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The medicinal fungus Cordyceps militaris: Research and development

Article  in  Mycological Progress · May 2012

DOI: 10.1007/s11557-012-0825-y

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Mycol Progress
DOI 10.1007/s11557-012-0825-y
REVIEW
The medicinal fungus Cordyceps militaris: research
and development
Bhushan Shrestha & Weimin Zhang & Yongjie Zhang &
Xingzhong Liu
Received: 20 February 2012 / Revised: 20 April 2012 / Accepted: 7 May 2012
# German Mycological Society and Springer 2012
Abstract Ophiocordyceps sinensis (syn. Cordyceps sinen-
sis) is a highly valued medicinal fungus. This entomopath-
ogen has a limited distribution, has been overharvested
in the wild, and its stromata have not been artificially
cultivated. Another entomopathogenic fungus, Cordy-
ceps militaris (commonly known as orange caterpillar
fungus), has chemical capacities similar to those of O.
sinensis, but unlike O. sinensis, its stromata can be
easily cultivated. Consequently, C. militaris is being
studied as an alternative to O. sinensis, and the large-
scale production of stromata is receiving substantial
attention. Significant research has been conducted on
the genetic resources, nutritional and environmental
B. Shrestha
Green Energy Mission/Nepal,
Ghatte Kulo, Anam Nagar,
Kathmandu, P.O. Box 10647, Nepal
W. Zhang
Guangdong Institute of Microbiology,
Guangzhou 510070, China
Y. Zhang
School of Life Sciences, Shanxi University,
Taiyuan 030006, China
X. Liu (*)
State Key Laboratory of Mycology, Institute of Microbiology,
Chinese Academy of Sciences,
No.3, 1st West Beichen Road, Chaoyang District,
Beijing 100101, China
e-mail: liuxz@im.ac.cn
Present Address:
B. Shrestha
Mushroom Research Division, National Institute of Horticultural
and Herbal Science, Rural Development Administration,
Suwon 441-707, Korea
requirements, mating behavior, and biochemical and
pharmacological properties of C. militaris. The complete
genome of C. militaris has recently been sequenced.
This fungus has been the subject of many reviews, but
few have focused on its biology. The current paper
reviews the biological aspects of the fungus including
host range, mating system, cytology and genetics,
insect- and non-insect nutritional requirements, environ-
mental influence on stroma development, and commer-
cial development.
Keywords Cultivation . Entomopathogenic fungus . Mating
system . Pharmaceutical application
Historical review
Medicinal fungi have long been an important part of human
culture and civilization, and species in the genus Cordyceps
are especially valued (McKenna et al. 2002). Most Cordy-
ceps species s.l. are parasites of insects or other arthropods
(Kobayasi 1941, 1982). Ophiocordyceps sinensis (Berk.)
G.H. Sung et al. (syn. Cordyceps sinensis (Berk.) Sacc.),
for example, is a parasite of caterpillars (Wang and Yao
2011) and is naturally distributed in the Tibetan Plateau of
China and surrounding high-altitude grasslands of Nepal,
Bhutan, and India (Shrestha et al. 2010; Zhang et al. 2012b).
As a highly valued medicinal fungus, O. sinensis has a long
history of being collected and traded (Jones 1997; Halpern
1999). Recent studies have shown that the natural popula-
tions of O. sinensis are decreasing because of over-
collection (Li et al. 2006c; Stone 2008; Zhang et al.
2012b). Because of the scarcity and high price of wild O.
sinensis specimens, many counterfeit materials including
cultured products are sold in the market, resulting in the

need for their authentication and quality control (Hsu et al.
2002; Hu et al. 2003; Li et al. 2004b, 2006c; Kuo et al.
2005, 2006; Zhao et al. 2006b; Ikeda et al. 2008; Paterson
2008; Chan et al. 2011; Chen et al. 2011a; Dong and Yao
2011; Liu et al. 2011; Au et al. 2012). Recently developed
novel molecular markers (Zhang et al. 2012a) will help
studying the population genetics of O. sinensis and will
possibly determine contaminants in its culture-based prod-
ucts. According to recent studies, there is substantial mor-
phological variation in the teleomorphic and anamorphic
stages of O. sinensis (Jiang and Yao 2002, 2003; Xiao et
al. 2009; Shrestha et al. 2010; Dong and Yao 2011; He et al.
2011a). Interestingly, stroma of O. sinensis in the wild is
found associated with a rich microflora (Jiang and Yao
2005; Wang et al. 2006; Zhang et al. 2010b, c) of which
the medicinal values are still not known.
A common Cordyceps species, C. militaris (L. : Fr.)
Link, is distributed worldwide from 0 to >2,000 m a.s.l.
(Kobayasi 1941; Mains 1958; Panigrahi 1995; Shrestha and
Sung 2005; Ma et al. 2007). Phylogenetically, C. militaris is
the type species of the newly emended genus Cordyceps in
the family Cordycipitaceae (Sung et al. 2007). This species,
commonly known as orange caterpillar fungus, is easily
cultured in both solid and liquid media and with a variety
of carbon and nitrogen sources. C. militaris has been in-
creasingly viewed as a substitute for O. sinensis because of
their similar chemical capacities and medicinal properties
(Gong et al. 2006a, b; Huang et al. 2006, 2009; Yu et al.
2006; Ni et al. 2007; Gao 2008; Yue et al. 2008; Zhou et al.
2009b; Das et al. 2010; Khan et al. 2010; Li et al. 2010;
Zheng et al. 2011b; Dong et al. 2012); however, more
systematic pharmacological studies should be carried out
and pure compounds should be identified in addition to
cordycepin (Paterson 2008). It is the stroma (plural stroma-
ta) of O. sinensis and C. militaris that is valued, which
contains other metabolites than the vegetative mycelium.
Because C. militaris can complete its life cycle when cul-
tured in vitro, it is considered a model organism for the
study of Cordyceps species in culture (Shrestha et al.
2004a; Gao 2008; Zhong et al. 2009; Xiong et al. 2010).
Unlike O. sinensis, C. militaris has worldwide distribu-
tion; however, its population density is quite insignificant.
Moreover, the stromata produced by C. militaris in the wild
are tiny in size. Thus, collection of specimens in the wild
cannot satisfy the human demand for this fungus, and there
has been substantial interest in the large-scale cultivation of
its mycelia and stromata for medicinal purposes (Dai et al.
2007; Gu et al. 2007). Earlier, the researchers have tried to
produce stromata of C. militaris on insects (de Bary 1867,
1887; Shanor 1936; Müller-Kögler 1965; Leatherdale 1970)
and later on different organic substrates (Pettit 1895; Sopp
1911; Kobayasi 1941; Basith and Madelin 1968; Yue et al.
1982) in the laboratory on an experimental basis. These
Mycol Progress
early studies have greatly contributed to the success of in
vitro and large-scale cultivation of C. militaris stromata. It
remains a mystery, however, how insect pathogens such as
Cordyceps species can grow and complete their life cycles
on non-insect media in vitro. Apart from stroma cultivation,
production of C. militaris mycelia in submerged culture is
also a promising field for the production of bioactive com-
pounds (Huang et al. 2006; Yu et al. 2006; Gu et al. 2007;
Kwon et al. 2009; Xie et al. 2009a, b; Das et al. 2010).
Many reviews have been published that focus more on
the cultivation practices of C. militaris (Wu et al. 2000a; Lin
et al. 2006a; Zheng and Kang 2006; Zhong et al. 2006; Dai
et al. 2007; He et al. 2011b). We have also tried to gather up-
to-date information on its biology, genetics and genome, as
well as on the optimum cultivation conditions in order to
make the information available to a wider audience in a
single review. We have, however, not touched upon the
pharmacological and biochemical aspects of C. militaris as
the current status of the knowledge has been excellently
reviewed from various laboratories (Paterson 2008; Zhou
et al. 2009b; Das et al. 2010). This text together with that by
Zheng et al. (2011a) will give a good overview of the
situation.
Life cycle of C. militaris
Host range and species affinity
Cordyceps militaris is the best known species of the genus
Cordyceps s.l. (Kobayasi 1941). Its morphological diversity
and adaption to a wide range of host insects probably
contribute to its presence in diverse geographical regions
and ecological zones in the world (Kobayasi 1941; Mains
1958; Sung and Spatafora 2004). Its most common hosts in
nature include lepidopteran larvae (caterpillars) and pupae;
other infrequent hosts being coleopteran, hymenopteran,
and dipteran insects. Lepidopteran hosts of C. militaris
belong to 12 different families (Table 1). Hosts in other
orders are Ips sexdentatus, Lachnosterna quercina, and Ten-
ebrio molitor (Coleoptera), Cimbex similis (Hymenoptera),
and Tipula paludosa (Diptera) (Table 1). There are rare reports
of C. militaris on coleopteran cockchafer (Roumeguère 1884;
Briard 1888 in Petch 1942) but it was considered an error
(Kobayasi 1941).
In addition to the above hosts, more than 25 species
of higher heteroceriid lepidopterans and unidentified
hymenopteran species have been recently reported as
hosts of C. militaris (Kryukov et al. 2011). Host iden-
tifications in the literature are often inadequate; for
example, most papers concerning hosts of C. militaris
identify the insects simply as lepidopteran larvae or
pupae (Kryukov et al. 2011). The identification of C.
Mycol Progress

Table 1 Host species of Cordyceps militaris belonging to different orders and families of insects

Order Family Species References

Coleoptera Scarabaeidae Lachnosterna quercina Farlow and Seymour 1888

Scolytidae Ips sexdentatus Kryukov et al. 2011
Tenebrionidae Tenebrio molitor de Bary 1867
Diptera Tipulidae Tipula paludosa Müller-Kögler 1965
Hymenoptera Cimbicidae Cimbex similis Kobayasi 1941
Lepidoptera Arctiidae Euprepia caja de Bary 1867
Bombycidae Andraca bipunctata Panigrahi 1995
Bombyx caja de Cesati 1861
B. mori Gu and Liang 1987
B. pythiocampa Durieu 1859 in Kobayasi 1941
B. rubi Tulasne and Tulasne 1865
Drepanidae Cymatophora duplaris Lagerberg 1922
C. flavicornis Lagerberg 1922
Palimpsestis duplaris Ulvinen 1969
Polyploca flavicornis Ulvinen 1969
Geometridae Biston falcata Ma et al. 2007
Culcula panterinaria Chen 1997
Lycia hirtaria Ulvinen 1969
Hepialidae Hepialus sp. Petch 1948
Lasiocampidae Dendrolimus latipennis Jing 1987
D. pini Sopp 1911
D. sibiricus and Macrothylacia rubi Kryukov et al. 2011
Gastropacha quercus de Bary 1867
Lymantriidae Dasychira pudibunda Ulvinen 1969
Leucoma salicis Kryukov et al. 2011
Noctuidae Apatele aceris Ulvinen 1969
Cocytodes coerulea Kobayasi 1941
Colocasia coryli Ulvinen 1969
Euxoa ochrogaster Kryukov et al. 2011
Hylophila prasiana Ulvinen 1969
Panolis flammea Siemaszko in Kobayasi 1941
Notodontidae Fentonia ocypete, Lampronadata cristata, Chen 1997
Phalera assimilis and Phalerodonta albibasis
Phalera bucephala Gray 1858
Quadricalcarifera punctatella Sato et al. 1994
Saturniidae Anisota senatoria Hitchcock 1961
Sphingidae Callambulyx tatarinovi Kobayasi 1941
Laothë populi Ulvinen 1969
Marumba sperchius Chen 1997
Mimas tiliae Ulvinen 1969
Sphinx euphorbiae de Bary 1867
S. pinastri Ulvinen 1969
Thyatiridae Tetheella fluctuosa Kryukov et al. 2011
Ochropacha duplaris
Tethea ocularis

militaris hosts, especially as reported in earlier publica- occasionally reported, e.g., on Syntypistis punctatella
tions, deserves revision in light of modern insect taxon- (Quadricalcarifera punctatella) and Dendrolimus pini
omy. Cases of C. militaris outbreaks have been (Kamata 1998, 2000; Sierpiñska 1998).

Many Cordyceps species are morphologically or other-
wise similar to C. militaris, and these include C. cardinalis
G.H. Sung & Spatafora, C. kyusyuensis A. Kawam., C.
pseudomilitaris Hywel-Jones & Sivichai, C. rosea Kobayasi
& Shimizu, C. roseostromata Kobayasi & Shimizu, C.
washingtonensis Mains, and others (Sung and Spatafora
2004; Sung et al. 2007; Wang et al. 2008). Although dis-
tinguishing C. kyusyuensis from C. militaris is difficult
based on morphology and cultural characteristics (unpub-
lished work), the larvae parasitized by C. kyusyuensis tend
to be larger and located deeper in soil than larvae parasitized
by C. militaris (Kawamura 1955; Sung 1996; see Table 1).
C. kyusyuensis, originally described from Japan on larvae of
the lepidopteran Clanis bilineata (family Sphingidae)
(Kawamura 1955), has been reported only from China
(Kobayasi 1981; Guo and Li 2000) and Korea (Sung
1996), besides Japan. According to Wang et al. (2008), C.
kyusyuensis is not a different species, rather a synonym of C.
militaris.
Mating system
Only a few cytological studies have been conducted on C.
militaris. Earlier studies showed that C. militaris has only
two chromosomes (Varitchak 1927; Jenkins 1934), while a
recent study reported seven chromosomes (Wang et al.
2010c). Despite the scarcity of cytological studies, the mat-
ing system of C. militaris has been studied in many instan-
ces at both cultural and molecular levels. Inoculations of
two single-ascospore strains together or on opposite sides on
a fruiting medium plate are the two major methods for
genetic analysis of mating system in ascomycetous fungi,
including Cordyceps spp. (Harris 2001). Gao et al. (2000b)
reported that single-ascospore strains of C. militaris were
unable to produce stromata when growing alone on media.
Later, Sung and Shrestha (2002) and Shrestha et al. (2004a)
showed that single-ascospore strains of C. militaris pro-
duced perithecial stromata when inoculated in combination
with a compatible isolate, but produced only deformed
stromata without perithecia when inoculated singly. Further,
it was shown that primordia developed in the meeting line
between two opposite mating-type strains that were inocu-
lated on opposite sides of an agar plate (Shrestha et al.
2004a). These results show that C. militaris is a bipolar
heterothallic fungus. The mating type character was found
to be stable for as many as 10 generations of subcultures of
the original strains (Shrestha et al. 2004a; Sung et al.
2006a). Each single ascospore strain was found to be her-
maphroditic but self-incompatible (Shrestha et al. 2004a). In
agreement with these studies, Liang et al. (2005) observed
that, when growing alone, most of the single-ascospore
strains could not produce well-developed stromata or pro-
duced only abnormal ones. Gao (2008), Wen et al. (2009)
Mycol Progress
and Zheng et al. (2011a) also recently confirmed bipolar
heterothallism in C. militaris. A molecular study has dem-
onstrated that C. militaris possesses opposite mating-type
idiomorphs, MAT1-1 and MAT1-2 (Yokoyama et al. 2006),
or two mating type genes, MAT-α and MAT-HMG (Wang et
al. 2010a). Zheng et al. (2011a) recently identified the
MAT1-1 mating type locus that included MAT1-1-1 and
MAT 1-1-2 genes.
Despite obvious heterothallism, homothallism has been
occasionally observed in C. militaris (Sato and Shimazu
2002a; Shrestha et al. 2004a; Liang et al. 2005; Wen et al.
2009; Zheng et al. 2011a). Homothallism in C. militaris
could be due to the presence of (1) both genotypes
(MAT1-1/2) in the chromosomes, (2) a disomic (diploid)
condition with an extra chromosome of the opposite mating
type locus, or (3) a heterokaryotic condition with more than
one nucleus with opposite mating type loci (Shrestha et al.
2004a; Wen et al. 2009). Intraspecific high genetic variation
seems common in C. militaris and may be associated with
mating-system instability (Wen et al. 2009). Recently, het-
erokaryosis and parasexuality were confirmed in single-
spore strains of C. militaris by RAPD analysis (Li et al.
2007a). Mating-type switching can also be another possible
reason for homothallism in C. militaris.
In contrast to mating-type loci in homothallism, mating-
type loci in heterothallic fungi exist in separate spores.
Mating compatibility in heterothallism can be controlled
by only one factor (bipolar) or by two factors (tetrapolar).
Basidiomycetous fungi are mostly heterothallic, while a
majority of ascomycetous fungi studied are homothallic.
Heterothallic ascomycetous fungi contain a single mating-
type locus with two alternate alleles (bipolar) (Poggeler
2001), while basidiomycetes have evolved multiple
mating-type loci (Kothe 2001). To determine mating com-
patibility in ascomycetes, a researcher must wait until fertile
reproductive structures are formed. There is no specific
indication in the somatic structure or the asexual reproduc-
tion structure in C. militaris strains that they have different
abilities of producing fruiting-bodies (Gao et al. 2000b; Wu
et al. 2000b). Liang (1990, 2001), however, distinguished
fruiting-body formation from ‘Paecilomyces-type’ but not
from ‘Acremonium-type’ strains. Li et al. (2006a) also
reported that fruiting-bodies were developed from a
‘Lecanicillium-type’ strain but not from an isolate with more
densely verticillate phialides. In another study, it was shown
that fast-growing isolates produced fewer fruiting-bodies
than slow-growing isolates (He et al. 2010).
Although C. militaris lacks distinctive phenotypic char-
acteristics both in the wild and in culture, one obvious
phenotypic characteristic in culture is the colony pigmenta-
tion that ranges from orange to yellowish-white on nutrient-
rich agar media (Shrestha et al. 2006). Either kind of pig-
mentation was observed in F1 progeny strains when two
Mycol Progress
parent strains with contrasting pigmentations (i.e, yellowish-
white vs. orange) were crossed (Shrestha et al. 2005a). The
F1 progenies either on back-crossing with the parents or
sister-crossing among themselves showed that colony pig-
mentation was independent of or distantly linked with the
mating-type locus (Shrestha et al. 2004a, 2005a).
The study of the mating system is more difficult with
clavicipitaceous fungi than with many other ascomycetous
fungi. Clavicipitaceous fungi are characterized by septate,
filamentous ascospores. From a cytological perspective,
each filamentous clavicipitaceous ascospore corresponds to
a unicellular ascospore of other ascomycetes. Ascospore
initials in C. militaris undergo asexual or somatic division
seven times and are ultimately filamentous in shape and
consist of 128 part-spores (Moore 1964; Ding et al. 1995;
Hywel-Jones 2002).
Several characteristics of C. militaris ascospores make
single-spore isolation difficult. Firstly, C. militaris asco-
spores that consist of as many as 128 part-spores, also
known as secondary spores, readily fragment into short
filaments upon discharge from mature perithecia, rendering
complete unfragmented ascospores difficult to find. Second-
ly, short filaments of the ascospores tend to overlap or attach
end-to-end when released (Shrestha et al. 2005c), which
often results in the isolation of multiple ascospores rather
than a single one. Thirdly, ascospores tend to germinate
quickly in agar media, and germinating hyphae can link
the genomes of nearby ascospores (Liang et al. 2005;
Shrestha et al. 2005c; Gao 2008). To avoid these problems,
researchers should pay attention to ascospore length during
isolation and should avoid isolating ascospores that appear
abnormally long. Our experience is to leave the ascospores
on the isolation medium for a few hours so that they swell
but do not germinate. The swelling makes it easier to ob-
serve individual ascospore filaments. Although individual
ascospore filaments can be isolated after some experience, it
is not possible to isolate all the eight ascospores from the
same ascus in a Cordyceps (s.l.) species. Isolation of indi-
vidual part-spores is the final goal that can be achieved to
establish a pure culture. Despite difficulty, it is hence urged ,
in future, to cultures from individual part-spores, probably
attainable by the dilution method, in order to overcome the
confusion of single-ascospore isolation.
Out-crossing has been successfully achieved for C. mil-
itaris in culture. For example, Sung et al. (2006b) reported
the formation of perithecial stromata from crossing between
different specimens of C. militaris. Although it has been
frequently studied, the mating system of C. militaris has
been described as unknown in fungal textbooks (e.g.,
Webster and Weber 2007).
It is clear that mating compatibility not only produces
fertile perithecia but also affects developmental and mor-
phological patterns of stromata. Stromata produced by
successful mating are regular, club-shaped, or cylindrical,
whereas those produced without mating are deformed and
abnormal in size (Shrestha et al. 2004a; Liang et al. 2005).
Usually, the cortex in the fertile clava of stromata becomes
loose and spongy at maturity whereas, in those produced
without mating, it remains hard and compact (unpublished
data). The reason might be that substantial materials and
energy are used for the development of perithecia in the case
of mating, leaving the interstitial cortex tissue weak. Al-
though the differences of biochemical composition between
perithecial and non-perithecial stromata remain to be deter-
mined, Wen et al. (2005) showed that the amounts of me-
dicinal components such as polysaccharide, adenosine,
cordycepin, and cordycepic acid differ between sclerotium
and stroma of C. militaris.
In addition to clarifying the life cycles of fungi, mating
studies are also important for strain improvement (Kothe
2001; Poggeler 2001). In addition, the mating system also
has evolutionary and taxonomic significance in Cordyceps.
Although researchers continue to debate whether homothal-
lism preceded heterothallism or vice versa (Geiser et al.
1998; Yun et al. 1999), it is generally assumed that the
mating systems might have switched from one to the other
type more than once in the course of evolution. Whether
most Cordyceps species are homothallic or heterothallic
remains unknown. A molecular study has shown that sev-
eral Cordyceps species studied contain opposite mating-type
idiomorphs, MAT1-1 and MAT1-2 (Yokoyama et al. 2006).
A recent study of the mating system in O. sinensis has
detected only the mating-type gene MAT1-2-1 of the idio-
morph MAT1-2 (Zhang et al. 2011), suggesting that O.
sinensis could be homothallic. The study of the mating
system in O. sinensis is hindered by inadequate understand-
ing of its biology, slow hyphal growth, difficulty in inducing
stromata in culture, and the scarcity of genetic and genomic
resources (Zhang et al. 2011).
In addition to increasing our understanding of fruiting-
body formation, information about mating compatibility can
also help with the identification of similar species, such as
C. militaris and C. kyusyuensis, according to a “biological
species concept”. It is important to understand the biological
species concept of C. militaris in the context of its world-
wide distribution and wide host range.
Nomenclature and status of the anamorph
Cordyceps militaris produces asexual spores (conidia) on
the tips of little-differentiated phialides and eight filamen-
tous sexual spores (ascospores) in an ascus (within a peri-
thecium) during its sexual phase (Kobayasi 1941; Feng et al.
1990; Liang 1990; Ding et al. 1995). Naming of the conidial
structure or anamorph began from the early phase of re-
search on C. militaris, but the name has frequently changed.

Gams (1971) removed C. militaris anamorph from Paecilo-
myces (now mainly Isaria) (Gams et al. 2005; Hodge et al.
2005) or Cephalosporium/Acremonium and transferred it to
Verticillium sect. Prostrata. Zare and Gams (2001) later
erected a new genus Lecanicillium to substitute for Verticil-
lium sect. Prostrata including the anamorph of C. militaris.
It is a characteristic of this fungus that conidia can arise in
the same strain either in chains or in heads which may be
partially modified by different media. The phialides can either
arise singly or in whorls again possibly dependent on the
medium, but no mutation is required for such a change.
Finally, starting on 1 January 2013, the confusion over
multiple naming of holomorphic fungi will be resolved
through the recent revision of Article 59 of the International
Code of Botanical Nomenclature (ICBN) [recently changed
to the International Code of Nomenclature for algae, fungi,
and plants (ICN)] that will validate only one name in a clade
based on priority (Hawksworth 2011; Miller et al. 2011;
Norvell 2011). The revision will help establish the concept
of one fungus 0 one name (Hawksworth 2011). Different
anamorph names associated with C. militaris can be stated
as morphological stages, e.g., acremonium, lecanicillium,
paecilomyces, verticillium, etc., with a lower case initial
letter and normal non-italic type, following Cannon and
Kirk (2000) and Hawksworth (2011).
Synonymous terms for stromata
A stroma is a compact, somatic structure or a cushion-like
matrix of mainly ascomycetous fungi on which or in which
spores or fruiting bodies are usually formed (Alexopoulos et
al. 1996; Kirk et al. 2001). The stroma is generally distin-
guished from the synnema, which carries conidia or asexual
spores at the tips of conidiogenous cells (phialides) or on
indistinct hyphae. Stromata of C. militaris are clavate, club-
shaped, or cylindrical with a lower sterile stipe and upper
fertile clava, also known as the apical part or head, and are
orange or yellow in color. In the wild, stromata are solitary
to few or in some cases gregarious. The literature on the
cultivation of Cordyceps spp., however, uses a variety of
terms interchangeably with stroma, and these include
fruiting-body (or fruitbody, fruit body, fruit-body), artificial
stroma, perithecial stroma, sporophore, sexual sporophyte,
sporocarp, ascostroma, and others. Among them, ‘fruit-
body’, ‘fruit body’, and ‘fruiting-body’ are most commonly
used and may be preferred to stroma for communication
with farmers, businessmen, and consumers. Stroma and
stromata, however, are the preferred terms for scientific
reports. Different words have also been used to describe
stromata produced in culture, and these include normal
versus abnormal, complete versus incomplete, regular ver-
sus irregular, stable versus unstable, mature versus imma-
ture, perfect versus imperfect, and uniform versus deformed.
Mycol Progress
For morphogenetic descriptions, the terms ‘perithecial’ and
‘non-perithecial’ or ‘aperithecial’ stromata should be used.
Cytology, genetics, and genomics
Only a few studies have been conducted on the cytology and
genetics of C. militaris. Moore (1964) showed that the
manner of somatic nuclear division is similar to that of cell
division. Electrophoresis karyotype analysis showed that its
chromosomal number is seven, and that chromosome size
ranges between 2.0 and 5.7 Mb (Wang et al. 2010c). How-
ever, recent whole genome sequencing of C. militaris has
shown that its total genome is exactly 32.2 Mb (Zheng et al.
2011a), which is smaller than that of two other entomopa-
thogenic fungi, Metarhizium anisopliae (39.04 Mb) and M.
acridum (38.05 Mb) (Gao et al. 2011). Conditions affecting
the formation and regeneration of protoplasts from C. mili-
taris have been extensively studied (Ma et al. 2008; Liu et
al. 2009; Zhou and Luo 2009; Li et al. 2011), and mutants
with superior traits, e.g., high production of cordycepin,
polysaccharide, and fruiting-bodies, have been obtained by
irradiation induction (Che et al. 2004; Zhou and Bian 2007;
Zhou et al. 2009a; Li et al. 2011).
The study of fungal genetics requires an efficient trans-
formation system. Zheng et al. (2011b) developed and opti-
mized Agrobacterium tumefaciens-mediated transformation
for C. militaris, which can facilitate the identification of
functional genes. Expressed sequence tag (EST) analysis
revealed different transcriptional patterns for C. militaris in
mycelia growing in liquid culture or on solid rice medium,
and in fruiting-bodies produced on rice medium or on silk-
worm pupae (Xiong et al. 2010). Further analysis showed
that genes involved in cell metabolism, energy metabolism,
and stress responses were upregulated during asexual devel-
opment, and that genes associated with cell wall structures
were upregulated during sexual development (Xiong et al.
2010). Many studies have described how conditions affect
fruiting-body formation and metabolite production (e.g.,
cordycepin, polysaccharide) (Cui and Zhang 2011; He et
al. 2011b), but the genetic basis for these processes is still
unclear.
Fortunately, the genome of C. militaris has recently been
sequenced (Zheng et al. 2011a). A total of 9,684 protein-
coding genes have been predicted, and 13.7 % of these
genes are species-specific, which is significantly higher than
the percentage for M. anisopliae (4.8 %) or M. acridum
(3.5 %). About 16 % of the C. militaris genes (1,547) are
related to pathogen–host interactions. No orthologs of
known human mycotoxins have been detected (Zheng et
al. 2011a), which is consistent with its safe usage as a
medicine. More than 63 % of the total 9,684 genes were
expressed during both mycelial growth and fruiting-body
Mycol Progress
formation. The Zn2Cys6-type transcription factors and
MAPK pathway were induced during fruiting, but not the
PKA pathway, which differs from the induction of these
pathways in other fungi. The complete sequencing of the
C. militaris genome will facilitate functional studies of
interesting genes and elucidate the genetic background for
biosynthesis of bioactive components, insect pathogenicity,
and isolate degeneration (Zheng et al. 2011a).
Significant genetic differences between wild-type strains
and degenerate strains have been detected (Li et al. 2003,
2007a). However, unlike the significant intraspecific genetic
diversity of O. sinensis (Zhang et al. 2009), the genetic
distance among C. militaris isolates from different localities
is extremely low based on nrDNA ITS sequences (K2P
distance value ≤0.01) (Wang et al. 2008).
Some C. militaris proteins have been purified, and some
C. militaris genes have been cloned. The purified proteins
include an extracellular trypsin-type serine protease P-1-1
(Hattori et al. 2005), a cytotoxic and antifungal protease
CMP (Park et al. 2009), an antifungal peptide cordymin
(Wong et al. 2011), a haemagglutinin with anti-proliferative
activity towards hepatoma cells (Wong et al. 2009), a lectin
(CML) that exhibits haemagglutination activity in mouse and
rat erythrocytes (Jung et al. 2007), fibrinolytic enzymes (Kim
et al. 2006; Cui et al. 2008; Choi et al. 2011), and superoxide
dismutase (Wang et al. 2005a). The cloned genes include a
glyceraldehyde-3-phosphate dehydrogenase (GPD) gene
(Gong et al. 2009), superoxide dismutase genes (Park et al.
2005; Wang et al. 2005b), and the β-1,3-glucan synthase
catalytic subunit gene (Ujita et al. 2006).
Cultivation of C. militaris
Nutritional requirements for stroma growth and production
Insects
The artificial growth and stroma production of C. militaris
has been studied in the laboratory on various insect pupae
and larvae, most often on the silkworm Bombyx mori, (Gu
and Liang 1987; Liang and Gu 1987; Gu et al. 1988; Gong
et al. 1993; Zhou et al. 2000; Chen and Ichida 2002; Li
2002; Pan et al. 2002; Sato and Shimazu 2002b; Zhang et al.
2003; Liu 2004; Wen et al. 2004; Li et al. 2006a; Zheng et
al. 2008a, b; Chai et al. 2010; Hong et al. 2010; Mu et al.
2010). Other insects used for artificial stroma production are
Antherea pernyi (Gu and Liang 1987; Liang and Gu 1987;
Gu et al. 1988; Yuan 1988, 1989; Feng et al. 1990; Wang et
al. 2002), Mamestra brassicae (Harada et al. 1995; Sato and
Shimazu 2002b), Tenebrio molitor (Sato and Shimazu
2002b; Lin et al. 2005), Ostrinia nubilalis (Liang and Gu
1987), Heliothis virescens, H. zea and Spodoptera
frugiperda (Sánchez-Peña 1990), Andraca bipunctata
(Panigrahi 1995), Philosamia cynthia (Jiang and Xun
1996), Spodoptera litura (Sato and Shimazu 2002b),
and Clanis bilineata (Song 2009). Chen and Ichida
(2002) documented a higher rate of infection and stroma
formation in silkworm pupae than in silkworm larvae.
Among three varieties of silkworm (Daeseungjam, Bae-
gokjam, and Keumokjam), the Daeseungjam variety was
found to be the most suitable for stroma formation of C.
militaris (Hong et al. 2010).
Natural organic substrates
Because insects are expensive and not always available (Lin
et al. 2006c), and because insects can be difficult to handle
and thus prone to microbial contamination, alternative or-
ganic substrates have been tested for commercial production
of C. militaris stromata. Fortunately, cereals with the addi-
tion of some organic substances have proven to be good
substitutes for insects. Kobayasi (1941) documented stroma
production of C. militaris on rice substrate. Since then, rice
has been used as the principal ingredient for growing stro-
mata of C. militaris (Basith and Madelin 1968; Chen and
Wu 1990; Liang 1990; Ma and Chen 1991; Sung et al. 1993,
1999; Pen 1995; Sung 1996; Wu et al. 1996; Zhang and Liu
1997; Choi et al. 1999; Li 2002; Zhang 2003; Li et al.
2006d; Lin et al. 2006b; Wen et al. 2008b; Chen et al.
2011b).
The porosity of the fruiting medium affects mycelial
growth and fruiting-body yield. Porosity increases with
grain size and declines with a higher ratio of water to grain
during rice medium preparation. In the absence of interstic-
es, mycelia mostly grow only on the surface of the medium
and thereby cannot absorb enough nutrition from the sub-
strate. Interstices permit hyphae to grow inside the medium
and to obtain sufficient nutrition from the substrate
(Kobayasi 1941). A ratio of rice to water from 1:1 to
1:1.35 or slightly higher has been reported to be optimal
for stroma production (Sung et al. 1999, 2002; Lin et al.
2006b; Zheng et al. 2008c; Yue 2010), but the optimal ratio
may depend upon the rice cultivar and its glutinous quality.
Husked rice (popularly known as brown or unpolished rice)
is usually used for cultivation of C. militaris. Maximum
fruiting-body yield has been obtained with whole rice grain
(Wen et al. 2008b).
Other organic materials used for the production of C.
militaris stromata include bean powder, corn grain, corn
cobs, cotton seed coats, jowar, millet, sorghum, fragments
of sunflower floral disks, and wheat grain (Chen and Wu
1990; Zhang and Liu 1997; Li 2002; Li et al. 2004a; Zhao et
al. 2006a; Gao and Wang 2008; Wei and Huang 2009). Rice
mixed with silkworm pupae has proven to be superior to
other substrates and is now routinely used (Ren 1998; Chen

et al. 2002; Shrestha et al. 2004a, b, 2005a, b; Sung et al.
2002, 2006a, b; Zhao et al. 2006a; Jin et al. 2009). C.
militaris strains require a relatively low level of nitrogen,
and excessive nitrogen might suppress differentiation of the
fruiting-body (Gao et al. 2000a). This probably explains
why yields have been observed to be less on insects than
on cereals in the culture. Xie et al. (2009a, b) have also
shown that brown rice, malt, and soybean are better sources
of nutrition for C. militaris than chemically defined media.
Agar media are usually not suitable for stroma production
(Basith and Madelin 1968; Yahagi et al. 2004).
Hormones and mineral components of the media
Plant hormones such as 2, 4-D, citric acid triamine, colchi-
cines, and others may enhance stroma production by C.
militaris (Li et al. 2004a; Wang et al. 2010b; Xiao et al.
2010). Similarly, mineral salts such as K+, Mg2+, and Ca2+
at a concentration of 0.1 g/l may increase fruiting yield (Li et
al. 2004a). Some elements may enhance the production of
bioactive compounds of C. militaris in culture (Dong et al.
2012).
Duration of stroma production and stromata yield
Commercial production must take into account the duration
of stroma production. Stromata are usually produced over a
period of 35–70 days (Zhang and Liu 1997; Sung et al.
1999; Yue 2010; Du et al. 2010). Zhang and Liu (1997)
reported the production period of 35–45 days on rice but
40–70 days on other substrates such as maize, millet, and
rice-tussah. Culture duration, however, depends upon the
shape and volume of the culture container and the amount
of medium. Stroma production has been quantified in some
studies. Wu et al. (1996) obtained 25 g of fresh fruit bodies
of C. militaris from 50 g of rice medium, while Zhang and
Liu (1997) reported a biological transformation rate of 61 %
on rice, 58–59 % on millet and rice-tussah media, and 42 %
on maize. Production of 18.0 g of stromata (fresh wt.) has
been recently obtained from 20 g of rice (Lin et al. 2006a).
Nearly 9 g of dry stromata (equivalent to about 68 g of fresh
wt.) was produced from 60 g of brown rice supplemented
with 10 g of silkworm pupae (Sung et al. 2006b).
Effect of environment
Whereas hormones govern morphogenetic and develop-
mental changes in plant tissue culture, it is environmen-
tal factors that govern the change from the somatic
phase to the reproductive phase in fungi. The optimal
environmental factors for growth of C. militaris stroma-
ta are briefly discussed below.
Mycol Progress
Temperature
Temperature greatly affects C. militaris stroma production.
High temperatures (about 25 °C) lead to maximal mycelial
growth, but lower temperatures induce and sustain stroma
production, viz. temperatures within the range of 18–22 °C
are reported as optimal (Sung et al. 1999, 2002; Gao et al.
2000a; Zhao et al. 2006a; Du et al. 2010; Sato and Shimazu
2002b), although a lower range of 14–17 °C (Du et al. 2007)
or a higher temperature of 25 °C (Li et al. 2004a; Yue 2010)
have also been reported to be appropriate for stroma pro-
duction. The maturation period of stromata is shorter at
25 °C than at 20 °C (Sato and Shimazu 2002b).
Light
Light is the most important environmental factor affecting
C. militaris stroma production and no stromata are produced
in darkness (Sato and Shimazu 2002b). Gao et al. (2000a)
obtained stromata under light intensities as high as 4,500 lx,
but Sato and Shimazu (2002b) indicated that the upper limit
was 1,400 lx; in general, 500–1,000 lx is considered optimal
(Sung et al. 1999; Gao et al. 2000a; Sato and Shimazu
2002b; Li et al. 2004a; Zhao et al. 2006a; Du et al. 2010),
with a 12-h light/dark cycle of 500–1,000 lx (Sung et al.
2002; Chen et al. 2011b). It has been found that 18 h of light
at 200 lx was optimal for pigmentation and primordium
initiation, whereas 10 h was suitable for fruiting-body
growth and yield (Mu et al. 2010). Longer and wider
fruiting-bodies were produced with light intensities of
500–1,000 lx than with an intensity of 100 lx (Hong et al.
2010).
Air exchange and humidity
High air exchange in the culture container favors mycelial
growth, primordium formation, and biomass yield (Zhang et
al. 2010a). Among materials tested for covering culture
bottles containing C. militaris, a hydrophobic fluoropore
membrane was the best (Zhang et al. 2010a). A high hu-
midity of 70–90 %, which probably matches the humidity
experienced by the fungus in nature, favors stroma produc-
tion. Low humidity causes the medium to dry too quickly.
Especially in dry, indoor environments, a humidifier will be
required to maintain sufficient humidity.
Inoculum preparation
For production of C. militaris stromata, media can be effi-
ciently inoculated with a liquid culture of the fungus (Sung
1996). The inoculum in liquid culture consists of conidia,
hyphal fragments, and hyphal pellets affected by nutrients,
culture duration, mode of culture, etc. The liquid culture
Mycol Progress
medium usually contains simple sources of carbon (usually
in the form of carbohydrates) and nitrogen (inorganic as
well as organic) along with mineral salts. The optimum
temperature and pH for liquid culture of C. militaris are
20–25 °C and 6.0–8.0, respectively. While C. militaris my-
celia produce yellowish-white to orange pigments on solid
media in light (Shrestha et al. 2006), liquid cultures of the
fungus usually remain colorless. Pellets in liquid culture
increase in size as the culture ages (Han et al. 2009). Pellets
are not a suitable form for inoculation purposes for three
reasons. First, and most importantly, pellets do not grow as
fast as conidial or hyphal suspensions on the fruiting medi-
um. Second, pellets frequently fail to induce stroma primor-
dia. Third, inoculum consisting of pellets is difficult to
quantify. For large-scale preparation of liquid inoculum,
shaking and aeration of the liquid are required for the
homogeneous growth of mycelium. The quantity of inocu-
lum added to the fruiting medium depends on the volumes
of the culture container and the fruiting medium (Sung et al.
2002; Lin et al. 2006b).
Industrial and commercial development
Although more than 400 Cordyceps s.l. species have been
described, only about 36 species have been artificially cul-
tivated for the production of fruiting-bodies (Wang 1995;
Sung 1996; Li et al. 2006b). Of those species that have been
artificially cultivated, only C. militaris has been commer-
cially cultivated; commercial development has focused on
C. militaris because of its excellent pharmaceutical effect
and short production period (Li et al. 2006b).
Cordyceps militaris has been cultivated in liquid media
for harvesting mycelia and on solid media for induction of
fruiting-bodies. While conditions for submerged cultivation
of C. militaris inoculum have been optimized (Kim et al.
2003; Liu et al. 2008), large-scale production of C. militaris
fruiting-bodies currently uses only solid media consisting of
artificial substrates or insects (e.g., the silkworm B. mori).
Because cultivation on insects is costly, fruiting-bodies of C.
militaris are mainly cultivated on artificial media in which
rice is the main component. Substrates used for industrial
cultivation of C. militaris in China have recently been
reviewed (Wang et al. 2009b). They include media for stock
culture, pre-culture spawn, and spawn. For each purpose,
the ingredients differ depending on the company (Wang et
al. 2009b). Although the cost for cultivation of several
insects has recently decreased greatly, fruiting-bodies culti-
vated on insects (1,000 RMB/kg) are twice as expensive to
produce as biomass cultivated in artificial media (500
RMB/kg) (Li et al. 2007b). Compared with the extremely
high price of O. sinensis, however, the price of C. militaris
is affordable.
In the cultivation of C. militaris fruiting-bodies, four
pivotal growth periods are usually identified: mycelial cul-
ture, pigment induction, stromata stimulation, and fruiting-
body production (Lu et al. 2005). Successful cultivation
requires proper control of temperature, humidity, and light
(Ren et al. 2009).
Cordyceps militaris cultures have two main uses. First,
the fruiting-bodies can be directly consumed as food. C.
militaris can be used in stewed chicken, stewed duck, soup,
hot pot, tea, and so on. Use of C. militaris in soup is very
popular in Southeast Asia, especially in Guangdong, Hong
Kong, and Taiwan, China. This use has been shown to be
safe if consumption is less than 2.5 g/kg of body weight
(Che 2003). Second, C. militaris fruiting-bodies and myce-
lia can be used as health products and drugs. In China, many
health products contain C. militaris, and these include oral
liquids, capsules, wines, vinegars, teas, yogurt, and soy
sauce (Wang and Yang 2006). Cultures of C. militaris are
also used to produce drugs for maintenance of kidney and
lung function, anti-aging, regulation of sleep, and chronic
bronchitis (Dai et al. 2007). Currently, more than 30 kinds of
C. militaris health products and drugs are available on the
market (Huang et al. 2010).
Problems and prospects
Degeneration of isolates
Degeneration of isolates is the main problem in C. militaris
cultivation. Degeneration is manifested as reduced growth
rate, mycelial density, pigmentation, and fruiting-body
yield, and also as changes in fruiting-body shape and size.
Degeneration can also be evident in the reduced production
of desired compounds from fermented cultures. Degenera-
tion in stroma production is related to the kind of material
that was used for isolation. If the isolate was derived from
multiple ascospores (multi-spores) or tissue, decline in stro-
ma production occurs soon after one or two subcultures
(Shrestha et al. 2004b). Degeneration can be delayed if
cultures are obtained from single ascospores (Shrestha et
al. 2004a; Sung et al. 2006a).
Compatible pairs of single-ascospore or single-conidium
strains should be used to study the effects of biotic and
abiotic factors on stroma production. With isolates derived
from multiple ascospores, multiple conidia, or tissue, stroma
production will vary even when culture conditions are con-
stant (Shrestha et al. 2002, 2004b; Liu et al. 2006). Unfor-
tunately, most publications concerning C. militaris culture
do not indicate how the isolate was obtained.
Degeneration in colony pigmentation has been observed
in C. militaris strains after several subcultures (Sung et al.
2006a). Lin et al. (2010) observed reduced dehydrogenase

activity and pigment in degenerated strains but no change in
mating-type or evidence of dsRNA infection. In addition,
the formation of heritable white synnemata has been
reported from C. militaris isolates in culture (Sato et al.
1997; Wang et al. 2009a). Formation of white synnemata
in C. militaris isolates may provide an opportunity for new
strain development as is the case for other cultivated mush-
rooms such as Flammulina velutipes. A few studies have
shown that preservation of C. militaris isolates at 4–10 °C
helps to maintain the ability of fruiting-body production for
up to 6 months (Sung et al. 2006a; Geng et al. 2009). An
increased mutation frequency at the DNA level has been
associated with degeneration of C. militaris isolates (Li et al.
2003).
The genes involved in C. militaris degeneration have not
been identified. In C. militaris, identification of genes and
their function is mostly lacking (Zheng et al. 2011b). So far
the glyceraldehyde-3-phosphate dehydrogenase gene, the β-
1,3-glucan synthase catalytic subunit gene, and the super-
oxide dismutase Cu, Zn-SOD gene are known to be in-
volved in phosphorylation, cell wall constitution, and
defense against oxidative damage, respectively (Wang et
al. 2005b; Ujita et al. 2006; Gong et al. 2009). Recent
research has demonstrated that Agrobacterium tumefa-
ciens-mediated transformation is useful to elucidate the
function of genes in C. militaris (Zheng et al. 2011b); this
method should also help in determining which genes are
responsible for degeneration of isolates.
Strain development and large-scale cultivation of C.
militaris
Cordyceps militaris cultivars with desirable properties such
as high production of stromata and high cordycepin content
have recently been developed (Sun et al. 2009; Du et al.
2010). Du et al. (2010) reported a new cultivar of C. mili-
taris with high cordycepin yield (24.98 mg/g of fruiting-
body dry wt.). Che et al. (2004) obtained a higher yielding
and more stable strain of C. militaris by UV mutagenesis.
Recently, regeneration of C. militaris from its protoplasts
has been studied (Zhou and Bian 2007; Liu et al. 2009;
Zhou and Luo 2009). C. militaris isolates from different
regions contain different concentrations of active com-
pounds (Wen et al. 2008a). Thus, superior isolates can be
selected for propagation.
Cordyceps militaris is usually cultured in small contain-
ers (0.5–1.0 l) because the fungus requires high humidity
and moisture, and because small containers such as bottles
and trays seem to provide an excellent environment for the
growth of primordia and stromata. Reusable light plastic
containers can reduce the cost of culture. Substantially in-
creasing production and reducing costs will require the
development of bed cultivation or other cultivation methods.
Mycol Progress
Conclusion
Xiong et al. (2010) considered insect-grown C. militaris to
be far superior to cereal-grown C. militaris. Huang et al.
(2009), however, reported that fruiting-bodies had higher
contents of cordycepin and adenosine when cultivated on
rice medium than on silkworm chrysalid or wheat medium.
The latter authors also observed that cordycepin and aden-
osine contents were higher in cultivated fruiting-bodies of
C. militaris than in natural O. sinensis fruiting-bodies, and
that the contents in cultured mycelium of C. militaris were
similar to those in O. sinensis fruiting-bodies. Thus, they
proposed that rice-grown C. militaris was the best substitute
for O. sinensis (Huang et al. 2009). Brown rice along with
malt and soybean has been shown to provide sufficient
nutrition for C. militaris (Xie et al. 2009a, b). Wu et al.
(2012) compared polysaccharides of C. militaris fruiting-
bodies cultivated on rice medium and silkworm pupae, and
found that the components and structures of polysaccharides
differ in fruiting-bodies cultivated in those two types of
media. Similarly, metabolomic studies at regular durations
is also useful in order to determine the optimum cultivation
period of C. militaris (Choi et al. 2010). There are diverse
views, however, regarding the medicinal properties of wild
and cultivated C. militaris as well as those grown on various
organic substances such as cereals and insects (Li et al.
2004b; Huang et al. 2009; Xiong et al. 2010). Future studies
on C. militaris will likely identify new organic substrates
that are economical and that support high stroma production
and a high concentration of bioactive compounds. The se-
quencing of its complete genome (Zheng et al. 2011a) will
facilitate research on the molecular basis of the biology and
medicinal qualities of C. militaris.
The phylogenetic classification has divided the genus
Cordyceps into five genera in three families that fit the
present concept of the fungal tree of life (Sung et al. 2007;
Kepler et al. 2012b). Phylogenetic classification is, howev-
er, in a state of flux and has resulted in the frequent transfer
of Cordyceps species from one genus to another in different
families (Sung et al. 2007; Kepler et al. 2012a). Further-
more, Cordyceps spp. growing on cicadas have been phylo-
genetically placed under three different genera (Sung et al.
2007; Sato et al. 2012). Outside the mycological world, the
transfer of C. sinensis from Cordyceps to the newly estab-
lished genus Ophiocordyceps has caused some confusion
among researchers in the biochemical and pharmacological
communities, because of the belief that O. sinensis (syn. C.
sinensis) would be different from Cordyceps spp. at the
generic level, though they have adapted to the same level
of insect-dependent nutrition. All Cordyceps spp. have sim-
ilar life cycles and have developed mechanisms to invade
insects and grow on them; the major difference being the
locality where they grow and the host insect they infect.
Mycol Progress
Given the diversity of host insects and geographical regions,
Cordyceps spp. differ widely in shape, size, color, texture,
position of fertile parts, and other micromorphological char-
acters. Among all Cordyceps spp. s.l., O. sinensis has a
distinct ecological niche in that it grows at high altitudes
in the alpine region. Cordyceps s.l. is a broad genus that
encompasses clavicipitaceous fungi that grow on insects as
well as on fungi. Cordyceps species that grow on fungi have
been phylogenetically placed in the new genera Elaphocor-
dyceps and Tyrannocordyceps (Sung et al. 2007; Kepler et
al. 2012b). The rest of the Cordyceps species grow on
insects and spiders. Cordyceps species remain dormant in
soil until they get in contact with a host. They all grow
inside the host body and form endosclerotium before emerg-
ing from the host body in the summer. Besides their mor-
phological diversity, they also differ widely in cultivability.
C. militaris is the most successfully cultivated species
(Kobayasi 1941; Sung 1996). As stated above, we believe
that C. militaris is worth promoting as a medicinal fungus,
being the best described and cultivable Cordyceps species.
Acknowledgments The first author acknowledges the support of the
Green Energy Mission/Nepal, Kathmandu, Nepal and a grant from the
Next-Generation BioGreen 21 Program (PJ008154 and PJ008321),
Rural Development Administration, Republic of Korea, during the
preparation of the manuscript. The second author acknowledges the
support of the National Natural Science Foundation of China (No.
30870450). The authors also sincerely thank the unknown reviewer
for the helpful comments and suggestions.
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