Biomolecules Produced by Trichoderma Species as Eco-Friendly Alternative Suppressing Phytopathogens and Biofertilizer Enhancing Plant Growth

Abstract

Olive (Olea europeae L.) is one of the most important fruit trees of the Mediterranean regions. Biotic factors such as phytopathogenic diseases have a significant negative impact on olive productivity in the Mediterranean Basin including Algeria. Currently, phytopathogens management is focus mainly on the use of chemical pesticides which is not recommended because it leads to environmental pollution, development of chemical resistance, and its low cost-efficiency. Eco-friendly methods and alternative disease control measures such as the use of biocontrol agents and biofertilizer should be opted as alternatives to the use of synthetic chemicals. Trichoderma species associated with olive roots are known for their ability to produce antimicrobial compounds, such as antibiotics, volatile organic compounds and lytic enzymes that restrict phytopathogenic strain growth. Besides, they are considered as plant growth promoting fungi (PGPF). This genus colonize the root systems of plants and promote their growth; it can increase nutrient availability and uptake in plants by fixing nitrogen, solubilizing phosphorus, producing several biomolecules and phytohormones. Moreover, it helps plants tolerate environmental stresses such as drought, salinity and diseases. In this work, we review pionnering and recent developments on several important biomolecules and functions that Trichoderma species isolated from olive rhizosphere soil exhibit to enhance plant growth and control phytopathogen diseases. Therefore, the use of highly competitive strains in open field in order to obtain consistent and better results in agricultural production activities.

1. Introduction

Olive is affected by a wide range of biotic constraints such as soil-borne diseases which can cause significant damage and economic losses. Currently, plants diseases are managed mainly through the use of chemical pesticides, which can generate negative effects, such as health problems, loss of ecological diversity, and the bioaccumulation of toxic substances [1]. Nowadays, a key practice to deal with plant pathogens in sustainable agriculture is the biological control, which is based on managing natural resources and developing antagonistic activities against harmful microorganisms [2] which make it an effective and eco-friendly approach against plant diseases [3]. Many microorganisms with antagonistic activity such as Trichoderma spp. offer an environment-friendly alternative to get out of chemical pesticides damages [4, 5, 6, 7]. Trichoderma spp. are known as promising fungal for the management of plant diseases, especially against soil-borne pathogens [8]. Therefore, Trichoderma spp. are most investigated and employed as biopesticide [9, 10, 11]. This genus has antagonistic activities and can act by various mechanisms against a wide range of soil-borne phytopathogenic fungi including competition for nutrients and the systemic activation of plant defense responses [12, 13, 14]. Thus, Trichoderma spp. are used as biopesticides in management of plant diseases worldwide [15]. Furthermore, they act by different modes of action against plant pathogens, including, mycoparasitism through the production of the cell wall degrading enzymes such as chitinases, glucanases, and proteases [16, 17], production of antibiotics and volatile organic compounds (VOCs) [18]. Trichoderma spp. produce wide spectrum of VOCs which are part from several chemical groups such as monoterpenes, sesquiterpenes, alcohols, aldehydes, aromatic compounds, esters, furans, hydrocarbons, ketones, and compounds containing S and N elements [19, 20]. These volatile compounds can diffuse through pores in the soil, move long distance, and affect pathogen without direct contact [21], which makes it more efficient at microbial interactions compared to non-volatile compounds [22]. Hence, it could be responsible for several biological processes such as biocontrol or communication between microorganisms [23]. Importantly, Trichoderma spp. are considered as plant growth-promoting fungi (PGPF) which can colonize and proliferate within the rhizosphere environment and enhancers of plant defense mechanisms. They display stimulation of plant growth because of their capacity to produce plant growth promoters [24, 25, 26]. Trichoderma species are able to promote plant growth through various mechanisms such as solubilizing insoluble phosphate, production of siderophore and plant hormone such as indole-3-acetic acid (IAA) [27].

2. Characteristics of Trichoderma spp.

Trichoderma is a genus of a heterogeneous group of fungal species. They are considered as anamorphic Hypocreales [28]. Trichoderma species are free-living and/or endophytic fungi that grow vigorously in soil and plant root ecosystems, they are known as ubiquitous saprophytic fungi [12, 29, 30, 31] as well as aboveground such as on rotting wood and other organic materials [17, 32, 33, 34, 35]. Further, Trichoderma strains produce a few pigments, ranging from a greenish-yellow up to a reddish tinge and sometimes colorless strains might likewise be available. The conidia can have different hues, going from drab to various tints of green or dim or earthy colored hints [28]. Microscopic identification criteria of Trichoderma are as follows: septate and translucent hyphae; conidiophores are short, translucent, branched often giving the pyramidal appearance, not verticillate, the phialides are attached at right angles to the conidiophores. Spores produced are conidia which are translucent, ovoid in shape, borne in small terminal clusters at the tips of phialides, some species can produce globose chlamydospores, which are intercalary or terminal. These chlamydospores are usually unicellular but can be multicellular for some species such as Trichoderma stromaticum [36].

3. Trichoderma spp. with biocontrol potentials against plant diseases

In recent years, Trichoderma species are considered as good alternatives for substituting chemical usages. They act by various mechanisms against a wide range of phytopathogenic fungi. Biocontrol by Trichoderma including mycoparasitism, antibiosis as well as competition for nutrients and space with plant pathogens [29, 32, 37, 38, 39]. Furthermore, their interaction with root induces the plant’s resistance to pathogens, and destruction of the root-knot nematode at the different stages of its growth phases [40].

3.1 Volatile organic compounds

Volatile organic compounds (VOCs) are low-molecular-weight molecules that have contain fewer than 12 carbon atoms, and may be associated with other elements such as nitrogen, sulfur, bromine, oxygen, fluorine, and chlorine [41]. These compounds exhibit antimicrobial activities, promote plant development, the induction of systemic resistance, and considered as chemical signaling in plants [42, 43]. Most of volatile compounds produced by Trichoderma species exerted antifungal activity against plant pathogens. It has been reported that [44] were identified more than 278 volatile compounds from liquid cultures of Trichoderma harzianum using GC–MS; these compounds are part of different chemical groups such as normal saturated hydrocarbons (C7–C30), cyclohexane, fatty acids, alcohols, cyclo-pentane, esters, sulfur-containing compounds, simple pyrane, and benzene derivatives. Similar compounds were detected by [45], suggesting that Trichoderma spp. strains isolated from the rhizosphere of healthy olive trees have antagonistic activity against plant pathogen by production of VOCs. Among compounds detected in this study, eicosane which has antifungal activity [46, 47]. On the other hand, several volatile compounds with antifungal activities were detected by [45] such as benzeneethanol, toluene, alcohols, phenols, cyclohexane. Palmitic acid, alkanes, octadecenoic acid, palmitic acid, and limonene. Many works revealed the antifungal effect of these compounds such as the finding of [48] who reported that Fatty acids (e.g., palmitic acid and octadecenoic acid) are known to possess antibacterial and antifungal activities. Furthermore, [49] report the production of palmitic acid by Trichoderma virens and T. harzianum. Compounds such as methoxyacetic acid and benzene were also detected; these compounds have been demonstrated to exhibit antimicrobial activities [50]. Moreover, in a previous study, [51] reported that the terpenoid and limonene were the main components which were observed as effective biological control compounds. Moreover, limonene is considered as a mediator of plant growth that leads to a change in the concentration of chlorophyll and the size of plants. In similar studies, alkanes with antifungal activity were also detected such as cyclohexane and cyclopentane, other alkanes were identified such as dodecane which has a role as antifungal agent [52].

3.2 Production of antimicrobial compounds

Plants and microorganisms are in constant competition for nutrients, microorganisms produce various antimicrobial compounds as a strategy to compete with other microorganisms for establishment in an ecological niche [53]. These compounds have bactericidal or bacteriostatic effect. Antimicrobial compounds produced by Trichoderma species can act by various mechanisms against a wide range of soil-borne phytopathogenic fungi including the production of antibiotics and/or hydrolytic enzymes, as well as competition for nutrients [12, 13, 14].

3.2.1 Lytic enzymes

The cell walls of fungi contain chitin, cellulase, and glucan. Therefore, phytopathogenic fungi are affected by some lytic enzymes, including 1,3-glucanases, lipases, cellulases, and chitinases [54]. Trichoderma species have been widely recognized for the production of extracellular enzymes with mycoparasitic effect such as glucanases, cellulases, and chitinases. Mycelium lysis was observed in the confrontation zone between the pathogen Verticillium dahliae and Trichoderma species suggesting the ability of these isolates isolated from the rhizosphere of healthy olive to produce enzymes involved on cell wall degradation process and lysis of the mycelium during the mycoparasitic activity [45]. Moreover, [12] suggested that Trichoderma isolates are able to produce cell wall degrading enzymes such as cellulase, xylanase, pectinase, glucanase, lipase, amylase, arabinase, protease, and chitinases that are the most important lytic enzymes playing a key role in the degradation of cell walls of other plant pathogenic fungi (Figure 1) [12].

Figure 1.

The different action stages of Trichoderma against Rhizoctonia solani through mycoparasitism are as follows: (A) appressorium-like structures, (B) Trichoderma wrapping around the hyphae of R. solani, (C) an enlargement of the interaction between Trichoderma and Rhizoctonia in which appressorium-like structures are observed. The scale bar equals 10 μm. (D) The hypha of R. solani, from which the Trichoderma hypha has been removed, shows the pores caused by the mycoparasite at the junction points between the two hyphae. R: hyphae of R. solani. T: hyphae of Trichoderma spp. [12].

3.2.2 Antibiotics

Fungi are able to produce compounds with antibiotic properties. They have low molecular weight and can interfere with the development of various microorganisms through inhibition [28]. This inhibition action called antibiosis. It is another mechanism found in Trichoderma which can restrict phytopathogens growth. There are more than 180 secondary metabolites of Trichoderma that have been identified into various classes of compounds [55]. These compounds have different effects against pathogens. Some secondary metabolites affect plant metabolism and growth. For example T. viride, T. harzianum, and Trichoderma koningii, are capable in the production and secretion of a volatile compound, 6-pentyl-α-pyrone (6-PAP) which exhibit antifungal activity against several pathogenic species such as Botrytis cinerea, R. solani, and Fusarium oxysporum [28].

Antibiotics produced by Trichoderma species inhibit the growth of other microorganisms. Most Trichoderma strains produce metabolites with antibiotic properties that prevent colonization by antagonized microorganisms; among these metabolites, the production of harzianic acid, alamethicins, tricholin, peptaibols, antibiotics, 6-PAP, massoilactone, viridin, gliovirin, glisoprenins, heptelidic acid, and others have been described [56].

3.2.3 Competition

Competition between fungi and pathogens is another mechanism of biocontrol. Trichoderma can compete for nutrients and restrict the growth of pathogens especially when nutrients become a limiting factor. They can easily compete the rhizosphere of different plants and cause changes in plant metabolism. Moreover, Trichoderma may colonize also space around infection sites [57]. Through chelators production such as siderophores which increase the absorption and concentration of certain nutrients (copper, iron, phosphorus, manganese, and sodium). As a result, iron will be less available for the pathogen. For this reason, competition in the soil between microorganisms is considered an indirect control mechanism for pathogens [58]. Studies conducted by [59] showed the important role of siderophores produced by Trichoderma asperellum in antagonism against F. oxysporum. They also play a role in stimulating plant growth by reducing oxidative stress. Besides, studies conducted by [59] showed the role of siderophores produced by T. asperellum T34 in controlling F. oxysporum, with a reduction in tomato infestation and stimulation of root plant growth.

3.2.4 Siderophores and the acquisition of iron

Siderophores are low-molecular-weight secondary metabolites produced by a wide range of plant and fungal species such as Trichoderma spp. [60]. They have the ability to capture metal ions with a high affinity for Fe (III) than Fe (II). Depending on the functional group that acts as the sequestrant, they can be classified into catecholates, hydroximates, and hydroxycarboxylates [61]. In the rhizosphere, crops may obtain iron through microbially produced siderophores [62]. There are more than 500 biomolecules that are classified as siderophores [62]. Iron deficiency can lead to severe biological inhibition for organisms by depriving them of this element because it is essential in cellular processes such as DNA synthesis, respiration, and free-radical detoxification [63]. Siderophores produced by Trichoderma spp. demonstrate various functions in the rhizosphere. In addition to conferring an advantage to take iron into the rhizosphere, under limiting conditions, siderophores may also inhibit the growth of pathogens that could potentially cause damage to the plant [64]. Trichoderma spp. producing siderophores in rhizospheres can restrict iron and make it less available to pathogens, indirectly promoting plant growth [65]. Studies conducted by [59] showed the role of siderophores produced by T. asperellum T34 in controlling F. oxysporum, reducing tomato infestation and stimulating plant root growth.

4. Biomolecules enhancing plant growth

4.1 Production of phytohormones

Phytohormones play an important role in agriculture [66]; they are synthesized by many rhizosphere microorganisms including Trichoderma spp. They have various roles such as modification of the physiological functions of plants to accelerate their growth by intensive cell division in callus tissue, promotion of phloem development, enhance lateral root development, plant growth stimulation and prevention of leaf aging by slowing down the breakdown of chlorophyll pigments in plants as well as improving metabolism even at low concentrations [67, 68, 69]. IAA and gibberellins (GAs) are among the most important phytohormones that regulate the plant’s development and enhance plant growth through several processes [70, 71, 72, 73, 74].

4.2 Acquisition and nutrients solubilization

Various fungi such as Trichoderma spp. are associated to the plant roots’ rhizosphere, they provide nutrients, protection against biotic and abiotic stresses, and stimulate plant growth [75, 76]. Trichoderma species have the ability to acquire nutrients in the rhizosphere through various mechanisms.

4.2.1 Phosphate solubilization

Phosphorus is important elements for plant growth. It can be found in two forms: organic phosphorus and inorganic phosphorus, which usually forms insoluble mineral compounds with calcium, aluminum, or manganese [77, 78]. The distribution of these forms in soils is influenced by several factors such as microbial activity, pH, soil type, and organic matter availability [1]. Recently, phosphate solubilizing microorganisms have attracted the attention of agronomists; these microorganisms were used as soil inoculum to improve plant growth [79]. Plants and fungi including Trichoderma spp. compete for the limited available phosphorus through various processes, such as solubilization, precipitation, absorption, and desorption. Inorganic phosphate and organic phosphorus can be mineralized through enzymatic action [1]. Trichoderma species have the ability to solubilize insoluble phosphate into soluble phosphate [80, 81]. In previous studies; [82] reported that Trichoderma atroviride LBM 112 and T. stilbohypoxyli LBM 120 revealed positive results for phosphate solubilization with formation of halo-zone on the solid medium containing insoluble inorganic phosphorus source. In addition, T. harzianum T11 (OL587563) isolated from rhizosphere soil of olive trees has several plant growth-promoting traits, such as the phosphate-solubilizing ability and the production of siderophores [74].

4.2.2 Nitrification and nitrogen fixation

Nitrogen fixation processes have significant ecological importance in various ecosystems, including those of agricultural interest. Nitrogen plays a critical role in plant synthesis as it is a component of important biomolecules such as nucleic acids, peptides, organic acids, and fatty acids, which are necessary for the structure and activity of all organisms. Nitrogen-fixing by microorganisms play a key role on growth-promoting plant. It has been suggested that the promotion effect on plant growth might be mediated by providing nitrogen through biological nitrogen fixation and hormones [83, 84]. Production of ammonia and nitrogen-fixing ability by Trichoderma strains are reported in previous findings. Ahemad and Kibret [85] reported that ammonia is useful for plants as directly or indirectly. Ammonia production by the Trichoderma isolates may influence plant growth indirectly; ACC synthesized in plant tissues by ACC synthase is released from plant roots and taken up by neighboring micro-organisms. Then, Trichodrema may hydrolyze ACC (1-aminocyclopropane-1-carboxylic acid) to ammonia. Besides, [74] reported that production of ammonia by Trichoderma species isolated from rhizosphere soil of olive is sustained with the results obtained by [86] who reported that among 20 Trichoderma spp. isolated from chili rhizosphere, 13 isolates were able to produce ammonia (Figure 2; Tables 1 and 2).

Figure 2.

Schematic description of the main mechanisms used by Trichoderma spp. to competitively colonize the rhizosphere of host plants [74].

Chemical nature	Secondary metabolites	Trichoderma species	Bio-activity observed	Reference
Alcohol	2-Phenylethanol	T. harzianum	Reduces the growth of Aspergillus flavus and aflatoxin production	[87, 88]
Anthraquinone	Pachybasin	T. harzianum	Increases the number of coils of the biocontrol agent against R. solani	[89]
	Emodin	T. viride	Antimicrobial and antineoplasic agent	[90, 91, 92]
Azaphilone	T22azaphilone	T. harzianum	Inhibits the growth of R. solani, Pythium ultimum and Gaeuman nomyces graminis	[93]
Bisorbicillinoid	Bisvertinolone	T. longibrachiatum	Antifungal properties via inhibition of β- (1,6)-glucan biosynthesis	[94]
Butenolide	Dehydro derivative of harzianolide	T. harzianum	Antifungal activity against Gaeumannomy ces graminis var. tritici	[95]
Hydrolytic enzymes	Cellulases	T. reesei	Degrades cellulase during root colonization to penetrate the plant tissue	[96]
	β-1,6- Glucanases	Trichoderma sp.	Hydrolyses fungal pathogen cell walls of B. cinerea, R. solani, Phytophthora citrophthora	[17]
	Chitinases	Trichoderma sp.	Hydrolytic enzymes of the fungal cell wall	[97, 98]
Indolic compound	Indole-3- acetic acid (IAA)	T. atroviride, T. virens	Controls a number of growth and development processes in plants	[99]
	Indole-3- acetaldehyde	T. atroviride, T. virens	Controls root growth in Arabidopsis thaliana	[99]
	Indole-3-; carboxaldehyde	T. atroviride, T. virens	Induces adventitious; root formation in A. thaliana	[100]
Koninginins	Koninginins A–E	T. koningii; T. harzianum	Antifungal activity against F. oxysporum, Fusarium solani, and Alternaria panax	[101, 102]
Monoterpene	β-Myrcene	T. virens	Regulates the expression of genes	[22, 103]
Nitrogen heterocyclic compound	Harzianic acid	T. arundinaceum; T. harzianum	Antimicrobial metabolite, siderophore and plant growth regulator	[104, 105, 106]
	Harzianopyridone	T. harzianum	Antifungal activity against B. cinerea, R. solani and inhibitor of the protein phosphatase type 2A; (PP2A)	[107]
	Melanoxadin	T. sp. strain ATF-451	Inhibits melanin formation in the larval hemolymph of the silkworm, Bombyx; mori	[108]
Peptide	Trichokonin VI (Tk VI)	T. longibrachiatum	Inhibits primary root growth in A. thaliana	[109]
Pyrane	Koninginin A	T. koningii	Plant growth regulator	[110]
	Koninginin D	T. koningii	Alters pathogen fungal growth of R. solani, Phytophthora cinnamomi, Pythium middletonii, F. oxysporum and Bipola ris sorokiniana	[111]
Pyridones	Hharzianopyridone	T. harzianum	antifungal activity against plant pathogenic fungi, such as P. ultimum, G.; graminis var. tritici, R. solani, and B. cinerea	[112]
Pyrones	6-Pentyl-2H- pyran—2-one	Trichoderma viride; T. atroviride	Antifungal activity against R. solani, F.	[113]
Siderophore	Fusarinine C	Trichoderma sp.	Fe-chelated, can be available to plants	[114]
	Ferricrocin	T. atroviride^a, T. virens^a, T. reesei^a	Key metabolite in the competition for iron in the rhizophere	[115]
	Coprogen B	Trichoderma spp.	Solubilizes iron unavailable to the plant	[116]
Steroidal compound	Viridin	T. koningii, T. virens, T. viride	Antifungal metabolite that alter the spore germination; of Botrytis; allii, Colletotrichum; lini and Fusarium caeruleum	[55]

Table 1.

Secondary metabolites secreted by Trichoderma sp. and their bio-active role.

^a

http://genomebiology.com/2011/12/4/R40

Compound	Strain	Crops	Application mode	Beneficial outcome	References
Biofertilizer	Trichoderma azevedoi	Lettuce	Simple exposure	Increases carotenoids and chlorophyll with reduction in the white mold attack to about 78.83%	[117]
	Trichoderma afroharzianum	Tomato	Seed inoculation or treatment	Helps in the secretion of; Phytohormones like; homeostasis, antioxidant; activity, phenylpropanoid; biosynthesis and glutathione metabolism	[118]
	T. harzianum,; T.asperellum,; Trichoderma; hamatum,; T.atroviride	Chinese cabbage	Irrigation	Increases soil enzyme activity, yield by 37%,; and increases the concentration of; inorganic nitrogen and phosphorus content of the; soil	[119]
	Trichoderma brevicompactum,; Trichoderma; gamsii, T. harzianum	Tomato	Seedling drenching	Improved growth and yield; due to the production of; IAA	[35, 120]
	T. harzianum; T. asperellum	Tomato	Seed treatment	Improves phosphorus; uptake	[121]
	T. brevicompactm,; T. gamsii,; T. harzianum	Tomato	Seed drenching	Improves phosphorus; solubilization	[120]

Table 2.

Trichoderma sp. as bio-fertilizers and their role in promoting plant growth and yield.

5. Conclusion

This report reviews the importance of Trichoderma spp. as a biocontrol agent suppressing the growth of the fungal pathogens and as biofertilizer enhancing plant growth. Therefore, the increase use of Trichoderma spp.as commercial mycofungicides and biofertilizers offers promising prospects for sustainable and environmentally friendly agriculture. These eco-friendly alternatives can substitute the excessive use of chemical products that can cause problems in the long term. The biotechnological advances from these microorganisms such as fungi are immense and yet to be explored. Thus, more studies need to be explored to elucidate the development of sustainable biotechnological applications of the Trichoderma species on soil–plant system.

Conflict of interest

The authors declare no competing interests.