TROPICAL ORCHID MYCORRHIZAE: POTENTIAL APPLICATIONS IN ORCHID CONSERVATION, COMMERCIALIZATION, AND BEYOND

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LANKESTERIANA 13(1–2): 57—63. 2013.
TROPICAL ORCHID MYCORRHIZAE: POTENTIAL APPLICATIONS
IN ORCHID CONSERVATION, COMMERCIALIZATION, AND BEYOND
Joel tuPAc otero1, 2*, AnA teresA MosquerA3 & nicolA s. flAnAgAn3
1Departamento de Ciencias Biológicas, Universidad Nacional de Colombia sede Palmira, Cra 32 12-00,
Palmira, Valle del Cauca, Caolombia
2Instituto de Estudios Ambientales IDEA, Universidad Nacional de Colombia sede Palmira, Colombia
3Programa de Biología, Ponticia Universidad Javeriana, Cali, Colombia
*Author for correspondence: jtoteroo@unal.edu.co
ABstrAct. Orchid mycorrhizae are unique interactions in the plant kingdom involving all the orchids and a
variety of fungi including Rhizoctonia. Orchids are one of the most charismatic plant families and include at
least 20,000 species widely appreciated by specialist growers and scientists. They also include Vanilla, source
of one of the most traded spices worldwide. Most mycorrhizal fungi belong to a group of basidiomycetes widely
known for their pathogenic interaction with many crop plants including potatoes, rice, and beans. The main
application of orchid mycorrhizal fungi is in the propagation of endangered and commercial orchid species, but
we have recently documented an alternative use. The fungal symbionts of orchids have the ability to induce
resistance to Rhizoctonia in rice plants, which opens new possibilities of biological control agents never
previously imagined.
Key Words: mycorrhiza, fungi, Orchidaceae, Rhizoctonia, biological control
Comprising 10% of angiosperm species,
Orchidaceae contains an estimated 25,000 different
species (Dressler 1993). Within Ecuador, 3784 orchid
species are reported, with 3264 species in neighboring
Colombia (Dodson 2003). The high diversity of
Orchidaceae in these countries reects the presence of
two biodiversity hotspots in the northwestern region
of South America: the tropical Andean region and
the Chocó-Darien biodiversity hotspot (Myers et al.
2002).
Orchids have attracted considerable scientic
interest since Darwin`s seminal publication (1877).
Unfortunately, the fascinating and diverse oral
morphology of orchids has also led in recent
decades to serious threats to the survival of many
orchid species. Particularly in regions with weak
law-enforcement, considerable pressure is exerted
on natural orchid populations from unsustainable
extraction of specimens from their natural populations
for commercialization. The value of ornamental trade
in orchids has been estimated at US $2 billion annually
(Harron et al. 2007). The ornamental orchid market
in Taiwan alone was reported to be valued at US
$114 million in 2012. Although the Taiwan industry
is largely based on Phalaenopsis species, breeders in
that and other countries grow and export species native
to the Neotropics. Orchid commercialization is also
a growing industry in Neotropical countries. While
many, if not most, orchid commercialization ventures
are based on the sustainable propagation of plants ex
situ, the novelty value of rare endemic species (which
are often difcult or impossible to cultivate) means
these species are subjected to continuing collection
pressures. Additionally, currently non-commercialized
species may provide valuable genetic resources in the
development of novel commercial hybrids. It is not
without reason that the whole of Orchidaceae with
the exception of articially propagated hybrids in
the genera Cymbidium, Dendrobium, Phalaenopsis,
and Vanda has been placed on Appendix II of the
Convention on International Trade in Endangered
Species of Wild Fauna & Flora (CITES), which lists
species that are not necessarily now threatened with
extinction but that may become so unless trade is
closely controlled.
Compounding the negative impact of wild
specimen collection on orchid populations is the fact
that many orchid species tend to be rare in the wild,
often with small, hyper-dispersed populations. This is
particularly true for epiphytic species, which comprise

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58 LANKESTERIANA
an estimated 70% of all orchid species (Zotz, in press),
with the great majority of these species occurring in
tropical regions. In addition to extraction pressures,
many orchid populations also face challenges to
their survival from habitat destruction and ecosystem
degradation, effects that are likely to be exacerbated
as a consequence of their specialized interactions
with pollinators, phorophytes, and mycorrhizal fungi.
Furthermore, a recent study indicates that other, as yet
unidentied factors related to microhabitat conditions
may be limiting orchid distribution and abundances
(McCormick et al. 2012). There is clearly much we
still have to learn regarding orchid biology in natural
habitats, in particular for tropical species, in order to
understand the evolutionary processes underlying
the high species diversity in this plant family and
also identify factors that determine distributions and
abundances of orchids in the wild so as to develop
effective conservation strategies for them.
The combination of commercial interest in tropical
orchids and the rapid rate of ecosystem degradation
in tropical regions has led to dramatic declines in
many orchid populations (Seaton 2007). Among many
conservation measures that need to be implemented,
key requisites for ensuring orchid population survival
are the maintenance of the levels of recruitment of
individuals into established populations as well as the
restoration of orchid populations in suitable sites.
Orchid research has principally focused on
taxonomy and the role played by pollination biology
in diversication (Micheneau et al. 2009; Schiestl
& Schluter 2009). However, other aspects of their
biology, in particular their mycorrhizal interactions,
are increasingly gaining prominence in the scientic
literature (e.g. Rasmussen 1995; Dearnaley 2007;
Kottke & Suarez 2009; Hossain et al. 2013). In addition
to the specialized relationships orchids often have with
their pollinators, accumulating studies have revealed
that some epiphytic orchids may also have strong
preferences for their host tree (phorophyte) as well
as their associated mycorrhizal fungi. Clear examples
of orchid distributions being limited with respect
to phorophyte have been reported (e.g. Gowland
et al. 2007, 2013; Crain 2012). Varying degrees of
mycorrhizal specicity have been observed throughout
Orchidaceae (Kottke et al. 2009; Yuang et al. 2010;
Valadares et al. 2012), in particular in achlorophyllous
species (Taylor & Bruns 1997; Selosse & Roy 2009).
In tropical taxa, studies of epiphytic orchids in
Oncidiinae have also revealed moderate to high levels
of preference for specic clades of Ceratobasidium
spp. (Otero et al. 2002, 2004, 2007; Valadares et al.
2012). More recently, Martos et al. (2012) showed a
general pattern for different fungal preferences among
orchids with terrestrial and epiphytic habits on the
island of Réunion.
Although historically the greater focus has been
placed on orchid pollination biology, it is likely
that seedling establishment, rather than seed set, is
the limiting step in orchid life cycles (Calvo 1993;
Otero & Flanagan 2006; Tremblay & Otero 2009).
One pollination event will yield millions of seeds
that are generally wind-dispersed. Seed germination
and seedling establishment, in contrast, may be
more challenging for the orchid plant. Lacking
endosperm, orchid seeds have minimal energy
resources for germination and rely instead upon
mycorrhizal symbioses to provide carbon and nutrients
(Bidartondo et al. 2004; Selosse & Roy 2009). Thus
the availability of a suitable mycorrhizal fungus
is crucial to orchid establishment in the wild, and
studies of orchid mycorrhizae across the family have
yielded a complicated panorama of orchid mycorrhizal
associations with varying degrees of specicity to one
or several different fungal taxa (e.g. Otero et al. 2002;
Martos et al. 2012).
Orchid mycorrhizal interactions also occur in
adult plants. In photosynthetic plants it has been
generally assumed that the provision of carbon to the
plant from the fungus is not essential, yet it seems
that mixotrophic nutritional strategies, in which the
plant receives carbon from both photosynthesis and
mycorrhizal interaction, are common (Selosse & Roy
2009; Roy et al. 2013). In this case it may be that
different parts of the plant receive carbon provision
from both sources. There is some indication that those
fungi involved in seed germination are not the same as
those that associate with adult plants. In tests of seed
germination efciency of different fungal isolates from
adult plants, fungi with provenance from other species
were sometimes more efcient (Otero et al. 2004;
Porras & Bayman, 2007).
Generally, mycorrhizal fungi in orchids belong
to the Rhizoctonia-like Basidiomycetes (Bayman

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otero et al. — Tropical orchid mycorrhizae 59
& Otero 2006), but recent studies have shown
that other groups of fungi can be involved. For
example, members of the Basidiomycete ‘rust’
lineage (Atractiellomycetes, Pucciniomycotina) are
mycobionts of orchids (Kottke et al. 2009). Epipactis
microphylla (Ehrh.) Sw. (Neottieae, Orchidaceae) is
associated with Ascomycete fungal species allied to
the ectomycorrhizal Septomycetes, including trufes
(Selosse et al. 2004).
Rhizoctonia-like fungi may be characterized by
certain hyphal morphological traits, including: a
lack of conidia; the hyphal branch at a right angle;
a septum located a short distance from a constricted
ramication; and presence of monilioid cells, a special
type of cell with a rounder shape, mainly in sclerotic
structures (Roberts 1999). The young hyphae can
be multi- or binucleate and rarely produce fruiting
bodies in culture, making it difcult to distinguish
different species from anamorph (asexual) cultures.
The teleomorph (sexual stage) of Rhizoctonia can
be Ceratobasidium, Thanatephorus (multinucleate),
Tulasnella or Sebacina, which differ morphologically
(Roberts 1999). However, it has proved extremely
difcult to promote sexual stages from asexual cultures
under laboratory conditions. The Rhizoctonia-like
fungi are grouped into anastomosis groups (AG) based
on their capacity for hyphal fusion. Multinucleate
Rhizoctonia (Thanatephorus) have 13 AG (AG1-
AG13), and binucleate Rhizoctonia (Ceratobasidium,
Tulasnella, and Sebacina) have been divided in 15
groups (AG-A to AG-O) (Sneh et al. 1991). Despite
these known traits, ne-scale characterization of fungal
strains based on morphological characters has been a
major obstacle to understanding orchid mycorrhizal
interactions.
Over the last decade or so, DNA sequence data have
been successfully applied to identify different fungal
species from pure fungal cultures or even directly from
fungal tissue present in the roots of adult plants (Otero
et al. 2002, 2004, 2005, 2007, 2011; Pereira et al.
2003, 2005, 2009; Suarez et al. 2008, 2009; Valadares
2012; Mosquera et al. 2013). The most frequent
gene region sequenced for the Rhizoctonia-like
fungi identication has been the internal transcribed
spacer (ITS) of ribosomal DNA. In tropical orchids,
four separate Ceratobasidium clades were reported
from epiphytic Oncidiinae in Puerto Rico (Otero et
al. 2002, 2004, 2005) and Central America (Otero
et al. 2007). Three further related clades have been
found in the Brazilian Coppensia doniana (Bateman
ex W.H.Baxter) Campacci (Oncidiinae), now
correctly known as Gomesa doniana (Bateman ex.
W.H.Baxter) M.W.Chase & N.H.Williams (Valadares
et al. 2012). Another occurs in Notylia (Oncidiinae),
Habenaria (Orchidinae), Cranichis (Cranichidinae),
Vanilla (Vanillinae), Epidendrum xanthinum Lindl.
(Laeliinae), Trizeuxis falcata Lindl. (Oncidiinae),
Maxillaria (Maxillariinae), and Dichaea (Dichaeinae)
(Mosquera et al. 2010). These Ceratobasidium clades
associated with Neotropical orchids are not too
distantly related to the fungi that are associated with
Pterostylis (Pterostylidinae) in temperate southeastern
Australia (Otero et al. 2011). Fungi belonging to the
teleomorph Tulasnella have also been isolated from
species of Pleurothallidinae (Suarez et al. 2006) and
Laeliinae (Pereira et al. 2001, 2005, 2006; Almeida et
al. 2007), as well as members of Vanilla in Puerto Rico
(Porras & Bayman, 2007) and Colombia (Mosquera-
Espinosa et al., unpubl.).
These studies represent an important step forward
for our understanding of the ecological interactions
between tropical orchids and their mycorrhizal
fungi at both the seed germination stage and in
adults. However, the nding of differing patterns of
mycorrhizal associations within the small number
of tropical orchid species studied so far indicates the
importance of continuing exploration of the diversity
and functional preferences of these symbioses across
the large number of orchids native to northwestern
South America. Clearly, those orchid species
facing the greatest conservation threats, either from
unsustainable extraction for commercialization or from
habitat degradation should be prioritized for orchid
mycorrhizal studies. Once the orchid mycorrhizal
fungi (OMF) have been characterized, studies can
then focus on their application in orchid conservation
programs for symbiotic orchid propagation from seed
for reintroduction and commercial purposes, as well
as for other potential applications which we discuss
below.
Commercially, there are two main orchid trades –
ornamental species and the cultivation of Vanilla. In
both trades, symbiotic seed germination could provide
considerable benets. While many species may be

LANKESTERIANA 13(1–2), August 2013. © Universidad de Costa Rica, 2013.
60 LANKESTERIANA
propagated from seed asymbiotically or vegetatively
in vitro, the presence of fungal mycorrhizae is likely
to enhance orchid plant hardening and establishment
success in reintroduction programs into the wild.
During symbiotic germination trials, fungi that give
a large advantage to seedling growth over asymbiotic
procedures have been identied. However, this is not
always straightforward. As mentioned above, fungi
isolated from the adult of the species are not always
the most effective at promoting seed germination in the
same species (Otero & Bayman 2009).
In Puerto Rico, seeds of both Tolumnia variegata
(Sw.) Braem and Ionopsis utriculariodes (Sw.) Lindl.
were symbiotically germinated using fungi isolated
from adults of the same species (Otero et al. 2004,
2005). Similarly, seeds of four groups of species of
Pterostylis s.l. were symbiotically germinated with
their mycorrhizal fungi (Otero et al. 2011). The three
Ceratorhiza and uninucleate Rhizoctonia anamorphs
isolated from Coppensia doniana Bateman ex
W.H.Baxter) Campacci [= Gomesa doniana (Bateman
ex W. H.Baxter) M.W.Chase & N.H.Williams]
were also used successfully in the in vitro symbiotic
germination of the same species (Valadares et al.
2012). However, Porras & Bayman (2007) found
that seeds of Vanilla species germinated better with
a Ceratobasidium fungus isolated from Ionopsis
utricularioides than from Tulasnella isolated from
adult Vanilla plants. These ndings indicate that
considerable study may be needed in order to identify
the most appropriate fungal partners for both orchid
conservation and commercialization.
Additionally, biotechnological techniques need
to be enhanced to improve efciency of symbiotic
germination and reduce possible contamination with
other microorganisms. Bayman (2012) developed
a formula for in-situ symbiotic propagation using
calcium alginate and combining orchid seeds and
mycorrhizal fungi for Epidendrum ibaguense Kunth to
facilitate control of the fungal presence under natural
conditions.
Vegetative propagation is almost universally used
in Vanilla cultivation. However, Vanilla crops face
serious threats from low genetic diversity, leading to
increased sensitivity to crop pathogens; propagation
from seed would help to promote genetic diversity.
At the Universidad Nacional de Colombia, Palmira
campus, Jazmin Alomia is inducing symbiotic
germination of Vanilla calyculata Schltr. with a
Rhizoctonia-like fungus (Alomía et al., unpubl.).
In addition to their functional role as orchid
mycorrhizae, some Rhizoctonia-like fungi are also
recognized as plant pathogenic fungi (Sneh et al.
1991; Roberts 1999). Rhizoctonia solani (teleomorph:
Thanatephorus cucumeris) is pathogenic in many
crop species with a worldwide distribution. It causes
damping off in beetroot, potato, beans, soya, and
cereals including rice and corn (Sneh et al. 1991)
among others, including some of the most important
crops in tropical regions.
The control of pathogenic Rhizoctonia-like fungi
generally involves chemical and biological control as
well as cultural practices. In recent years the use of
biocontrol against pathogenic Rhizoctonia fungi of
human food products has become more widespread as
a means of limiting the use of synthetic agrochemicals
that may be detrimental to both consumers and the
environment. Ideally, a fungal biocontrol agent should
specically target the pathogenic fungus and not those
that may be benecial, such as mycorrhizal and pest
pathogens. There are many examples of potential
biological control of pathogenic Rhizoctonia using
mycopathogenic fungi such as Trichoderma sp. and
Chaetomium sp. (Gao et al. 2005). A hypovirulent
Rhizoctonia-like fungus has been used to induce
systemic resistance against pathogenic Rhizoctonia
(Gressel 2001).
In our studies in Colombia, Rhizoctonia-like
fungi were isolated from a number of different orchid
species and evaluated for biocontrol potential against
the pathogenic Rhizoctonia solani in rice (Mosquera
et al. 2010). The orchid mycorrhizal fungi OMF were
identied using ITS sequences and found to form four
discrete groups. The rst included fungi from tropical
epiphytic orchids; the second included plant pathogenic
Rhizoctonia species (Thanatephorus spp.) used as a
positive control; the third included mycorrhizal fungi
from terrestrial orchids; and the nal group included
mycorrhizal fungi from Vanilla species (Mosquera et
al. 2013).
To evaluate whether the OMF were pathogenic
on rice, isolates were inoculated on healthy plants
in controlled glasshouse conditions designed to
favor the pathogenicity of Rhizoctonia solani in rice

LANKESTERIANA 13(1–2), August 2013. © Universidad de Costa Rica, 2013.
otero et al. — Tropical orchid mycorrhizae 61
(high humidity and temperature). The OMF induced
some symptoms of pathogenicity in rice, but the
severity was signicantly lower that those induced
by the positive Rhizoctonia solani control isolated
from local rice plantations (Mosquera et al. 2013).
However, when the effect on inducing resistance
against pathogenic fungi was evaluated, those healthy
rice plants inoculated with OMF two days before
inoculation with the pathogenic fungi had signicantly
fewer pathogenicity symptoms than the control plants
inoculated only with the pathogenic Rhizoctonia
solani but without OMF (Mosquera et al. 2013). These
ndings show that non-pathogenic Rhizoctonia species
that form mycorrhizae with tropical epiphytic orchids
may have potential application as biocontrol agents.
Further work is needed to understand the biological
mechanism through which this effect is mediated and
also to develop the technological application.
Conclusions
While the application of OMF in orchid
propagation for research, conservation, and
commercial purposes has been broadly recognized, the
true potential of this application is still to be realized
in Neotropical regions. Effective application of OMF
for conservation and commercial purposes requires
a considerable amount of a priori study in order to
determine which combination of plant species and
fungal partner is most effective.
Our studies in Colombia are now showing that
OMF are potentially valuable biocontrol agents for
important crop pathogens, thus providing another
tool for reducing the application of agrochemicals on
already sensitive tropical ecosystems. This deserves
greater exploration and study over the coming years.
AcKnoWledgMents. We thank DIPAL and Hermes from
Universidad Nacional de Colombia for support. ATM was
supported by a fellowship from Colciencias “Doctorados
Nacionales” 2006.
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