**4. Diversity of** *Trichoderma* **in tropical America**

Comparatively few comprehensive studies have been undertaken to assess the diversity of *Trichoderma* in neotropical regions. Since agriculture is a vital segment of local economies in the neotropics, most research in this region on *Trichoderma* has been directed to their biological control activities against phytopathogens. Studies have focused on biological control of plant pathogens with economical importance in cacao plantations,

among 96 isolates tested (Bissett et al., 2003). The *T. harzianum* complex was equally prevalent, exhibiting high metabolic and morphological variability that may explain the wide distribution of this species aggregate over different habitats (Kubicek et al., 2003). Sadfi-Zouaoui et al., 2009, in a study encompassing four different bioclimatic zones in Tunisia, assessed the genetic diversity of endemic species of *Trichoderma* and their association to bioclimatic zones. *T. harzianum*, divided into six clades, was the prevalent species complex. *T. harzianum* and *T. longibrachiatum* were predominant in forest soils in north Tunisia; *T. harzianum, T. saturnisporum* and *Trichoderma* sp. indet. were isolated from forest soils in central Tunisia; *T. atroviride* and *T. hamatum* were found in cultivated fields in northeast Tunisia; and *T. harzianum* and *T. hamatum* were present in oasis soils in south Tunisia. Zhang et al. (2005) assessed the biodiversity and biogeography of *Trichoderma* in China, sampling four disparate regions: north (Hebei province), south-east (Zhejiang province), west (Himalayan, Tibet) and south-west (Yunnan province). *T. asperellum, T. koningii, T. atroviride, T. viride, T. velutinum, T. cerinum, T. virens, T. harzianum, T. sinensis, T. citrinoviride,* and *T. longibrachiatum* were identified along with two putative new species. This study revealed a north-south gradient in species distribution in eastern Asia. Tsurumi et al. (2010) explored the biodiversity of *Trichoderma* in Mongolia, Japan, Vietnam and Indonesia, isolating 332 strains and finding T*. harzianum, T. hamatum, T. virens* and *T. crassum* in most habitats. *T. koningiopsis, T. atroviridae* and *T. stramineum* also were frequently isolated, except in cool sites where they were replaced by *T. polysporum* and *T. viridescens.* In tropical areas *T. ghanense, T.brevicompactum* and *T. erinaceum* were prevalent. In addition they discovered five unidentifiable isolate groups and 26 singular unidentified strains. The most comprehensive survey over any one biogeographic region was performed by Jaklitsch (2009, 2011). He employed three genetic markers to identify 620 specimens of *Hypocrea* occurring in 14 countries in temperate regions of Europe, identifying 75 species including 29 previously undiscovered, thus greatly expanding the number of species known in that region. His observations suggest that the biodiversity of *Hypocrea/Trichoderma* above soil exceeds the diversity residing in soil. He also speculated that the majority of species may be nectrotrophic on other fungi colonizing wood and bark. It now appears that the majority of *Trichoderma* species are capable of sexual recombination and form a teleomorph,

and a comparatively smaller number may be clonally propagating agamospecies.

Sardinia, which may or may not involve the displacement of native strains.

**4. Diversity of** *Trichoderma* **in tropical America** 

As a result of these recent discoveries, generalizations on the distribution of *Trichoderma*  have become increasingly problematic. Their occurrence will be modulated by microclimatic components, substrate availability, rhizosphere associations, soil chemistry, complex ecological interactions and many other factors. The introduction of invasive species, biocontrol agents, and agricultural perturbations result in changes in specific patterns of distribution that cannot be clearly identified, as suggested by Migheli et al. (2009) in finding the colonization of pan-European pan-global *Hypocrea/Trichoderma* species on the island of

Comparatively few comprehensive studies have been undertaken to assess the diversity of *Trichoderma* in neotropical regions. Since agriculture is a vital segment of local economies in the neotropics, most research in this region on *Trichoderma* has been directed to their biological control activities against phytopathogens. Studies have focused on biological control of plant pathogens with economical importance in cacao plantations, orchards, coffee, beans, cotton, flowers and rubber tree plantations, (Castro, 1996, Carsolio *et al*., 1994, Hebbar *et al*., 1999; Hoyos-Carvajal et al., 2008; Rivas & Pavone, 2010; Samuels *et al*, 2000, Samuels *et al.,* 2006, ), to control the symbiotic fungus of the leaf-cutting ant *Atta cephalotes* (López & Orduz, 2003), as well as to study the ability of *Trichoderma* to improve plant vigour and stimulate crop growth (Bae et al., 2009, Hoyos-Carvajal et al., 2009b, Resende et al., 2004).

Our knowledge of the distribution of *Trichoderma* species is constantly evolving in the context of current advances toward resolving the taxonomy of the genus. As a consequence, we can anticipate in future years to better understand the biogeography of *Trichoderma* species as research is pursued in new regions and to resolve complex species aggregates. For example, Samuels et al. (2006) determined that the species commonly cited in literature, *Trichoderma koningii,* in the strict sense is a relatively uncommon species restricted to temperate Europe and North America. From within the *T. koningii* aggregate he erected numerous new species, describing *T. caribbaeum* var*. aequatoriale, T. koningiopsis,* and *T. ovalisporum* as endophytes of *Theobroma* species in tropical America, and *T. ovalisporum* also from the woody liana *Banisteropsis caapi* in Ecuador. *T. koningiopsis* (previously identified as *T. koningii*) was determined to be common in tropical America, occurring also on natural substrata in East Africa, Europe and Canada, from ascospores in eastern North America, and as an endophyte in *Theobroma. T. stilbohypoxyli*, described as a parasite of *Stilbohypoxylon* species in Puerto Rico, was found to be more common in the tropics. Samuels et al. (1998) reported on the diversity of *Trichoderma* section *Longibrachiatum,* revealing diversity in neotropical areas resulting in the description of new species in this section. Jaklitsch et al. (2006), in revising the *T. viride* species complex, reported *T. viridescens* as a species found in Peru at high elevation, and *T. neokoningii* in a tropical region in Peru. He also described, as new species, *T. scalesiae* isolated as an endophyte from the trunk of daisy tree (*Scalesia pedunculata*) in the Galapagos Islands of Ecuador, *T. paucisporum* as a mycoparasite of *Moniliophthora roreri* on pods of *Theobroma cacao* in Ecuador, and *T. gamsii*, an apparently cosmopolitan species that has been found in Italy, Rwanda, South Africa, and Romania as well as Guatemala. Recent studies undertaken to find biocontrol agents in specific crops such as cocoa also has resulted in the determination of other new species in neotropical regions (Jaklitsch et al., 2006, Samuels et al., 2000, Samuels et al., 2006).

### **4.1 Can we generalize on the soil-inhabiting species of** *Trichoderma* **occurring in the neotropics?**

Hoyos-Carvajal et al. (2009a) carry out a systematic survey of *Trichoderma* species in seven countries: Mexico, Guatemala, Panama, Peru, Ecuador, Brazil and Colombia, isolating primarily from soil and employing complementary observations on morphology, metabolism and sequences of ITS 1 and 2 and the 5' region of *tef*-1a encompassing four introns. They identified 182 *Trichoderma* isolates finding a wide diversity of species over this region of the neotropics - *T. asperellum* (26 isolates), *T. asperelloides* (34 isolates, as *T. asperellum* 'B') *T. atroviride* (3), *T. brevicompactum* (5), *T. crassum* (3), *T. erinaceum* (3), *T. gamsii* (2), *T. hamatum* (2), *T. harzianum* (49), *T. koningiopsis* (6), *T. longibrachiatum* (3), *T. ovalisporum* (1), *T. pubescens* (2), *T. rossicum* (4), *T. spirale* (1), *T. tomentosum* (3), *T. virens* (8), *T. viridescens* (7), *T. parareesei* (3, as *H. jecorina*), along with 11 presumptive new species that have not yet been described. Analyses of variance were performed on metabolic data for the Colombian isolates. Highly significant differences (P < 0.0001) in assimilation were observed for 42 substrates among the 12 species isolated from Colombia (*T. asperellum*, *T. atroviride*, *T.* 

Biodiversity of *Trichoderma* in Neotropics 311

as glucose and N-acetyl-D-glucosamine were essentially the same. *T. asperelloides* had faster growth only on the disaccharide gentobiose. All differences between the two species were a matter of rate of growth, rather than growth/no growth. Notably, *T. asperellum* had significantly higher growth on substrates that are usually not at all assimilated by fungi, such as D-psicose, sedoheptulosan and γ-hydroxybutyric acid. *T. asperellum* was isolated from soils with poorly incorporated litter or crop residuals in colder climate zones (15ºC), growing on a wider variety of poorly metabolized substrates, and concomitantly under more difficult nutritional conditions in forest soils or debris. The pattern of growth on recalcitrant substrates for *T. asperellum* may be correlated with occurrence in relatively undisturbed, forested soils or other natural habitats. *T. asperelloides* was associated with agricultural soils crops, and in Colombia displayed a pattern of affinity for readily available substrates such as sugars, from comparatively warmer climates (23–28 ºC), and the rhizosphere soils where this strain was collected are associated with tropical crops such as African palm (*Elaeis guineensis*), coffee (*Coffea arabiga*), black mulberry (*Morus* sp.), avocado (*Persea americana*) and some grasses, in areas with a comparatively high organic matter contents. Despite the apparent different habitat preferences, *T. asperellum* and *T. asperelloides* did not exhibit differences in growth rates over the range 5-40°C, both species with

In the neotropics, the second most prevalent species from neotropical soils studied by Hoyos-Carvajal et al. (2009a) was *T. harzianum*, commonly associated with the rhizosphere of cultivated plants and frequently employed as a biocontrol agent against soil-borne phytopathogens. The predominance of *T. harzianum* in many different environments might be explained by its ability to assimilate a comparatively wide array of carbon substrates. The concept of *T. harzianum* as a genetically variable complex, comprising reproductively isolated biological species, recent agamospecies and phylogenetically unresolved relict lineages as determined by Druzhinina et al. (2010) is coherent with the adaptive range of this taxon. In the study of neotropical *Trichoderma* by Hoyos-Carvajal et al. (2009a), *T. harzianum* A was characterized by growth on poorly metabolized substrates, and strains from Colombia were isolated from a variety of environments, but commonly Andean soils associated with *Impatiens* sp. Group A also included strains from Mexico, consistent with the distribution found by Chaverri et al. (2003) for *H. lixii* lineage 1, and also includes isolates from Panama and Peru. The second clade, *T. harzianum* B, comprised mostly strains from the rhizosphere of tobacco, and sequences are coincident with lineage 5 of Chaverri et al. (2003) that included strains from Japan, Mexico and Cameroon, and with lineage 6 from Europe. *T. harzianum* C comprised nine strains from Colombia together with lineage 3 identified by Chaverri et al. (2003) from the United States, lineage 7 from Japan and Mexico, lineage 2 from Europe, and lineage 4 from Cameroon. *T. harzianum* A had fastest growth on L serine, i-erythriol, glycerol, D-sorbitol, D-ribose, α-D-lactose, L-threonine, L-proline, D-mannitol, and L-sorbose; however, there was no significant difference among the three genotypes on glucose, and all three genotypes had similar linear growth rates in culture on PDA over the temperatures range 5-40ºC. Significantly higher growth rates for *T. harzianum* A on the monosaccharide polyols i-erythritol, D-sorbitol, and D-mannitol, and on the fatty acid glycerol could indicate the presence of dehydrogenases allowing an adaptation to relatively dry habitats. *T. harzianum* B was the only genotype able to assimilate the disaccharide sucrose, although it exhibited relatively poor growth on the disaccharide lactose which was

temperature optima near 28C.

readily assimilated by genotypes A and C.

*brevicompactum*, *T. erinaceum*, *T. hamatum*, *T. harzianum*, *T. koningiopsis*, *T. longibrachiatum*, *T. virens*, *T. viridescens*, *T. parareesei* and *Trichoderma* sp. 210). The highest growth rates were observed on 23 substrates for *T. viridescens* isolated from mostly from rhizosphere of *Impatiens*, for *T. asperellum* obtained from a broad range of substrates on five substrates, and for *T. harzianum* from varied habitats and *T. parareesei* from African palm on three substrates. Seven substrates for which *T. viridescens* had the fastest growth rate were substrates not or scarcely assimilated by any other species (sedoheptulosan, glucuronamide, 2-aminoethanol, D-lactic acid methyl ester, putrescine, L-alaninamide, γ-hydroxyphenylacetic acid), perhaps indicating an ability to grow on recalcitrant substrates, although a similar pattern has been observed in other studies, contrasting isolates from undisturbed forests habitats, capable of growing on recalcitrant substrates, with isolates from agricultural habitats (Bissett unpublished). *Trichoderma viridescens* and *T. harzianum* showed positive growth on the largest number of significant substrates (41 and 34 respectively), indicating possible adaptation to a relatively broad range of habitats or niches and reflected in their wide distibutions. Slowest growth rates were observed for *T. erinaceum* from maize rhizosphere on 15 substrates, and for *Trichoderma* sp. 210 from river sand on 11 substrates. *T. longibrachiatum* and *Trichoderma* sp. 210 in section Longibrachiatum, along with *T. erinaceum* assimilated the fewest substrates (19-25 substrates).

The considerable biodiversity of *Trichoderma* in neotropical regions was evident in the study by Hoyos-Carvajal et al. (2009a). Nineteen species were identified from 182 isolations, and eleven so far undescribed species were discovered from rainforest soils and other specific habitats such as river sand, humus and wood in Peru, Mexico, Guatemala and Colombia. In a study of *Trichoderma* in Venezuelan soils the most abundant species was *T. harzianum*, followed by *T. virens, T. brevicompactum, T. theobromicola, T. koningiopsis, T. ovalisporum, T. asperellum, T. pleurotum* and *T. koningiopsis* (Rivas & Pavone, 2010). These observations are added to new species of *Trichoderma* from neotropics described in recent years, mainly as endophytes in plants (Jaklitsch et al., 2006, Samuels et al., 2006), and are evidence of the significant biodiversity of *Trichoderma* in the tropical region (table 1).

Unlike the studies conducted by Kullning et al. (2000) and Kubicek et al. (2002), assessing *Trichoderma* biodiversity of specific geographic areas, where the most common species was *T. harzianum*, in contrast in neotropical areas studied by Hoyos-Carvajal et al. (2009a) was *T. asperellum* (33% of strains) and *T. harzianum* the second most common (27%). Genetic variation was evident for both species, and two (*T. asperellum*) or three (*T. harzianum*) distinct genotypes were evident in the analysis of *tef* sequences and metabolic profiles. *T. asperellum,* which is often isolated from tropical regions (Druzhinina et al., pers. comm.), could be divided into two groups (A and B), which more recently have been described as separate species, *T. asperellum* and *T. asperelloides* respectively (Samuels et al., 2009, 2010). *T. asperellum* includes isolates from Brazil, Peru and Colombia originating in soils with poorly degraded materials such as fallen leaves or crop residuals in colder climate zones. T. *asperelloides* includes strains collected in Colombia and Ecuador that exhibit a preference for soils and substrates with high organic content, and often adapted to the rhizosphere of crops in Andean zones. These two species could not be differentiated by morphological characters or by growth rates, suggesting the development of ecologically or geographically isolated lineages as has been reported for *T. harzianum* (Chaverri et al., 2003) and *T. koningii* (Samuels et al., 2006). Metabolic differences were apparent between the closely related species, with *T. asperellum* better able to assimilate a wider range of C-substrates including some organic acids, polyols and amino acids, although growth rates on readily assimilated substrates such

*brevicompactum*, *T. erinaceum*, *T. hamatum*, *T. harzianum*, *T. koningiopsis*, *T. longibrachiatum*, *T. virens*, *T. viridescens*, *T. parareesei* and *Trichoderma* sp. 210). The highest growth rates were observed on 23 substrates for *T. viridescens* isolated from mostly from rhizosphere of *Impatiens*, for *T. asperellum* obtained from a broad range of substrates on five substrates, and for *T. harzianum* from varied habitats and *T. parareesei* from African palm on three substrates. Seven substrates for which *T. viridescens* had the fastest growth rate were substrates not or scarcely assimilated by any other species (sedoheptulosan, glucuronamide, 2-aminoethanol, D-lactic acid methyl ester, putrescine, L-alaninamide, γ-hydroxyphenylacetic acid), perhaps indicating an ability to grow on recalcitrant substrates, although a similar pattern has been observed in other studies, contrasting isolates from undisturbed forests habitats, capable of growing on recalcitrant substrates, with isolates from agricultural habitats (Bissett unpublished). *Trichoderma viridescens* and *T. harzianum* showed positive growth on the largest number of significant substrates (41 and 34 respectively), indicating possible adaptation to a relatively broad range of habitats or niches and reflected in their wide distibutions. Slowest growth rates were observed for *T. erinaceum* from maize rhizosphere on 15 substrates, and for *Trichoderma* sp. 210 from river sand on 11 substrates. *T. longibrachiatum* and *Trichoderma* sp. 210 in section Longibrachiatum, along with *T. erinaceum*

The considerable biodiversity of *Trichoderma* in neotropical regions was evident in the study by Hoyos-Carvajal et al. (2009a). Nineteen species were identified from 182 isolations, and eleven so far undescribed species were discovered from rainforest soils and other specific habitats such as river sand, humus and wood in Peru, Mexico, Guatemala and Colombia. In a study of *Trichoderma* in Venezuelan soils the most abundant species was *T. harzianum*, followed by *T. virens, T. brevicompactum, T. theobromicola, T. koningiopsis, T. ovalisporum, T. asperellum, T. pleurotum* and *T. koningiopsis* (Rivas & Pavone, 2010). These observations are added to new species of *Trichoderma* from neotropics described in recent years, mainly as endophytes in plants (Jaklitsch et al., 2006, Samuels et al., 2006), and are evidence of the

Unlike the studies conducted by Kullning et al. (2000) and Kubicek et al. (2002), assessing *Trichoderma* biodiversity of specific geographic areas, where the most common species was *T. harzianum*, in contrast in neotropical areas studied by Hoyos-Carvajal et al. (2009a) was *T. asperellum* (33% of strains) and *T. harzianum* the second most common (27%). Genetic variation was evident for both species, and two (*T. asperellum*) or three (*T. harzianum*) distinct genotypes were evident in the analysis of *tef* sequences and metabolic profiles. *T. asperellum,* which is often isolated from tropical regions (Druzhinina et al., pers. comm.), could be divided into two groups (A and B), which more recently have been described as separate species, *T. asperellum* and *T. asperelloides* respectively (Samuels et al., 2009, 2010). *T. asperellum* includes isolates from Brazil, Peru and Colombia originating in soils with poorly degraded materials such as fallen leaves or crop residuals in colder climate zones. T. *asperelloides* includes strains collected in Colombia and Ecuador that exhibit a preference for soils and substrates with high organic content, and often adapted to the rhizosphere of crops in Andean zones. These two species could not be differentiated by morphological characters or by growth rates, suggesting the development of ecologically or geographically isolated lineages as has been reported for *T. harzianum* (Chaverri et al., 2003) and *T. koningii* (Samuels et al., 2006). Metabolic differences were apparent between the closely related species, with *T. asperellum* better able to assimilate a wider range of C-substrates including some organic acids, polyols and amino acids, although growth rates on readily assimilated substrates such

assimilated the fewest substrates (19-25 substrates).

significant biodiversity of *Trichoderma* in the tropical region (table 1).

as glucose and N-acetyl-D-glucosamine were essentially the same. *T. asperelloides* had faster growth only on the disaccharide gentobiose. All differences between the two species were a matter of rate of growth, rather than growth/no growth. Notably, *T. asperellum* had significantly higher growth on substrates that are usually not at all assimilated by fungi, such as D-psicose, sedoheptulosan and γ-hydroxybutyric acid. *T. asperellum* was isolated from soils with poorly incorporated litter or crop residuals in colder climate zones (15ºC), growing on a wider variety of poorly metabolized substrates, and concomitantly under more difficult nutritional conditions in forest soils or debris. The pattern of growth on recalcitrant substrates for *T. asperellum* may be correlated with occurrence in relatively undisturbed, forested soils or other natural habitats. *T. asperelloides* was associated with agricultural soils crops, and in Colombia displayed a pattern of affinity for readily available substrates such as sugars, from comparatively warmer climates (23–28 ºC), and the rhizosphere soils where this strain was collected are associated with tropical crops such as African palm (*Elaeis guineensis*), coffee (*Coffea arabiga*), black mulberry (*Morus* sp.), avocado (*Persea americana*) and some grasses, in areas with a comparatively high organic matter contents. Despite the apparent different habitat preferences, *T. asperellum* and *T. asperelloides* did not exhibit differences in growth rates over the range 5-40°C, both species with temperature optima near 28C.

In the neotropics, the second most prevalent species from neotropical soils studied by Hoyos-Carvajal et al. (2009a) was *T. harzianum*, commonly associated with the rhizosphere of cultivated plants and frequently employed as a biocontrol agent against soil-borne phytopathogens. The predominance of *T. harzianum* in many different environments might be explained by its ability to assimilate a comparatively wide array of carbon substrates. The concept of *T. harzianum* as a genetically variable complex, comprising reproductively isolated biological species, recent agamospecies and phylogenetically unresolved relict lineages as determined by Druzhinina et al. (2010) is coherent with the adaptive range of this taxon. In the study of neotropical *Trichoderma* by Hoyos-Carvajal et al. (2009a), *T. harzianum* A was characterized by growth on poorly metabolized substrates, and strains from Colombia were isolated from a variety of environments, but commonly Andean soils associated with *Impatiens* sp. Group A also included strains from Mexico, consistent with the distribution found by Chaverri et al. (2003) for *H. lixii* lineage 1, and also includes isolates from Panama and Peru. The second clade, *T. harzianum* B, comprised mostly strains from the rhizosphere of tobacco, and sequences are coincident with lineage 5 of Chaverri et al. (2003) that included strains from Japan, Mexico and Cameroon, and with lineage 6 from Europe. *T. harzianum* C comprised nine strains from Colombia together with lineage 3 identified by Chaverri et al. (2003) from the United States, lineage 7 from Japan and Mexico, lineage 2 from Europe, and lineage 4 from Cameroon. *T. harzianum* A had fastest growth on L serine, i-erythriol, glycerol, D-sorbitol, D-ribose, α-D-lactose, L-threonine, L-proline, D-mannitol, and L-sorbose; however, there was no significant difference among the three genotypes on glucose, and all three genotypes had similar linear growth rates in culture on PDA over the temperatures range 5-40ºC. Significantly higher growth rates for *T. harzianum* A on the monosaccharide polyols i-erythritol, D-sorbitol, and D-mannitol, and on the fatty acid glycerol could indicate the presence of dehydrogenases allowing an adaptation to relatively dry habitats. *T. harzianum* B was the only genotype able to assimilate the disaccharide sucrose, although it exhibited relatively poor growth on the disaccharide lactose which was readily assimilated by genotypes A and C.

Biodiversity of *Trichoderma* in Neotropics 313

were isolated from other neotropical countries, notably Peru (6 new species) and Guatemala (3 new species) from which there were far fewer isolates. However, sampling in these countries was selective for unusual substrates above ground, resulting in the high proportion of novel strains. Therefore, the study by Hojos-Carvajal et al. (2009) would not account for the above ground biodiversity of species in Colombia on account of a relatively selective (but typical) sampling regime, although it is indicative of the wide distribution of *T. asperellum* and *T. harzianun* in soils, as reported in previous studies for other regions

The various studies of *Trichoderma* in the neotropics have expanded the known biogeographical and ecological distribution of many *Trichoderma* species. For example, *T. virens* (rain forest in Perú; rotten wood, rhizosphere of rice, tobacco and grassland in Colombia), *T. pubescens* (rain forest soil in Perú*), T. strigosum* (Perú rain forest soil), and *T. tomentosum* (cloud forest soil, Guatemala), were originally described from North America and Europe where they are relatively uncommon (Bissett, 1991 b). *T. ovalisporum*, previously reported from Ecuador as an endophyte in *Banisteriopsis caapi* and *Theobroma* sp. (Samuels et al., 2006), was isolated as an apparent saprophyte from soil in Panama. The infrequent isolation of these species also from neotropical soils suggests that these species may be restricted to specific ecozones, habitats or niches (Hoyos-Carvajal et al., 2009a). Samuels et al. (1998) reported *H. jecorina* (anam.: *T. reesei*) to be common in the pantropical region, and it is an important producer of cellulase enzymes. Hoyos-Carvajal et al., 2009a reported the species in typically warm soils cultivated with African palm in Colombia, but these strains did not assimilate sucrose, which had been reported for isolations of *H. jecorina* from the eastern Pacific (Samuels et al., 1998). We now know that the species reported by Hoyos-Carvajal et al. (2009) was in fact *T. parareesei,* recently differentiated from the sympatric

Eleven neotropical clades were differentiated from known *Trichoderma* species by Hoyos-Carvajal et al. (2009a) based on morphologic, metabolic and molecular differences and these remained undescribed. These are presumed to represent new taxa in *Trichoderma* and are the subject of ongoing investigations. The high proportion of apparently new species in this study is an indication that we have only begun to explore the biodiversity of *Trichoderma* in

*Trichoderma* species represent a major component of soil biodiversity with an important role in maintaining soil and plant health. The numbers, diversity, roles, and interactions of *Trichoderma* species in the environment are only now being discovered as tools are developed to distinguish the anamorph forms most commonly encountered. Significant and novel biodiversity of *Trichoderma* in the neotropics has been demonstrated, although we have only begun to explore the diversity of regions, habitats and substrates that exist in the region. We are now able to account taxonomically for a significant component of their biological diversity, to begin to predict biological activities, and to communicate results through the use of accurately determined names. The identification of *Trichoderma* species, as for species in other economically important and species-rich genera, is increasingly reliant on molecular data as the limits of phenotypic characters to distinguish species are reached. Many new species of *Trichoderma* will undoubtedly be distinguished as molecular tools are developed for ecological and metagenomic studies. Agriculture is the main economic

(Kullnig et al., 2000, Kubicek et al., 2003, Chaverry et al., 2003).

species *H. jecorina* by Druzhinina et al. (2010a).

the neotropic regions.

**5. Conclusions** 

In the study carried out by Hoyos-Carvajal et al. (2009a), Colombia becomes the most intensively surveyed neotropic region for *Trichoderma* biodiversity to date, comprising 116 isolates, representing 11 described species and one new taxon. As was mentioned, the most commonly isolated species from Colombia were *T. asperellum* (inclusive of more recently distinguished *T. asperelloides)* and *T. harzianum*. The prevalence of these species in Colombia may be explained by their genetic variability, seen in the several distinct genotypes found for each species, and their corresponding ability to grow on a wide variety of carbon substrates. However, the isolation methods used in this study, which are commonly used to isolate from the soil and rhizosphere, would be selective for soil-inhabiting species such as *T. asperellum* and *T. harzianum,* which are fast growing and sporulate early, allowing them to be recognized ahead of slower developing species. The majority of new species in this study


Table 1. *Trichoderma* species currently identified from the tropics with references to morphological descriptions*.*

In the study carried out by Hoyos-Carvajal et al. (2009a), Colombia becomes the most intensively surveyed neotropic region for *Trichoderma* biodiversity to date, comprising 116 isolates, representing 11 described species and one new taxon. As was mentioned, the most commonly isolated species from Colombia were *T. asperellum* (inclusive of more recently distinguished *T. asperelloides)* and *T. harzianum*. The prevalence of these species in Colombia may be explained by their genetic variability, seen in the several distinct genotypes found for each species, and their corresponding ability to grow on a wide variety of carbon substrates. However, the isolation methods used in this study, which are commonly used to isolate from the soil and rhizosphere, would be selective for soil-inhabiting species such as *T. asperellum* and *T. harzianum,* which are fast growing and sporulate early, allowing them to be recognized ahead of slower developing species. The majority of new species in this study

*T. harzianum* Chaverry *et al.*, 2003; Gams & Bissett, 1998

**Species Reference** 

*T. asperellum* Samuels *et al.*, 1999 *T. asperelloides* Samuels *et al.*, 2010 *T. atroviride* Bissett, 1992. *T. brevicompactum* Krauss *et al.*, 2004. *T. caribbaeum* Samuels *et al.* 2006 *T. caribbaeum var. aequatoriale* Samuels *et al.* 2006 *T. crassum* Bissett, 1991 a *T. erinaceum* Bissett *et al.*, 2003 *T. evansii* Samuels & Ismaiel, 2009 *T. gamsii* Jaklitsch *et al.*, 2006 *T. hamatum* Gams y Bissett, 1998

*T. koningiopsis* Samuels *et al.*, 2006 *T. lieckfeldtiae* Samuels & Ismaiel, 2009

*T. pleurotum* Komoń-Zelazowska et al., 2007

Table 1. *Trichoderma* species currently identified from the tropics with references to

*T. reesei (H. jecorina)* Gams and Bissett, 1998 *T. rossicum* Bissett *et al.*, 2003. *T. scalesiae* Jaklitsch *et al.*, 2006 *T. spirale* Bissett 1991a *T. stilbohypoxyli* Samuels *et al.* 2006a *T. theobromicola* Samuels *et al.* 2006b *T. tomentosum* Bissett 1991a *T. virens* Bissett 1991ª *T. viridescens* Jaklitsch *et al.*, 2006

*T. longibrachiatum* Gams y Bissett *T. neokoningii* Jaklitsch *et al.*, 2006 *T. ovalisporum* Holmes *et al.*, 2004 *T. parareesei* Atanasova *et al.*, 2010 *T. paucisporum* Samuels *et al.* 2006b

*T. pubescens* Bissett 1991a

morphological descriptions*.*

were isolated from other neotropical countries, notably Peru (6 new species) and Guatemala (3 new species) from which there were far fewer isolates. However, sampling in these countries was selective for unusual substrates above ground, resulting in the high proportion of novel strains. Therefore, the study by Hojos-Carvajal et al. (2009) would not account for the above ground biodiversity of species in Colombia on account of a relatively selective (but typical) sampling regime, although it is indicative of the wide distribution of *T. asperellum* and *T. harzianun* in soils, as reported in previous studies for other regions (Kullnig et al., 2000, Kubicek et al., 2003, Chaverry et al., 2003).

The various studies of *Trichoderma* in the neotropics have expanded the known biogeographical and ecological distribution of many *Trichoderma* species. For example, *T. virens* (rain forest in Perú; rotten wood, rhizosphere of rice, tobacco and grassland in Colombia), *T. pubescens* (rain forest soil in Perú*), T. strigosum* (Perú rain forest soil), and *T. tomentosum* (cloud forest soil, Guatemala), were originally described from North America and Europe where they are relatively uncommon (Bissett, 1991 b). *T. ovalisporum*, previously reported from Ecuador as an endophyte in *Banisteriopsis caapi* and *Theobroma* sp. (Samuels et al., 2006), was isolated as an apparent saprophyte from soil in Panama. The infrequent isolation of these species also from neotropical soils suggests that these species may be restricted to specific ecozones, habitats or niches (Hoyos-Carvajal et al., 2009a). Samuels et al. (1998) reported *H. jecorina* (anam.: *T. reesei*) to be common in the pantropical region, and it is an important producer of cellulase enzymes. Hoyos-Carvajal et al., 2009a reported the species in typically warm soils cultivated with African palm in Colombia, but these strains did not assimilate sucrose, which had been reported for isolations of *H. jecorina* from the eastern Pacific (Samuels et al., 1998). We now know that the species reported by Hoyos-Carvajal et al. (2009) was in fact *T. parareesei,* recently differentiated from the sympatric species *H. jecorina* by Druzhinina et al. (2010a).

Eleven neotropical clades were differentiated from known *Trichoderma* species by Hoyos-Carvajal et al. (2009a) based on morphologic, metabolic and molecular differences and these remained undescribed. These are presumed to represent new taxa in *Trichoderma* and are the subject of ongoing investigations. The high proportion of apparently new species in this study is an indication that we have only begun to explore the biodiversity of *Trichoderma* in the neotropic regions.
