubescens,
975
Boletus edulis, Craterellus cornucopioides, Hypholoma
capnoides, some Russula species, and Xerocomus badius.
Unexpectedly, the fruiting body production of non-
mycorrhizal species also increased after thinning (Egli
et al. 2010). Chanterelle production had been decreased
by thinning but recovered in the following 6 years (Pilz
et al. 2006). In woods with a majority of Abies alba Miller
(four stations located on the Sirna side of Mt Amiata,
southern Tuscany, Italy), the major number of B. edulis
fruiting bodies were counted in medium thinning stations,
which indicates that this species does not need a dense
canopy but an open sunny wood habitat to maintain and/or
increase its productivity (Salerni and Perini 2004).
Unfavourable climatic conditions could also be specifi-
cally corrected by human action if the needs of the species
are known. Thanks to recordings for more than 10 years in
Aquitaine, southwest of France, it was observed that the
main factor inducing the fructification of Boletus species is
a rainfall leading at least a 40% increase in soil moisture,
and that the gap between the rainfall and the fruiting body
production can vary from 6 to 25 days, depending on the
soil temperature. For B. edulis, a cold shock of more than
5°C is necessary for having the benefit of the inducing
rainfall (Leprince and Guinberteau, pers. comm.). Water
and temperature requirements vary with the fungal species
(Martínez de Aragón et al. 2007) and could be modified by
thinning practices inducing modification of rain and
sunshine penetration in the forest canopy but also by
watering to improve the productivity of fungi of important
economic value.
Removal of herbaceous vegetation, litter, or humus influ-
ences the production of mushrooms by having positive effects
on species richness of ECM fungi (Baar and Kuyper 1998;
Smit et al. 2003) but negative effects on the fruiting process
of a specific species as B. edulis (Salerni and Perini 2004).
Liming, a forestry practice used to counteract forest
decline in acidic soils, durably impacts ECM fruiting body
development. Fifteen years after treatment in parts of
declining stands of 35-year-old Picea abies and 60-year-
old Fagus sylvatica (Vosges mountains, France), liming has
resulted in a shift from a typical acidophilic forest fungal
community to a less typical one, and apparition of many
late-stage forest species (Rineau et al. 2010).
Dynamics of population composition of forest fungi is
influenced by the age of the associated host trees. Reports
of forest mushroom yields by age class are extremely
variable across forest types (Bonet et al. 2004), so advices
for forest management practices would be restricted to the
studied areas. For instance, L. sanguifluus has higher
production in young pine forests (<50 years) of the pre-
Pyrenees mountains. Consequently, in this region, the forest
management for this species must be directed as maintain-
ing young forest stands (Martínez de Aragón et al. 2007).
976
Pinus reforestation in degraded soils in abandoned
farmlands in Mediterranean dry areas appeared to be means
to achieve a natural forest which provides fungal produc-
tion and diversity as high as those found in surrounding
natural forest stands of Quercus and Populus (Oria-de-
Rueda et al. 2010). Nevertheless, differences in fungal
communities were observed. Laccaria laccata, Lepista
sordida, Suillus bellini, Suillus collinitus, S. granulatus,
and S. luteus were edible and/or marketable fungi found
only in the reforestation stands, whilst Agaricus campestris,
Agaricus sylvicola, Coprinus comatus, Coprinus disemina-
tus, and Lepista nuda, collected in Quercus and/or Populus
forests, were absent. This study illustrates the fact that the
quality of the soil and history of its cultivation influence the
natural development of ECM mushrooms that might
compete with the inoculated ones in new forest stands.
Mycorrhization and cultivation of new species
In recent years, successful artificial mycorrhizations were
developed with edible ECM fungi currently known as
uncultivable. For instance, under in vitro conditions, the
edible fungi Tricholoma portentosum AT615, Tricholoma
saponaceum AT616, Rhizopogon rubescens AT630, and
Lactarius akahatsu AT583 formed mycorrhizas with Pinus
densiflora. Basidiocarps developed 5–9 months after
transplantation of the mycorrhized seedlings in open-pot
soil (Yamada et al. 2001). A method that could provide a
means to initiate, propagate, transplant, and sustain mycor-
rhizas of T. matsutake was developed by Yamada et al.
(2006). In vitro T. matsutake–P. densiflora mycorrhizas
grew and developed for up to 12 months and formed
mycelium–soil aggregates called “shiro” that is essential for
the production of fruiting bodies.
Astraeus is the most popular and the most expensive (7–
10 US$/kg) edible ECM mushroom group consumed in
Thailand. Fangfuk et al. (2010) reported the first in vitro
mycorrhization between Astraeus hygrometicus and P.
densiflora. Either 1-week-old sterilised P. densiflora seed-
lings transplanted into autoclaved forest soil and inoculated
with A. hygrometicus spores, or 3–7 day-old aseptically
germinated seedlings placed in vermiculite-peat moss
mixture-MNC medium containing A. hygrometicus myceli-
um formed ectomycorrhizas.
B. edulis sensu lato (B. edulis Bull.:Fr. sensu stricto,
Boletus aereus Bull.; Fr., Boletus pinophilus Pilat et
Dermek, and Boletus reticulatus Schaeff.) is a species
complex of commercial mushrooms consumed worldwide
that have defied cultivation to date. World production and
in-season retail market are estimated to be 20,000–
100,000 t and >250 million US$ (Hall et al. 2003). The
interest in its cultivation is increased by the fact that B.
Appl Microbiol Biotechnol (2011) 89:971–979
edulis dry mushroom extract showed high antioxidant
properties, enabling possibility of its use as natural source
of antioxidant in pharmaceutical or food industry (Vidović
et al. 2010). Plantations of Castanea sativa and Pinus
uncinata inoculated with B. edulis and B. aereus in the
Pyrénées atlantiques and Dordogne regions (Olivier et al.
1997) failed to significantly produce Boletus fruiting
bodies. Meotto et al. (1999) observed that 6 years after
ouplanting, B. edulis mycorrhizas persisted on 82% of the
Boletus-inoculated C. sativa, and Amanita caesarea my-
corrhizas on 88% of the Amanita-inoculated plants, but no
fruiting body production was observed. At the same time,
Scleroderma areolatum mycorrhizas increased notably over
the first sampling performed 4 years after outplanting. The
proliferation of indigenous ECM fungi probably explains
why fruiting body production has not been reported after
that. However, Peintner et al. (2007) characterised the
fungal communities in a C. sativa forest producing large
quantities of B. edulis fruiting bodies and concluded that
factors triggering formation of mycorrhizas and fructifica-
tion of B. edulis s.l. appear too complex to be simply
explained on the basis of the amount of fungal mycelia in
the soil. Recently, in an experiment without mycorrhization,
high production of these mushrooms was found associated
with 3-year-old Cistacea plants in Northwestern Spain
(Oria-de-Rueda et al. 2008). This much earlier onset
compared to fruiting associated with Castanea, Quercus,
and Pinus that are generally observed in older plantations
suggests behaviour as early-stage mycorrhizal mushroom
when associated with Cistacea. Águeda et al. (2008)
succeeded in the in vitro mycorrhization of Cistus ladanifer
and Cystus albidus with B. aereus, B. edulis, and B.
reticulatus. That might be a promising way to cultivate
these mushrooms with less risk of mycorrhizas replacement
by competitors after outplanting of inoculated Cistus
compared to Pinus or Castanea plantations. Cistaceae are
also interesting host for Terfezia spp., called desert truffles.
They where shown in semi-arid ecosystems of the
Mediterranean Basin, especially in the Iberian Peninsula
and in Italy, in association with members of the Cistaceae,
mainly with Helianthemum species (Díez et al. 2002; Rana
et al. 2007). Association of Terfezia spp. was successfully
obtained with various Cistaceae (Fortas and Chevalier
1992; Morte et al. 2000).
Wang and Hall (2004) observed that rhizomorphs of B.
edulis were often closely wound around Amanita root tips,
forming composite mycorrhizas. The authors hypothesised
that nutrients may flow between the two species, which
might be necessary for B. edulis to fruit. Other fungal
associations favourable to the production of B. edulis
mushrooms might be expected as B. edulis, and other
ECM with an ecological convergence are frequently found
fruiting close to each other. Co-mycorrhization of a late-
Appl Microbiol Biotechnol (2011) 89:971–979
stage edible mushroom with an early-stage partner have
been obtained in vitro (Guinberteau, pers. comm.) and
could be a new research way for improving the efficiency
of mushroom production in forest.
Mycosilviculture for the production of edible ECM
mushrooms
The development of a mycosilviculture for the production
of edible ECM mushrooms in forests is dependent on the
possibility to cultivate new species that have defied
cultivation to date. It will be possible with improvements
in controlled mycorrhization and adaptation of forest
management to the ecological requirements of the selected
species. In addition to works on the fungal biodiversity in
order to find efficient strains, research efforts should focus
on the role of mycorrhiza helper bacteria in controlled
mycorrhization before tree planting in forests and in the
sustainability of the symbiosis. Taken into account the
ecology of the targeted mushrooms being late-stage ECM,
recent works show that one can develop avoiding strategies
based on co-inoculation with early-stage ECM for a
favourable association or identification of plant species
having a short life cycle and favourable to early colonisa-
tion by late-stage ECM with less risk of mycorrhizas
replacement by competitors. Cloning the host plant could
improve the quality of the controlled mycorrhization and
the sustainability of the symbiosis. The combination of
these biotechnological progresses and forest managements
based on the ecology of the ECM offers new opportunities
for the production of edible mushrooms in forest.
Acknowledgments We thank our colleague J. Guinberteau for
providing information on Boletus edulis production and his unpub-
lished data.
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