**6. Cross-talk by and response to secondary metabolites in mycoparasitic interactions**

Several environmental cues like temperature, light, carbon, nitrogen, pH and competing or synergistic organisms are known to influence the transcriptional regulation of secondary metabolism-associated gene clusters (**Figure 1**). Suboptimal environmental conditions thereby often facilitate and promote transcriptional activation or transcriptional reprogramming events [70]. In media containing chitin or *B. cinerea* cell walls, the predicted cutinase transcription factor 1 encoding gene of *T. harzianum* (*Thctf1*) was up-regulated. *Thctf1* deletion mutants exhibited reduced antagonistic and antifungal ability, and the mutant strain did not synthesize two 6-PP derivatives, indicating a role of *Thctf1* in secondary metabolism of *T. harzianum* [71]. Furthermore, the overexpression of the gene encoding multiprotein bridging factor 1 (*Thmbf1*) of *T. harzianum* – a transcriptional co-activator of *Thctf1* – negatively regulated

Like known for the production of mycotoxins in non-mycoparasitic species [73], secondary metabolite production in mycoparasitic fungi is governed by heterotrimeric G protein signaling and the associated cAMP-pathway, as well as mitogen-activated protein kinase (MAPK) cascades [74, 75]. *T. atroviride* mutants, lacking the MAPK-encoding gene *tmk1* showed an enhanced production of peptaibols and of 6-PP [74]. First evidence for a positive regulation of the secondary metabolism by cAMP signaling came from *T. virens* ∆*tac1* mutants bearing a deletion of the adenylate cyclase-encoding gene. The mutants were unable to offend *Sclerotium rolfsii* and *R. solani,* but showed a clear inhibition zone in direct confrontation with *Pythium* sp., pointing to a host-dependent expression of secondary metabolism-associated

**Figure 1.** Overview on mycoparasitism-influencing factors and pathways in secondary metabolite biosynthesis of

mycoparasitic fungi.

the antifungal abilities, as well as the expression of VOCs [72].

44 Secondary Metabolites - Sources and Applications

In bacteria, it has been shown that at sub-inhibitory concentrations antibiotics serve as mediators of microbial communication and interaction with one of the outcomes being the production of cryptic metabolites [81]. Accordingly, the interaction with other fungi may shape the secondary metabolite profile of a specific fungus, making co-cultures a valuable tool for eliciting the activation of silent secondary metabolism-associated gene clusters.

Studies on the mutual effects of secondary metabolites produced during mycoparasitic interactions are rare however. *Trichoderma*-derived 6-PP was shown to suppress the synthesis of the *Fusarium* mycotoxins fusaric acid and deoxynivalenol (DON) [82–85], suggesting that 6-PP acts as communication molecule that elicits biological responses in the interaction partners. On the other hand, fusaric acid and DON modulate 6-PP production as well as chitinase gene expression in *T. atroviride* and recent studies provided evidence that *Fusarium* mycotoxins induce defense mechanisms in mycoparasites such as *T. atroviride* and *C. rosea* which results in mycotoxin detoxification [59, 86]. *C. rosea* was shown to open the ring structure of zearalenone (ZEN), while *Trichoderma* spp. seem to convert ZEN into its reduced and sulfated forms and metabolize DON to deoxynivalenol-3-glucoside, a detoxification product of DON previously identified in plants [87, 88]. In the interaction of the mycoparasite *T. arundinaceum*

with *B. cinerea, Botrytis-*derived mycotoxins botrydial and botcinins attenuated trichothecene biosynthesis gene expression in *Trichoderma* while botrydial production was repressed by *Trichoderma-*derived harzianum A and aspinolide [89–91].

**References**

[1] Jeffries P. Biology and ecology of mycoparasitism. Canadian Journal of Botany.

Secondary Metabolites of Mycoparasitic Fungi http://dx.doi.org/10.5772/intechopen.75133 47

[2] Karlsson M, Atanasova L, Jensen DF, Zeilinger S. Necrotrophic mycoparasites and their genomes. Microbiology Spectrum. 2017;**5**:FUNK-0016-2016. DOI: 10.1128/microbiol-

[3] Renshaw JC, Robson GD, Trinci APJ, Wiebe MG, Livens FR, Collison D, Taylor RJ. Fungal siderophores: Structures, functions and applications. Mycological Research.

[4] Mukherjee PK, Horwitz BA, Kenerley CM. Secondary metabolism in *Trichoderma* – A genomic perspective. Microbiology. 2012;**158**:35-45. DOI: 10.1099/mic.0.053629-0

[5] O'Brien PA. Biological control of plant diseases. Australasian Plant Pathology.

[6] Fischer GJ, Keller NP. Production of cross-kingdom oxylipins by pathogenic fungi: An update on their role in development and pathogenicity. Journal of Microbiology.

[7] Vinale F, Sivasithamparam K, Ghisalberti EL, Marra R, Barbetti MJ, Li H, Woo SL, Lorito M. A novel role for *Trichoderma* secondary metabolites in the interactions with plants. Physiological and Molecular Plant Pathology. 2008;**72**:80-86. DOI: 10.1016/j.

[8] Viterbo A, Inbar J, Hadar Y, Chet I. Plant disease biocontrol and induced resistance via fungal Mycoparasites. In: Kubicek CP, Druzhinina IS, editors. Environmental and Microbial Relationships: The Mycota. Vol. 4. Berlin, Heidelberg: Springer; 2007. pp. 127-

[9] Schmoll M, Dattenböck C, Carreras-Villaseñor N, Mendoza-Mendoza A, Tisch D, Alemán MI, Baker SE, Brown C, Cervantes-Badillo MG, Cetz-Chel J, Cristobal-Mondragon GR, Delaye L, Esquivel-Naranjo EU, Frischmann A, Gallardo-Negrete JJ, García-Esquivel M, Gomez-Rodriguez EY, Greenwood DR, Hernández-Oñate M, Kruszewska JS, Lawry R, Mora-Montes HM, Muñoz-Centeno T, Nieto-Jacobo MF, Nogueira Lopez G, Olmedo-Monfil V, Osorio-Concepcion M, Piłsyk S, Pomraning KR, Rodriguez-Iglesias A, Rosales-Saavedra MT, Sánchez-Arreguín JA, Seidl-Seiboth V, Stewart A, Uresti-Rivera EE, Wang C-L, Wang T-F, Zeilinger S, Casas-Flores S, Herrera-Estrella A. The genomes of three uneven siblings: Footprints of the lifestyles of three *Trichoderma* species. Microbiology

and Molecular Biology Reviews. 2016;**80**:205-327. DOI: 10.1128/MMBR.00040-15 [10] Mukherjee PK, Horwitz BA, Herrera-Estrella A, Schmoll M, Kenerley CM. *Trichoderma* research in the genome era. Annual Review of Phytopathology. 2013;**51**:105-129. DOI:

[11] Quandt CA, Bushley KE, Spatafora JW. The genome of the truffle-parasite *Tolypocladium* ophioglossoides and the evolution of antifungal peptaibiotics. BMC Genomics.

1995;**73**(S1):1284-1290. DOI: 10.1139/b95-389

2002;**106**:1123-1142. DOI: 10.1017/S0953756202006548

2017;**46**:293-304. DOI: 10.1007/s13313-017-0481-4

2016;**54**:254-264. DOI: 10.1007/s12275-016-5620-z

146. DOI: 10.1007/978-3-540-71840-6\_8

10.1146/annurev-phyto-082712-102353

2015;**16**:553. DOI: 10.1186/s12864-015-1777-9

spec.FUNK-0016-2016

pmpp.2008.05.005

Co-culturing of mycoparasites with prey fungi simulates the conditions occurring during the mycoparasitic interaction in natural or agricultural systems and could hence encourage the production of secondary metabolites via communication and signaling molecules. Accordingly, pairwise interactions of *Aspergillus niger, Fusarium verticilliodes* and *C. rosea* led to metabolites that occurred in single cultures but were suppressed in dual cultures, and many new metabolites not present in single cultures were found in dual cultures [92]. Similar results were obtained in co-culturing experiments of *T. harzianum* and *Talaromyces pinophilus* with the accumulation of siderophores being induced in both interaction partners and the production of *Talaromyces*-derived 3-*O*-methylfunicone and herquline B being reduced. In addition, the novel substance harziaphilic acid was detected in the co-culture only [93].

Based on these studies, it is evident that secondary metabolites contribute to mycoparasitic interactions in various ways including inhibition of the activity or synthesis of mycoparasitism-relevant enzymes and other substances, by eliciting defense and detoxification responses or by triggering the production of cryptic metabolites. In most cases, however, information on the spatial distribution of the secreted substances is lacking and it is hence difficult to assign novel secondary metabolites specifically induced during the co-cultivation to its actual producer. Recently, mass spectrometry-based imaging (MSI) turned out as a valuable tool for in situ visualizing the dynamics and localization of small molecules released during microbial interactions [94]; however, reports on its application to mycoparasitic fungus-fungus interactions are still rare. By applying matrix-assisted laser desorption/ionization (MALDI)-based MSI for visualization and identification of secondary metabolites being exchanged during the mycoparasitic interaction of *T. atroviride* with *R. solani* [95], the diffusion of *Trichoderma*derived peptaibols toward the prey and the accumulation of *Rhizoctonia*-derived substances at the borders of fungal interaction was tracked. Monitoring of the *T. harzianum* interaction with the fungal phytopathogen of cacao plants *Moniliophthora roreri* by MSI lead to the detection of T39 butenolide, harzianolide, sorbicillinol and an unknown substance specifically produced in the co-culture with a spatial localization in the interaction and overgrowth zones [96].
