Mycol. Res.

100 (8): 923-935 (1996) Printed in Great Britain 923

CENTENARY REVIEW
Trichoderma: a review of biology and systematics of the genus

GARY J. SAMUELS
United States Department of Agriculture, Agriculture Research Service, Systematic Botany and Mycology Laboratory, Room 304, B-OI1A,
BARC-W, Beltsville, MD 20705-2350, USA.

Species of Trichoderma, a genus of hyphomycetes, are ubiquitous in the environment, but especially in soils. They have been used or
encountered in many human activities, including commercial applications in production of enzymes and biological control of plant
disease. They are the cause of disease in commercially produced mushrooms, and have been identified as causal agents of disease in
immunosuppressed humans. Knowledge about what constitutes a species of Trichoderma, or about interspecific relationships, has not
kept pace with the expanding number of applications or frequency of encounter of Trichoderma by biotechnologists, plant
pathologists and medical personnel. This review presesents an overview of the interaction between humans and Trichoderma, and a
more intensive review of knowledge of systematics and taxonomy of the genus.

Two-hundred years ago, when it was first described,
mycologists mistook Trichoderma Pers.: Fr. for a Gastero-
mycete (Persoon, 1794). It was only half a century later that
the true nature of the genus was realized, but movement
beyond the point of recognizing the class affinities of
Trichoderma to development of a species monograph has been
slow. While the only comprehensive taxonomic monograph
for the genus is admitted by its author to be artificial (Rifai,
1969 b), researchers in applied fields have been attributing to
Trichoderma species novel biological properties and activities.
Between the years of 1992 and 1995 alone, approximately
550 articles that cited the genus were catalogued in the United
States Department of Agriculture database AGRICOLA. I will
not review all those papers here, but refer the reader to the
excellent reviews available for the fields of biological control
(Papavizas, 1985; Chet, 1987, 1993; Lumsden, 1992) and
enzymology (Kubicek et aL 1990). A comprehensive bib-
liography for Trichoderma up to 1980 is found in Domsch,
Gams & Anderson (1980). In the present review I will present
some of the major advances in our knowledge of the biology
of Trichoderma, followed by a more intense review of the steps
taken to understand the systematics of the genus.
BIOLOGY
Trichoderma has not yet, given us any 'wonder drugs' such as
penicillin, but the ability of some species to produce enzymes
and/or to attack or inhibit other fungi has attracted major
research efforts in several areas, including biological control of
plant disease, and enzyme production, as well as in studies of
genetic control and manipulation in filamentous fungi. At a
time in human history when we acknowledge the need to
appreciate the economic, if not aesthetic, value of our
biological diversity, Trichoderma stands as a readily exploitable
source. Whether and how this source can be utilized will
depend upon understanding the biology of the species.
Trichoderma is a genus of filamentous deuteromycetes. Its
members are generally found in all soils including forest
humus layer (Wardle, Parkinson & Waller, 1993) as well as in
agricultural and orchard soils (see Chet, 1987; Roiger, Jeffers
& CaldwelL 1991 and references therein). Davet (1979) and
Smith, Wilcox & Harman (1990) have described media
selective for Trichoderma. Trichoderma species are rarely
reported to occur on living plants and have not been found as
endophytes of living plants (Petrini, 1986 & pers. comm.).
Danielson & Davey (1973) reported that individual species
showed preference for soil temperature and moisture content
of forest soils. Widden & Aribtol (1980) found a seasonality
to species distribution, although the seasonal occurrence of
individual species may be influenced by competition provided
by other Trichoderma species. Trichoderma viride Pers. does
seem to be adapted to cooler climates (see Roiger et al., 1991,
and references). Trichoderma species may be sensitive to some
environmental pollution, as is indicated by the low rate of
recovery of T. viride from coniferous forests that had been
subjected to alkaline dust for a period of 25 years; the high pH
(6.6) of the humus layer was blamed (Fritze & Baath, 1993).
While Trichoderma species are generally considered to be
aggressive competitors, Wardle et al. (1993) found that the
ability to compete was species dependent. Thus, Mucor
hiemalis Wehmer displaced T. harzianum Rifai from the forest
soiL but was itself induced to produce more conidia at the
expense of biomass by T. polysporum (Link: Fr.) Rifai in forest
humus. Some niche differentiation was suggested, as both of
the species mixtures had higher biomass of the fungi after 47
days than the monocultures of each species.
Physiological activities of Trichoderma species have been
Trichoderma: a review of biology and systematics of the genus
both beneficial and harmful to human enterprise. The ability
of Trichoderma reesei E. G. Simmons to break down cellulosic
materials through the production of cellulase enzymes has
lead to their commercial exploitation (see Reese & Mandels,
1989; El Gogary et al., 1990; Kubicek et aI., 1990; Cuevas,
Culiat & Manaligod, 1991; Bedford & Classen, 1993) in areas
as diverse as clothes-washing detergent. animal feed and fuel
production. Cellulases from T. reesei also have the potential of
bleaching kraft pulp, thereby providing an alternative to
conventional bleaching with chlorine (Buchert et al., 1994).
The effluent from chlorine bleaching contains environmentally
problematiC chlorinated hydrocarbons. Cellulolytic enzymes
produced by T. viride and T. reesei help the sugar industry to
dispose of sugar by-products, molasses and sugar beet pulp by
transforming the sugars to protein, that can be fed to
ruminants (Nigam, 1994). Katayama & Matsumura (1993)
have found that T. harzianum possesses an oxidative system
that is capable of degrading the organochlorine pesticide
endosulfan. Chitinolytic enzymes from T. harzianum and T.
virens O. H. Miller, Giddens & A. A. Foster) Arx (formerly
known as Gliocladium virens J. H. Miller, Giddens & A. A.
Foster) are thought to be responsible for degradation of
fungus cell walls and, thereby, effective in biological control
of the fungal pathogen Botrytis cinerea Pers.: Fe. (Cruz et al.,
1992; Di Pietro et al., 1993; Lorito et al., 1993, 1994). On the
other hand, those same enzymes are responsible for failure of
the shi'itake (Lentinula edodes (Berk.) Pegler) crop in Japan
(Komatsu, 1976; Tokimoto, 1982) through disruption of the
mushroom mycelium.
Trichoderma harzianum and T. virens (mostly under the name
Gliocladium virens) are the most commonly cited species in
biological control (see reviews in Papavizas, 1985; Chet,
1987). Trichoderma harzianum, alone or in combination with
other Trichoderma species or chemical adjuvants, has been
used in control of several diseases. Some of these include:
Rhizoctonia damping-off in radish (Lifshitz, Lifshitz & Baker,
1985), com and soybean (Kommedahl et a!" 1981); grey-
mould on tomato (Migheli et al., 1994), grapes and straw-
berry (Elad ef al., 1995; Harman et aI., 1995); Colletotrichum
storage rot of apple (Tronsmo & Hjeljord, 1995); cucumber
fruit rot caused by Rhizoctonia solani J. G. Kuhn (Lewis &
Papavizas, 1980). take-all disease in wheat (Ghisalberti &
Sivasithamparam, 1991), and sclerotinia sclerotiorum (Lib.) de
Bary in pea (Knudsen & Eschen, 1991) and 5. minor Jagger in
lettuce drop (Vannacci et al., 1991); and a wilt-complex,
predominantly caused by Sclerotium rolfsii Sacc., Rhizoctonia
solani and Fusarium oxysporum Schldl. in lentil and chickpeas
(Mukhopadhyay, 1995). In addition to a direct role of T.
harzianum in individual applications, it is effective in restoring
suppressiveness to heat-treated media amended with com-
posted hardwood bark (Nelson & Hoitink, 1983). Soils amended
with T. harzianum can be disease suppressive, have greener
turf, probably by enhanced root growth, and reduced brown
patch (Rhizoctonia soIani). dollar spot (Sderotinia homeocarpa
F. T. Benn.), and Pythium blight (Lo, Nelson & Harman, 1995).
Trichoderma harzianum also produces volatile chemicals that
arrest wood rot induced by basidiomycetes (Morrell. 1990).
Various Trichoderma species gave control of Phytophthora
crown rot of apple seedlings in Wisconsin (Roiger, Jeffers &
924
Caldwell, 1991). Trichoderma virens and T. longibrachiatum
Rifai were useful in control of groundnut root and stem rot
diseases caused by Rhizoctonia solani in India (Sreenivasaprasad
& Manibushanrao, 1993). Trichoderma harzianum, T. parcer-
amosum Bissett, and T. viride showed in vitro antagonism
against Cryphonectria parasitica (Murrill) M. E. Barr, the cause
of chestnut blight (Arisan-Atac, Heidenreich & Kubicek,
1995). Trichoderma species, especially T. viride, when used in
combination with fumigants, were positively correlated with
a decline in viability of Phellinus weirii (Murrill) R. W. Gilb.
in live roots of Douglas-fir (Pseudotsuga menziesii) (Nelson,
Theis & McWilliams, 1995). The authors also suggested the
use of Trichoderma strains in combination with lower levels of
fumigant for control of root rot in individual trees of high
value.
Trichoderma species have been used in commercial prepara-
tions for biological control of fungal-induced plant diseases
(Lumsden, Lewis & Locke, 1993). Trichoderma harzianum is the
active ingredient in TRICHODEX<tB>, which is used against
postharvest rot of apple, and is combined with T. polysporum
in the product BINAB-T<tB>, which is used in control of wound
decay and wood rot (Ricard, 1981). Trichoderma harzianum
AG2, prepared from protoplast fusion. protects against a wide
range of soil-borne plant diseases (Harman, 1990). GlioGard<tB>
has T. virens as its active ingredient and is used to prevent
damping-off in seedlings caused by Pythium and Rhiwctonia
species (Lumsden & Locke, 1989). Unidentified species of
Trichoderma are active in two products used in control of
deadarm disease of vines and stone fruit in New Zealand
(Trichoject and Tricho Minidowels; Hunt & Gale, 1995).
No single mode of action of Trichoderma species against
fungal plant parasites is known; for a review of this subject see
Chet (1987). Control of Rhizoctonia solani and Pythium ultimum
by Trichoderma species, including T. harzianum, may be
effected through direct penetration of host hyphae (Dennis &
Webster, 1971; Benhamou & Chet, 1993). Inbar & Chet
(1992) have found that mycelium of T. harzianum is
preferentially attached to nylon fibres that had been coated
with a plant lectin (concanavalin A) or a lectin derived from
the cell wall of Rhiwctonia solani. Extracellular enzymes.
including j3-1,3-glucanase, chitinase and cellulase (Elad, Chet
& Henis, 1979; Cruz et aI., 1993; Lorito et al., 1994; Harman
et aI., 1995) are effective in disrupting the mycelium of the
pathogen. On the other hand, mycoparasitism is not the
primary mechanism of biocontrol of Pyfhium ultimum and R.
solani by T. virens (Howell. 1987; Lumsden et al., 1992).
Howell (1987) showed that mutants of strains of T. virens
normally parasitic on R. solani had lost their mycoparasitic
ability but retained the antibiotic capability of the parent
strains. Later Howell & Stipanovic (1991) divided strains of T.
virens into two groups on the basis of antifungal antibiotic
production. One group. designated as 'Q' strains, produced
gliotoxin and were effective against Rhizoctonia solani but
inactive against Pythium ultimum Trow. The other group,
designated as 'P' strains. produced gliovirin and were strongly
active against P. ultimum but were inactive against R. solani.
Inhibitory volatile substances were also suggested as the
means of biocontrol of Botrytis cinerea mould of snap beans by
T. hamatum (Nelson & Powelson. 1988). Ghisalberti &
G. J. Samuels
Rowland (1993) found several antifungal metabolites produced
by T. harzianum to suppress growth of take-all (Gaeumanno-
mycesgraminis (Sacc.) Arx & D. L. Olivier). Faull, Graeme-Cook
& Pilkington (1994) studied production of an isonitrile antibiotic
by a uv-induced mutant of T. harzianum and its activity
against soil-borne plant pathogens (Faull & Scarseletti, 1994).
Antifungal antibiotics produced by Trichoderma species are
reviewed by Ghisalberti & Sivasithamparan (1991).
While Trichoderma species are generally considered either
useful to humans or at least ecologically neutraL this is not
always the case. Taylor (1986) reported that members of
Trichoderma and Gliocladium produce some of the most toxic
substances known. These include trichothecenes (Adams &
Hanson, 1972; Corley, Miller-Wideman & Durley, 1994) and
'pentaibols', polypeptides containing a high proportion of a-
aminoisobutyric acid that exhibit toxicity to experimental
animals (see Scott, 1991).
Trichoderma species are not generally known to affect
human health, but there are scattered reports indicating
pathogenicity to humans. Trichoderma viride and T. Iongi-
brachiafum, respectively, have been involved in complications
of renal dialysis (Loeppke ef al., 1983; Tanis ef aI., 1995). A
liver transplant recipient developed infection of a perihepatic
haematoma due to Trichoderma viride, and despite surgical
removal and treatment with amphotericin B, abundant
quantities of the Trichoderma were recovered from the
haematoma (Jacobs ef aI., 1992). More recently Trichoderma
species (T. Iongibrachiafum, T. pseudokoningii Rifai) were
identified as the cause of death in immunosuppressed humans
in the U.s.A. (D. A. McGough, pers. comm.).
Trichoderma species are not involved in plant parasitism or
in postharvest crop loss. However, because of its mycoparasitic
abilities, T. harzianum does cause serious losses in commercial
mushroom production. This includes shi'itake and Agaricus
mushrooms in Japan (Komatsu, 1976) and other parts of the
world (Muthumeenakshi ef al., 1994; Speranzini ef al., 1995; P.
Romaine, pers. comm.).
Most Trichoderma strains are not encountered in association
with sexual stages, and are considered to be strictly mitotic,
clonal fungi. The apparent lack of sexuality is a barrier to
understanding interrelationships within and among Tricho-
derma species. Moreover, the absence of outcrossing denies a
means of genetic manipulation which has caused dismay for
authors such as Papavizas (1985). One of the most important
developments in Trichoderma biology has been the discovery
of the ability to fuse protoplasts of genetically diverse strains
or even species. Improved biological control and industrial
strains have been found among the nonparental progeny
resulting from the fusion of isolated protoplasts (Pe'er & Chet,
1990; Stasz & Harman, 1990; Harman & Hayes, 1993; Meza
ef al., 1995); and genetic linkage data concerning enzyme
markers have been obtained from protoplast fusions in T. reesei
(Bawa & Sandhu, 1994). Stasz & Harman (1990) could not
explain the resultant variation in progeny of crosses involving
T. viride, T. hamafum (Bonord) Bainier, and T. koningii Oudem.
in terms of classical parasexuality. However, Bawa & Sandhu
(1994) used auxotrophic and cellulolytic characterization to
demonstrate the presence of haploid parasexual progeny.
Strains identified as T. harzianum have also been improved for
925
biocontrol applications through transformation with bacterial
DNA, and the transformants have remained stable in the
environment (Sivan & Harman, 1991; Migheli ef al., 1994).
Meza ef al. (1995) transferred a benomyl resistance marker
between strains of T. reesei, and selected progeny of protoplast
fusion were higher in cellulolytic ability than either parent.
Shin & Cho (1993) obtained recombinant, interspecific fusion
progeny when protoplasts of biological control strains of T.
virens and T. harzianum were crossed. The ability to generate
protoplasts has also provided an opportunity to separate
chromosomes and to localize genes on individual chromo-
somes, as has been done for T. reesei (Gilly & Sands, 1991;
Mantyla ef al., 1992).
Sexual reproduction is known in Trichoderma in the sense
that the only known teleomorphs of Trichoderma are species
of Hypocrea Fr. and closely related genera, members of the
ascomycete order Hypocreales. There are few documented
cases of Hypocrea species undergoing the sexual cycle in
artificial culture; the best known is Hypocrea spinulosa Fuckel
(as Chromocrea spinulosa (Fuckel) Petch). In this species sexual
reproduction is under control of one locus and two alleles, but
half of the progeny appear to be self-fertile owing to
unidirectional shifting to the compatible mating type in some
mycelial nuclei (Mathieson, 1952; Perkins, 1987). A similar
phenomenon was suggested by Canham (1969) for H. cifrina
(Fr.) Fr. var. cifrina and var. americana Canham, and by Samuels
& Lodge (1996) for H. poronioidea Moller. In the case of H.
poronioidea, however, Samuels & Lodge (1996) were not able
to induce crossing between the self fertile and self sterile
strains and suggested that half of the progeny of meiosis were
sexually incompetent. It should be added that Samuels &
Lodge (1996) did not use auxotrophic mutants or other
genetic markers, so that the possibility of crossing between
self fertile and self sterile strains cannot be excluded. On the
other hand, Samuels, Petrini & Manguin (1994) observed a
normal bipolar segregation of mating type in asci of H. jecorina
Berk. & Broome. Rehner (pers. comm.), using amplified
fragment length polymorphisms of nuclear rONA, demon-
strated the ability of widely geographically separated strains
of H. jecorina to undergo meiotic recombination. Hypocrea
jecorina, the teleomorph of T. reesei (Kuhls ef al., 1996a),
represents an excellent model system for classical genetic
studies in Trichoderma.
SYSTEMA TICS
Toward a concept of genus
While it is difficult to define, there is a general concept of
a 'basic' Trichoderma morphology (see for example Rifai,
1969; Bissett, 1984, 1991a-c, 1992; Fig. 4): rapid growth;
abundant powdery, green conidia; and ill-defined conidio-
phores. That this morphology has been unmistakable is
attested to by the paucity of generic synonyms: only four
taxonomic synonyms were uncovered by Rifai (1969). The
most persistent of the generic synonyms was Pachybasium
Sacco (1885). Pachybasium was proposed for species of
Verficillium Nees that have sterile elongations on the
conidiophores (Fig. 2), but Bainier (1906) soon recognized
Pachybasium to be synonymous with Trichoderma. Within
Trichoderma: a review of biology and systematics of the genus 926

3
00
00

.0Cbo

Figs 1-5. Trichoderma and Hypocrea. Fig. 1. Tolypocladium niveum, ex neotype (ARSEF 3280). Fig. 2. Trichoderma polysporum-aggr.
anamorph of Hypocrea sp. (G]S 98-121). Fig. 3. Trichoderma minutisporum Bissett, anamorph of Hypocrea sp. (CBS 901.72). Fig. 4.
Trichoderma viride anamorph of Hypocrea rufa (G]S 90-97). Fig. 5. Hypocrea pulvinata anamorph (G]S 91-220). Scale bars = 10 11m.

Trichoderma, the Pachybasium-types can be distinguished from
Trichoderma sensu stricto in having white conidia produced
from doliform phialides that are held in botryose clusters, and
often with a sterile or terminally fertile extension (Fig. 2).
Although most authors, including Hughes (1958) in his critical
evaluation of hyphomycete genera, did not accept Pachy-
basium, a few new species were described in the genus as
recently as 1961. While Pachybasium is not now recognized as
a genus, Bissett (1991a, b) accepted the section Pachybasium in
Trichoderma.
The morphological concept of Trichoderma is not completely
settled. Although the Trichoderma morphology is usually
unmistakable, there is morphological intergradation with
other hyphomycete genera. Carmichael et ai. (1980) questioned
whether Toiypocladium W. Cams could be a synonym of
Trichoderma, possibly because of a similarity in arrangement of
G. J. Samuels 927

'.
3

'.

Figs 6-9. Trichoderma and Gliocladium. Fig. 6. Trichoderma virens. ex type culture. Fig. 7. Gliocladium viride (J. P. Jones Gl 114). Fig. 8.
Gliocladium prnicillioides, anamorph of Sphaerostilbella aureonitens (GJS 83-286). Fig. 9. Trichoderma anamorph of H. flavovirens (GJS
95-154). Scale bars = 10 11m.

phialides of T. inflatum W. Gams (= To. niveum (0. Rostr.)
Bissett; Fig. 1) to that of Trichoderma polysporum (Link:Fr.)
Rifai (Fig. 2). However. the morphological comparison is only
superficial as To. niveum is now known to be the anamorph of
Cordyceps facis Kobayashi & Shimizu (Hodge & Krasnoff,
1995; Hodge, Krasnoff & Humber, 1996), a member of the
Clavicipitales.
The inclusion of Gliocladium virens (Fig. 6), one of the most
frequently cited of the biological control fungi, in Trichoderma
by Arx (1987) has only recently been accepted following
DNA sequence analysis (Rehner & Samuels, 1994, 1995). The
penicillus of phialides, combined with conidia held in slime, all
on a more or less discrete conidiophore (Fig. 6) are generally
characteristic of Gliocladium Corda. Trichoderma virens is not
Trichoderma: a review of biology and systematics of the genus
the only Trichoderma to have a generalized Gliocladium-type
conidiophore, which is found among anamorphs of Hypocrea
species such as H. gelatinosa (Webster, 1964; Bissett, 1991b)
and various other Hypocrea species (Doi, 1972; e.g. H.
flavovirens Berk. & Broome, Fig. 9). When viewed in light of
the different teleomorphs and biologies, the morphological
similarity between T. virens and the type species of Gliocladium,
G. penicillioides Corda (Fig. 8) is not indicative of close
phylogenetic relationship.
Gliocladium penicillioides, and similar Gliocladium species, are
anamorphs of species of Sphaerostilbella (Seifert, 1985).
Sphaerostilbella Henn. species occur on basidiomata of members
of the Aphyllophorales and the genus is more closely related
to Hypomyces Tul. than it is to Hypocrea (Rehner & Samuels,
1995). Further, many unrelated hypocreaceous anamorphs are
classified in Gliocladium because they have penicillately
arranged phialides (e.g. G. roseum Bainier, which is a
Cionostachys Corda; see illustrations in Domsch et al., 1980).
Bissett (1991 b) included T. virens in Trichoderma sect.
Pachybasium Bissett on the basis of morphological comparison
to other species that he included in that section. From the
point of view of teleomorphs, T. virens is probably closely
related to T. aureoviride Rifai, the anamorph of H. aureoviridis
Plowr. & Cooke, a species that is morphologically and
anatomically similar to H. gelatinosa Tode: Fr.
Rehner & Samuels (1994) compared various hypocrealean
ascomycetes that have Gliocladium anamorphs using sequences
of large subunit nuclear ribosomal DNA and found T. virens,
G. viride, G. roseum, and G. penicillioides to be widely dispersed
in the Hypocreales. Trichoderma virens was arranged among
Hypocrea species, and can thus be regarded as a species of
Hypocrea. The consequence of this finding for Trichoderma
taxonomy is that the morphological stereotype of Trichoderma
has to be modified to accept this stereotypical Gliocladium
morphology. In the case of G. virens, however, it is not too
difficult to rationalize the Gliocladium morphology within
Trichoderma because (i) of the intergrading Hypocrea anamorphs
and (ii) the branching pattern of G. virens can be seen to be a
modification of a more typical Trichoderma, (iii) the formation
of typical Hypocrea-like chlamydospores (Figs 5, 6), and (iv)
the formation of green conidia.
In his subdivision of Trichoderma into sections, Bissett
(1991 b) tended to the philosophy that Hypocrea anamorphs
are, a priori, species of Trichoderma. However true that may be
in phylogenetic terms, inclusion of all Hypocrea anamorphs in
Trichoderma may require an unacceptable expansion of the
morphological concept of the genus. Gliocladium viride Matr.
(formerly known as G. deliquescens Sopp, Fig. 7) is the
anamorph of Hypocrea Iutea (Tode: Fr.) Petch, although the
species is found commonly in its anamorph form from soil
isolations. Hypocrea lutea is morphologically, biologically and
genetically similar to H. gelatinosa (Rehner & Samuels, 1994,
1995). Although this Gliocladium-like anamorph is not difficult
to distinguish from G. penicillioides, it does not bear much
morphological similarity to more typical Trichoderma species.
While acknowledging that G. viride behaves, genetically, like
a Hypocrea species, and thus like any other Trichoderma
species, it represents the diagnosticians worst nightmare: it
would be impossible to determine this species as a Trichoderma;
928
in any current taxonomic treatment it would be placed in
Gliocladium. For a general discussion of the integration of
anamorph fungi into the ascomycetes see Cams (1995).
Bissett (1991 a) expanded the morphological concept of
Trichoderma by including anamorphs of species of Hypocrea
that have effused stromata in sect. Hypocreanum Bissett. These
Hypocrea species include H. pulvinata Fuckel (Rifai & Webster,
1966b; Fig. 5), H. citrina (Rifai & Webster, 1966b; Canham,
1969), and others. Their anamorphs have colourless conidia
borne in conspicuous clear and uncoloured liquid on more or
less verticillately branched, or unbranched and then Acrem-
onium- or Verticillium-like conidiophores. Bissett (1991a)
envisioned a progression of forms among the Hypocrea
anamorphs that ranges from these Acremonium-or Verticillium-
like to more typical Trichoderma conidiophores (e.g. H. rufa
(Pers.: Fr.) Fr., Fig. 4).
Preliminary results of sequence analysis of rDNA (Rehner.
peTS. comm.) place these Hypocrea species with effused
stromata within the larger genus Hypocrea. However, care
should be taken before accepting the placement of the
anamorphs of these fungi in Trichoderma, as it is at least
possible that rather than being Trichoderma anamorphs, they
are synanamorphs or even spermatia. Hypocrea poronioidea, a
Hypocrea species with a discrete, turbinate stroma, produces
both a typical Trichoderma anamorph as well as a synanamorph
that is morphologically similar to the anamorphs of H.
pulvinata. In H. poronioidea, however, these colourless conidia
apparently only form on incipient stromata, and this suggests
that they have a sexual function (Samuels & Lodge, 1996).
Furthermore, on the basis of rDNA sequence similarity, H.
poronioidea is basal to the species of Hypocrea that have effused
stromata, and closer to them than it is to other Hypocrea
species (Rehner, pers. comm.). Conceivably the formation of
spermatia, or at least Acremonium-like anamorphs, is an
apomorphy for this group. Hypocrea poronioidea, hypothesized
as being primitive, retains the ability to form Trichoderma
conidia as well as the Acremonium-like conidia whereas the
ability to form Trichoderma is lost among the more derived
members.
Toward a concept of species
As is usually the case, species of Trichoderma have been
defined on the basis of their morphology. Morphological
characters used in species recognition in Trichoderma have
been outlined by Rifai (1969b) and Bissett (1984, 1991a-c,
1992). Unfortunately, some of the richest characters for
species recognition in hyphomycetes in general are either not
variable enough, or are difficult to describe in Trichoderma.
Conidial size and shape, an extremely useful character in other
genera, are of limited value in Trichoderma. Conidia of most
species are less than 5 I.lm long and wide. Conidia may be
globose, subglobose, ellipsoidal or oblong, and the shape is
useful in recognizing groups of species, but within groups its
value is diminished. Conidia may be some shade of green or
greenish yellow, or colourless. The differences in shade of
green, which may range from a deep green to nearly grey,
may be taxonomically significant, but difficult to interpret and
communicate. Conidial ornamentation, which may be smooth,
G. J. Samuels
warted or tuberculate, is certainly a useful species character
but may require scanning electron microscopy to demonstrate.
There are usually no definable conidiophores in Trichoderma
(see, for example, Figs 2-4). The conidia tend to aggregate
into pulvinate masses, the aggregates formed of intertwined
hyphae bearing phialides and often appearing to be attached
to the substrate at one or a few points. An ontogenetic study
of such aggregates could be fascinating, and could also help in
defining a conidiophore. For example, do aggregates originate
with a single hypha that proliferates7 The width of the hyphae
within the aggregate is significant. Bissett (1991 b) has made
extensive taxonomic use of fertile or sterile elongations of
conidiophores (Fig. 2). The arrangement of phialides on
hyphae within aggregates is characteristic but is virtually
impossible to define and communicate (compare Figs 2-4).
Phialide shape is characteristic of a species but, again, is
difficult to define. Perhaps new mathematical definitions can
be developed that would reflect form of phialides and patterns
of branching more accurately than the available words. Seifert,
Wingfield & Wingfield (1995) have suggested that branching
in Trichoderma resembles 'iterated functional systems' that are
derived from mathematical formulae.
Chlamydospores in Trichoderma tend to be globose to
subglobose, terminal or intercalary in hyphae, smooth, green,
and less than 15 ~m in diameter (Figs 5-7). While these
chlamydospores are characteristic of Trichoderma and Hypocrea,
their form is not diagnostic of species but their presence may
be (Samuels, Doi & Rogerson 1990; Bissett 1991 b).
Synanamorphs, apart from chlamydospores, are rare in
Trichoderma. Hypocrea suIawesensis Yoshim. Doi (Samuels et al.,
1990) produces solitary' macroconidia' at the tips of branches
of conidiophores of its Trichoderma anamorph. In H. poronioidea
(Samuels & Lodge, 1996), hyaline conidia held in drops of
colourless liquid at the tips of sparingly branched conidio-
phores form in addition to a typical Trichoderma anamorph.
As was discussed above, these hyaline conidia may be
spermatia.
Characteristics of colonies grown on agar media are subtle.
Most species grow rapidly and conidiate readily on common
media. Unlike comparable genera such as Penicillium Link or
Fusarium Link, where cultures present many diagnostic
characters, in Trichoderma there is virtually no aerial mycelium
and most diffusable pigmentation is typically in shades of
yellow. Yellow pigment is a characteristic for some species of
sect. Longibrachiatum. Trichoderma colonies often show zo-
nation in conidial development; Schrufer & Lysek (1990)
demonstrated that rhythmic growth and sporulation are
heterogeneous in populations. They also reported a de-
pendence on light for induction of sporulation. Whether
conidial production occurs in aggregates, or predominantly on
'mononematous' conidiophores diffused throughout the
colony, may be a species character. It has been my observation
that a relatively weak medium such as cornmeal agar with 2 %
dextrose permits better observation of conidial aggregation
when compared to media such as potato dextrose or oatmeal
agar, where conidial production is generally much more
profuse. However, diffusable pigment forms more reliably on
potato dextrose agar than cornmeal agar with dextrose or
oatmeal agar. Some cultures develop sweet odours that are
929
sometimes reminiscent of coconut. The coconut odour
produced by some strains of T. viride and T. harzianum has
been attributed to an antifungaL unsaturated pyrone, 6-
pentyl-a-pyrone (Serrano-Carreon et aI., 1992; see also
Claydon et ai., 1977).
Rifai (1969) concluded, not surprisingly, that anamorph
characters alone might not provide a useful taxonomy of
Trichoderma. He took the attitude that there is no real way to
define a biological species in morphological terms, that the
morphological characters were continuously variable, and that
there is no way of knowing the degree of variation tolerable
within an individual species. One way to chart variation
within a biological species of Trichoderma is to use anamorphs
of known Hypocrea species, so that there would be no doubt
that all strains, derived from ascospores as they are, would
represent only one species. Because, prior to 1969, the number
of Hypocrea species that had been grown in pure culture was
smalL Rifai refrained from proposing narrow species concepts
in Trichoderma. He adopted instead the concept of 'aggregate'
species, which he defined as ' aggregations of morphologically
very similar and often hardly separable species.' Of these
aggregates, five were based on Hypocrea species, viz. T.
aureoviride Rifai (H. aureoviridis), T. hamatum Rifai (H. semiorbis
(Hook.) Berk.); T. piluIiferum Webster & Rifai (H. piIuIifera
J. Webster & Rifai), T. pseudokoningii (H. d. schweinitzii (Fr.)
Sacc.), and T. viride (H. rufa). Although he considered that his
work was preliminary and not to be taken as a complete
taxonomic treatment, it has become the primary source of
species identification for Trichoderma.
Bissett (1984, 1991a-c, 1992), like Rifai, adopted a
morphological approach to taxonomy of Trichoderma. He
essentially elevated each of Rifai's aggregate species to
sectional level (Bissett, 1991a) and proposed one new section
for Acremonium-like or Verticillium-like anamorphs of the
Hypocrea species, such as H. puIvinata, that have effused
stromata. Doi, Abe & Sugiyama (1987) added an additional
section, sect. Saturnisporum for species that have warted,
wrinkled or nearly winged conidia. Bissett has published
revisions of sections Longibrachiatum (Bissett, 1984) and
Pachybasium (Bissett, 1991 b) and recognized several pre-
sumably biological species in each.
In the publications of Rifai (1969) and Bissett (1984, 1991
a, b) we have taxonomies that reflect differing taxonomic
philosophies. Bissett has perceived noncontinuous morpho-
logical characteristics of biological species where Rifai saw a
continuum within a few basic morphologies. Is one system
any more reflective of reality than the other? Independently
derived, macromolecular, data have been applied in tests of
the morphotaxonomic hypotheses.
Macromolecular approaches to Trichoderma taxonomy
The first efforts at macromolecular characterization of
Trichoderma strains and species were undertaken not by
professional taxonomists, but by frustrated users of the
existing taxonomy. Stasz et aI. (1988, p. 170) plaintively
remarked that, ' ... methods are lacking to differentiate among
strains for patent purposes, or to determine the variability and
abundance of strains in natural ecosystems'. Morphology
alone in Trichoderma has not led to a satisfactory taxonomy or,
Trichoderma: a review of biology and systematics of the genus
at any rate, to a taxonomy that has been useful to many users.
Characters derived from nucleic acids and enzymes are
attractive because, with cladistic analysis, they seem to offer
the possibility of greater objectivity than do traditionally
observed and analyzed data. Whether these techniques
increase taxonomic objectivity may be debated, but the
additional characters are certainly welcome.
Macromolecular analyses based on enzymes and nucleic
acids have shown that at least some of the aggregate species
are phylogenetically based (e.g. Okuda, Fujiwara & Fujiwara,
1982; Stasz et aI., 1989; Leuchtmann, Petrini & Samuels, 1996)
at the same time confirming genetic diversity of the individual
aggregates (Zamir & Chet, 1985; Stasz et aI., 1989; Meyer,
1991; Meyer et al., 1992; Fujimori & Okuda, 1994;
Muthumeenakshi ef al., 1994; Samuels ef al., 1994; Zimand ef
aL 1994; Kuhls et al., 1996a, b; Leuchtmann ef aI., 1996).
Stasz et al. (1989) evaluated five aggregate species of
Trichoderma using enzyme polymorphisms. While they
concluded that morphological species are not characterized by
specific alleles at single loci, or specific patterns of alleles at
multiple loci, they demonstrated what they called 'core
groups' of morphological species. Despite individual strains
that gave widely divergent allozyme patterns, the core groups
coincided well with Rifai's aggregate species T. pseudokoningii,
T. koningii, T. hamafum, and T. viride. Strains used under the
name of T. harzianum aggregate fell into one of two clusters.
These results indicate that, in part. there is a genetic basis for
the morphological aggregate species, and also confirm Rifai's
own contention that each aggregate is genetically het-
erogeneous.
Meyer & Plaskowitz (1989) observed two types of conidial
ornamentation among twelve strains referable to T. viride, a
member of sect. Trichoderma. Meyer (1991) later found that
mitochondrial DNA restriction fragment data distinguished
between those two groups and he suggested that each group,
characterized by conidial ornamentation and mtDNA type,
could represent a distinct species. This is an interesting
correlation that should be confirmed by the study of more
strains if only because T. viride is the type species of the genus.
Okuda et al. (1982) found that the aggregate species T.
hamatum, T. harzianum, T. koningii, and T. viride could be
subdivided on the basis of production of isonitrile antibiotics.
There was some correspondence between the respective
groups and phenotypical characters of the strains.
Zamir & Chet (1985) divided twenty-three strains of the T.
harzianum-aggregate among five different types according to
their enzyme profiles. Fujimori & Okuda (1994) and
Muthumeenakshi et al. (1994), also working with the T.
harzianum-aggregate, used various molecular techniques to
distinguish, respectively, two and three groups within the
aggregate. Muthumeenakshi et ai. (1994) found three distinct
types of ITS-I, and strains characterized by ITS type 2 were
aggressive antagonists in commercial mushroom production.
In this regard it is interesting that Stasz et al. (1989), using
isozyme analysis, also found two enzyme profiles in T.
harzianum.
Fujimori & Okuda (1994) successfully used RAPD's to
identify duplicate Trichoderma strains in microbial screening.
Kuhls, Lieckfeldt & Borner (1995) used RAPD's of T. reesei to
930
determine that T. 'todica' (ATCC 36936), an unpublished
strain patented for the production of antiviral antibiotics, is
actually T. parceramosum, a member of sect. Longibrachiafum
Bissett. Schlick et ai. (1994) used DNA fingerprinting to
identify patent strains of T. harzianum. They also found that
strains that had been produced by gamma irradiation had the
same ITS-1 and ITS-2 sequences as the parent strains. Zimand
et al. (1994) used the RAPD procedure to identify biocontrol
strains of Trichoderma species.
Bissett (1984) elevated the T. Iongibrachiatum-aggregate
species to status of section and included the species T.
citrinoviride, T. Iongibrachiafum, T. parceramosum, T. pseudo-
koningii, and later the anamorph of H. schweinitzii (Bissett,
1991c). Samuels et ai. (1994), using isozymes, and Kuhls et ai.
(1996a, b), using a variety of DNA techniques, confirmed that
sect. Longibrachiatum is distinct from other sections. Bissett
(1984) included the species T. pseudokoningii, which Rifai
(1969) regarded as an aggregate species, in sect. Longi-
brachiatum. Trichoderma pseudokoningii was originally isolated
from ascospores of Hypocrea d. schweinifzii in eastern Australia,
and Leuchtmann et a/. (1996) found that T. pseudokoningii is
similar to other Hypocrea strains found in New Zealand. They
also concluded that T. pseudokoningii, a species commonly
reported in the literature, is most likely limited in distribution
to Australia and New Zealand and has probably been widely
misinterpreted (e.g. Bissett, 1984).
Trichoderma sect. Longibrachiafum includes T. reesei, a species
that is well known for cellulase production (Reese & Mandels,
1989) and that accounted for about half of the approximately
550 articles that cited Trichoderma in the USDA AGRICOLA
data base for 1992-1995. Bissett (1984) synonymized T. reesei
with T. Iongibrachiafum on morphological grounds. Meyer et
al. (1992), using DNA-fingerprint analysis, and Samuels ef ai.
(1994), combining morphometries and isoenzyme profiles,
distinguished between T. reesei and T. Iongibrachiatum. In
another isozyme study (Leuchtmann et al., 1996), using a much
larger set of strains than was used by Samuels et al. (1994), T.
reesei and H. jecorina were closely joined to each other and
formed a group that was distinct from all other species in sect.
Longibrachiatum, including other associated Hypocrea teleo-
morphs. Kuhls et al. (1996a), using rDNA sequence analysis
and PCR-fingerprints, observed T. reesei to behave exactly as
any strain of H. jecorina, concluded that T. reesei is the
anamorph of H. jecorina, differing in minor phenotypic
characters and in its inability to undergo sexual reproduction
(Samuels et aI., 1994).
Doi et al. (1987) proposed the new sect. Saturnisporum for
two species, T. ghanense Y. Doi, Y. Abe & Sugiy. and T.
safurnisporum HammilL species that have conspicuously
warted, sometimes alate, conidia. Branching patterns of both
species are similar to branching in sect. Longibrachiafum and,
like sect. Longibrachiatum, both have ellipsoidal conidia. Kuhls
et al. (1996b) and Turner et ai. (1996) used nucleic acid
analyses to place, respectively, T. safurnisporum and T.
ghanense in sect. Longibrachiatum.
The teleomorphs of Trichoderma
Trichoderma has all the essential characteristics of anamorphs
of the ascomycete order Hypocreales (Samuels & Seifert.
G. J. Samuels
1987): brightly to lightly coloured conidia, conidiophores and
colonies, and conidia formed from phialides. By the time the
first volume of the Transactions of the British Mycological Society
was published in 1902, the specific link between T. viride and
H. ntfa was an accepted fact (Smith, 1902), thus establishing
the generic link between Hypocrea and Trichoderma. Berkeley,
as early as 1860, suspected a link between l. viride and some
unnamed ascomycete when he enigmatically noted that the
Trichoderma species was' probably not autonomous.' In that
same year L.-R. Tulasne (Tulasne, 1860) proved that l. viride
and the ascomycete Hypocrea rufa are expressions of one life
cycle. The brothers Tulasne (Tulasne & Tulasne, 1865)
illustration of T. viride is remarkably accurate in the
representation of the phialides and their disposition on the
conidiophore. The observation of the relationship between H.
rufa and l. viride, later proven by single ascospore cultures by
Brefeld (1891), represented a major advance in our under-
standing of the interrelationship between deuteromycetes
and ascomycetes.
The only proven teleomorphs for Trichoderma have been
closely related genera, viz. Hypocrea with fewer in Podostroma
P. Karst. and sarawakus Boedijn. Our knowledge of all of
these genera is slowly developing, but a picture is emerging
wherein species of Podostroma and sarawakus that have
Trichoderma anamorphs could be placed in Hypocrea. Since
1860 many Hypocrea species have been grown in pure culture
and their Trichoderma anamorphs described (see list in Doi &
Doi, 1979).
Because only members of the Hypocreales have been
proven to have Trichoderma anamorphs, there is no doubt that
Trichoderma is a genus of that order. It is, however, somewhat
more difficult to assign Trichoderma to a single teleomorph
genus because the three, albeit closely related, ascomycete
genera have Trichoderma anamorphs. Ribosomal DNA sequen-
ces (Rehner & Samuels, 1994; Kuhls et al., 1996a), PCR-
fingerprints (Kuhls et al., 1996a, b; Turner et al., 1996) and
isozyme studies (Samuels et al., 1994; Leuchtmann et al., 1996)
have integrated individual Trichoderma species, not known to
reproduce sexually, among Hypocrea species.
Bisby (1939), unable to distinguish the Trichoderma
anamorphs of H. rufa and H. gelatinosa, referred to both as l.
viride. Because, in his estimation, the anamorphs were the
same, he considered the green-spored H. gelatinosa to be a
growth form of the white-spored H. ntfa. Bisby can, perhaps,
be forgiven for confusing the anamorphs of H. rufa and H.
gelatinosa because, in Bisby's time, several species of Hypocrea
could have been confused with H. gelatinosa, and all of them
have a Trichoderma anamorph that could have been confused
with l. viride (for example H. f/avovirens, Fig. 9). In 1964 the
anamorph of true H. gelatinosa was recognized to be a
Trichoderma with slimy conidia (Webster, 1964) similar to l.
virens. Ten years later Webster and Rifai published a series of
papers detailing connections between Hypocrea species and
their Trichoderma anamorphs (Webster, 1964; Rifai & Webster,
1966a, b; Webster & Rifai, 1968). In addition to H. gelatinosa,
this series established the current holomorphic concept of the
common species H. pulvinata and H. ntfa, and culminated in
the publication of Rifai's revision of Trichoderma in 1969.
The work of Y. Doi and his collaborators, which includes
931
Japanese, Asian Pacific and tropical American species, is the
most comprehensive study of Hypocrea available (Doi &
Doi, 1979, 1986; Doi, 1966 - 1968, 1971 - 1973a, b, 1975,
1976, 1978, 1980; Doi, Doi & Toyozawa, 1984; Doi &
Yamatoya, 1989). Unfortunately, Doi did not publish keys to
either teleomorphs or anamorphs. Domsch et al. (1980)
assigned many of the anamorphs described in Doi's work to
Rifai's aggregate species, but any aggregate could be linked to
more than one Hypocrea species. Doi (1972), in his revision of
Japanese Hypocrea, recognized two subgenera. He further
divided one of those subgenera into sections, subsections, and
series based on stromal anatomy. While there was broad
consistency within the subsections and series as to the type of
anamorph, anamorphs in sect. Hypocrea subsect. Hypocrea,
which includes the type species H. rufa, could be assigned to
one of four of Rifai's aggregate species. Thus, at least in the
case of one subsection, the anamorphs and teleomorphs are
telling different phylogenetic stories because it is not possible
to predict from most Hypocrea species the aggregate species of
the Trichoderma anamorph.
Hypocrea perithecia do not form in cultures of Trichoderma
strains isolated from natural substrates; however, Trichoderma
anamorphs occasionally develop in cultures derived from
Hypocrea ascospores. One can question whether Trichoderma
strains encountered in the absence of a teleomorph are parts
of Hypocrea life cycles. Some of the aggregate species
described by Rifai (I969) are Hypocrea species in the sense that
they are based on ascospore isolations from Hypocrea species
(see above).
However, given the difficulty of understanding what
constitutes a species of Trichoderma, it is still unclear whether
Trichoderma strains encountered in nature are actually Hypocrea
anamorphs. As was discussed above, T. reesei, while apparently
incapable of sexual reproduction and culturally distinct from
H. jecorina, behaves exactly like any strain of H. jecorina (Kuhls
et ai., 1996a). However, l. reesei is known only from its
original isolation and the mutants that have been derived from
it (EI Gogary et al., 1990).
Leuchtmann et al. (1996), Kuhls et ai. (1996 b), and Turner et
al. (1996) studied the relationship between T. citrinoviride (sect.
Longibrachiatum) and H. schweinitzii (Fr.) Sacc. Unlike l. reesei,
l. citrinoviride is a common and cosmopolitan species. In each
study, four strains of H. schweinitzii, three from eastern USA
and one from France, were interspersed with strains of l.
citrinoviride, some HypocrealTrichoderma pairs or clusters more
closely interrelated than individual pairs or clusters were
related to other individual pairs or clusters. There also
remained several l. citrinoviride strains that clustered singly or
alone, but not with Hypocrea strains. Unfortunately because H.
schweinitzii has not been induced to reproduce sexually in
vitro, no comment can be made about interfertility among the
various Hypocrea and Trichoderma strains or whether the l.
citrinoviride strains are sexually competent. There is little
doubt, however, that H. schweinitzii is the teleomorph of T.
citrinoviride. Thus, Hypocrea teleomorphs have been proven for
at least two commonly occurring species of Trichoderma, viz.
l. citrinoviride and T. viride. Bissett (1991 c) illustrated an
anamorph for H. schweinitzii that was based on a New Zealand
collection (CBS 243.63). New Zealand H. schweinitzii strains
Trichoderma: a review of biology and systematics of the genus
are genetically diverse, comprising at least two populations
(Kuhls et al., 1996b; Leuchtmann et al., 1996), both of which
may be distinct species and different from typicaL north
temperate H. schweinitzii.
How many species of Trichoderma are there 7
Just how many species are included in Trichoderma has been a
continuing matter of uncertainty. The genus was originally
proposed by Persoon in 1794 with four species, of which only
one, T. uiride, remains. In all of Saccardo's Sylloge fungorum
only 27 species of Trichoderma were included, and prior to
1980 there were fewer than fifty described species. The only
key to Trichoderma prior to 1939 was that of Gilman & Abbott
(1927), who recognized Pachybasium with one species, and
distinguished four species of Trichoderma on the basis of
colony characters, and conidial shape and pigmentation. Bisby
(1939) considered that colony characters and conidial shape
and ornamentation were too variable to define species, and
concluded that there was only one species, T. uiride. He noted
that while a small number of isolates might exhibit
characteristic colonies or morphology, apparent differences
were blurred on examination of many more cultures. Between
1939 and 1969 Trichoderma was basically seen to include only
one species. Obviously, literature about the biology of
Trichoderma from that period should be viewed carefully.
Today there are about seventy-five described species of
Trichoderma. Doi & Doi (1986) listed the species described
before 1986.
How many species are there really? The answer to this
question will depend upon the operational definition of a
species. If the example of T. reesei given above is representative,
then there could be an endless number of morphologically
distinct clonal species that are genetically very close to other
morphospecies. If one assumes that the majority of Hypocrea
represent 'good' species of Trichoderma, and if only half of
more than 200 described species of Hypocrea are 'good'
species, then there must be at least 100 biological species of
Trichoderma. But this would be an underestimation as it does
not account for the many Hypocrea species still to be
described.
The state of our knowledge of Hypocrea taxonomy is not
much further advanced than it is for Trichoderma itself. An idea
of the described us unknown species diversity in Hypocrea is
seen in Doi's (1972) monograph of the genus for Japan. Of the
fifty-two taxa (including forms) that he accepted, 31
(approximately 60%) were described as new. The majority of
these had Trichoderma anamorphs; while many could be
assigned at least to one of the aggregate species, it is evident
that there is more morphological variation than is accounted
for in Rifai's aggregates. So, while Domsch et al. (1980)
correctly noted that each of Rifai's nine species aggregates
could be connected with more than one teleomorph species,
much of the phenotypical diversity in Trichoderma remains
undescribed.
THE FUTURE?
Trichoderma has been shown to be a valuable source for the
commercial production of enzymes, for example, that can be
932
helpful in recycling cellulosic waste materials while producing
useful by-products. Trichoderma species are also a valuable
tool for the biological control of plant pathogens, thus helping
to reduce the need for polluting chemicals. A comprehensive
taxonomic scheme is urgently required to provide rapid
identifications, to distinguish deleterious strains, and to
adequately and safely survey the biodiversity, and realize the
commercial potentiaL of Trichoderma.
Emerging evidence has shown that some of Rifai's (1969)
aggregate species, such as T. longibrachiatum, are mono-
phyletic. Other aggregates, such as T. harzianum have been
proven to be perhaps even more diverse than Rifai had
envisioned. There has only been limited testing of more
narrowly circumscribed morphological species, but at least in
sect. Longibrachiatum the species proposed by Bissett (1984)
have been supported.
We still do not know what a species of Trichoderma is.
Development of a species concept in Trichoderma requires a
combination of phenotypic and genetic information derived
from repeated collection of Hypocrea and Trichoderma strains.
It may be possible to use phenotypic characters in the
identification of many species, but it must always be borne in
mind that strains that look different, and that in more 'naIve
times,' before the development of isozyme and nucleic acid
analyses, might have been described as distinct taxa, are not
necessarily genetically distinct. This is certainly the case with
T. reesei and H. jecorina. We cannot even begin to guess at the
number of so-called Trichoderma species that are nothing more
than clonal lines, genetically like but phenotypically distinct
from other lines. A challenge for the future will be to
understand how speciation occurs in Hypocrea and Trichoderma.
I am indebted to Ms Kathie Hodge for letting me see her
unpublished manuscript on Cordyceps facis, and for providing
the illustration of Tolypocladium inflatum. Dr S. A. Rehner
allowed me access to his unpublished data on Hypocrea and
Trichoderma. Drs M. E. Palm, C. T. Rogerson and A. Y.
Rossman read various drafts of this work and provided helpful
comments.
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