A soil fungus confers plant resistance against a phytophagous insect by disrupting the symbiotic role of its gut microbiota.

Plants generate energy flows through natural food webs, driven by competition for resources among organisms, which are part of a complex network of multitrophic interactions. Here, we demonstrate that the interaction between tomato plants and a phytophagous insect is driven by a hidden interplay between their respective microbiotas. Tomato plants colonized by the soil fungus Trichoderma afroharzianum, a beneficial microorganism widely used in agriculture as a biocontrol agent, negatively affects the development and survival of the lepidopteran pest Spodoptera littoralis by altering the larval gut microbiota and its nutritional support to the host. Indeed, experiments aimed to restore the functional microbial community in the gut allow a complete rescue. Our results shed light on a novel role played by a soil microorganism in the modulation of plant-insect interaction, setting the stage for a more comprehensive analysis of the impact that biocontrol agents may have on ecological sustainability of agricultural systems.

Trichoderma: a multipurpose, plant-beneficial microorganism for eco-sustainable agriculture.

Trichoderma is a cosmopolitan and opportunistic ascomycete fungal genus including species that are of interest to agriculture as direct biological control agents of phytopathogens. Trichoderma utilizes direct antagonism and competition, particularly in the rhizosphere, where it modulates the composition of and interactions with other microorganisms. In its colonization of plants, on the roots or as an endophyte, Trichoderma has evolved the capacity to communicate with the plant and produce numerous multifaceted benefits to its host. The intricacy of this plant-microorganism association has stimulated a marked interest in research on Trichoderma, ranging from its capacity as a plant growth promoter to its ability to prime local and systemic defence responses against biotic and abiotic stresses and to activate transcriptional memory affecting plant responses to future stresses. This Review discusses the ecophysiology and diversity of Trichoderma and the complexity of its relationships in the agroecosystem, highlighting its potential as a direct and indirect biological control agent, biostimulant and biofertilizer, which are useful multipurpose properties for agricultural applications. We also highlight how the present legislative framework might accommodate the demonstrated evidence of Trichoderma proficiency as a plant-beneficial microorganism contributing towards eco-sustainable agriculture.

Selection of Endophytic Beauveria bassiana as a Dual Biocontrol Agent of Tomato Pathogens and Pests.

Endophytic fungi (EF) can enhance both plant growth and defense barriers against pests and pathogens, contributing to the reduction of chemical pesticides and fertilizers use in agriculture. Beauveria bassiana is an entomopathogenic fungus showing endophytism in several crops, often associated with a good capacity to limit the development of pests and disease agents. However, the diversity of the protective efficacy and plant response to different strains can be remarkable and needs to be carefully assessed for the successful and predictable use of these beneficial microorganisms. This study aims to select B. bassiana strains able to colonize tomato plants as endophytes as well as to control two important disease agents, Botrytis cinerea and Alternaria alternata, and the pest aphid, Macrosiphum euphorbiae. Nine wild-type isolates and one commercial strain were screened for endophytism, then further characterized for plant-growth promotion plus inhibition of disease development and pest infestation. Four isolates proved to have a good control activity against the biotic stressors tested, but only Bb716 was also able to promote plant growth. This work provides a simple workflow for the selection of beneficial EF, paving the way towards more effective use of B. bassiana in Integrate Pest Management (IPM) of tomato.

Bioformulations with Beneficial Microbial Consortia, a Bioactive Compound and Plant Biopolymers Modulate Sweet Basil Productivity, Photosynthetic Activity and Metabolites.

Increasing attention is being given to the development of innovative formulations to substitute the use of synthetic chemicals to improve agricultural production and resource use efficiency. Alternatives can include biological products containing beneficial microorganisms and bioactive metabolites able to inhibit plant pathogens, induce systemic resistance and promote plant growth. The efficacy of such bioformulations can be increased by the addition of polymers as adjuvants or carriers. Trichoderma afroharzianum T22, Azotobacter chroococcum 76A and 6-pentyl-α-pyrone (6PP; a Trichoderma secondary metabolite) were administrated singularly or in a consortium, with or without a carboxymethyl cellulose-based biopolymer (BP), and tested on sweet basil (Ocimum basilicum L.) grown in a protected greenhouse. The effect of the treatments on basil yield, photosynthetic activity and secondary metabolites production was assessed. Photosynthetic efficiency was augmented by the applications of the bioformulations. The applications to the rhizosphere with BP + 6PP and BP + T22 + 76A increased the total fresh weight of basil by 26.3% and 23.6%, respectively. Untargeted LC-MS qTOF analysis demonstrated that the plant metabolome was significantly modified by the treatments. Quantification of the profiles for the major phenolic acids indicated that the treatment with the T22 + 76A consortium increased rosmarinic acid content by 110%. The use of innovative bioformulations containing microbes, their metabolites and a biopolymer was found to modulate the cultivation of fresh basil by improving yield and quality, thus providing the opportunity to develop farming systems with minimal impact on the environmental footprint from the agricultural production process.

Temperature Differentially Influences the Capacity of Trichoderma Species to Induce Plant Defense Responses in Tomato Against Insect Pests.

Species of the ecological opportunistic, avirulent fungus, Trichoderma are widely used in agriculture for their ability to protect crops from the attack of pathogenic fungi and for plant growth promotion activity. Recently, it has been shown that they may also have complementary properties that enhance plant defense barriers against insects. However, the use of these fungi is somewhat undermined by their variable level of biocontrol activity, which is influenced by environmental conditions. Understanding the source of this variability is essential for its profitable and wide use in plant protection. Here, we focus on the impact of temperature on Trichoderma afroharzianum T22, Trichoderma atroviride P1, and the defense response induced in tomato by insects. The in vitro development of these two strains was differentially influenced by temperature, and the observed pattern was consistent with temperature-dependent levels of resistance induced by them in tomato plants against the aphid, Macrosiphum euphorbiae, and the noctuid moth, Spodoptera littoralis. Tomato plants treated with T. afroharzianum T22 exhibited enhanced resistance toward both insect pests at 25°C, while T. atroviride P1 proved to be more effective at 20°C. The comparison of plant transcriptomic profiles generated by the two Trichoderma species allowed the identification of specific defense genes involved in the observed response, and a selected group was used to assess, by real-time quantitative reverse transcription PCR (qRT-PCR), the differential gene expression in Trichoderma-treated tomato plants subjected to the two temperature regimens that significantly affected fungal biological performance. These results will help pave the way toward a rational selection of the most suitable Trichoderma isolates for field applications, in order to best face the challenges imposed by local environmental conditions and by extreme climatic shifts due to global warming.

Coordination Properties of the Fungal Metabolite Harzianic Acid toward Toxic Heavy Metals.

Some Trichoderma strains are known for their capacity to produce harzianic acid, a metabolite belonging to the tetramic acid derivatives. Harzianic acid has interesting biological properties, such as antimicrobial activities against phytopathogenic fungi and promotion of plant growth. It also possesses remarkable chemical properties, including the chelating properties toward essential transition metals, which might be related to the biological activities. Increasing knowledge on chelating properties might be relevant for understanding the various beneficial effects of harzianic acid in the interaction between the producer fungi and plants. In this work, the coordination capacity of harzianic acid was studied to evaluate the formation and stability of complexes formed with toxic heavy metals (i.e., Cd2+, Co2+, Ni2+, and Pb2+), which might have a crucial role in the tolerance of plants growing in metal-contaminated soils and in abiotic stress.

Metabolites of a Drechslera sp. endophyte with potential as biocontrol and bioremediation agent.

Endophytic fungi have several well-established beneficial effects on plant health and growth, and are a huge source of bioactive compounds. The endophyte Drechslera sp. strain 678, isolated from the roots of an Australian native grass Neurachne alopecuroidea, demonstrated efficacy against four plant pathogens (Pythium ultimum, Rhizoctonia solani, Botrytis cinerea, Alternaria alternata). In addition, strain 678 was capable of degrading a common additive used in gasoline, known as methyl tertiary-butyl ether (MtBE). Thus, the organic extracts obtained from the culture filtrate of strain 678 were studied. Metabolomic analysis revealed the presence of two major bioactive metabolites, monocerin and an alkynyl substituted epoxycyclohexenone derivative, which showed good antifungal activity. The Drechslera sp. strain 678 and its compounds show promise for applications in biocontrol and bioremediation activities in agriculture or as a remediation option for MtBE contamination in soil.

Antimicrobial activity of harzianic acid against Staphylococcus pseudintermedius.

The emerging concern about the increase of antibiotic resistance has encouraged research efforts to develop effective alternatives to counteract bacterial infections. Herein, we studied a new perspective to therapeutic treatment against Staphylococcus pseudintermedius, an opportunistic pathogen documented as the major cause of skin, ear, and post-operative bacterial infections in dogs and cats. Antimicrobial activity of secondary metabolites produced by selected microbial strains belonging to Trichoderma, Talaromyces, Clonostachys and Coniothyrium fungal genera has been tested against S. pseudintermedius. Several extracts, particularly those obtained from Trichoderma harzianum E45 and ET45, showed a significant antimicrobial activity towards S. pseudintermedius methicillin-resistant (MRSP) and methicillin-susceptible (MSSP) strains. Bioassay-guided fractionation of E45 and ET45 extracts allowed to isolate harzianic acid as the major compound responsible for biological activities (e.g. antimicrobial, antibiofilm formation and biofilm disaggregation).

An Innovative Olive Pâté with Nutraceutical Properties.

Food plays a central role in health, especially through consumption of plant-derived foods. Functional foods, supplements, and nutraceuticals are increasingly entering the market to respond to consumer demand for healthy products. They are foods, supplements, and ingredients which offer health benefits beyond the standard nutritional value. Some benefits are associated with phenolic compounds and phytochemicals with antioxidant properties. An olive pâté (OP) was added with antioxidants derived from olive mill wastewater (OMWW) to obtain a functional product rich in phenolic compounds. The olive pâté is produced from the ground olive pericarp, which shows an excellent natural antioxidant content. The OMWW is a waste product from oil processing, which is also rich in phenolic compounds. The result was a product rich in trans-resveratrol, OH tyrosol, and tyrosol in concentrations such as satisfying the European community's claims regarding the possible antioxidant action on plasma lipids with excellent shelf-life stability. The total phenolic content was assayed by a colorimetric method, the antioxidant activity by the ABTS [(2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)] test, the phenolic profile by Q Exactive Orbitrap LC-MS/MS. The shelf-life stability was confirmed by yeast, molds, and total microbial count, pH, and water activity determinations, and the best pasteurization parameters were determined. The palatability was judged as excellent.

Trichoderma Applications on Strawberry Plants Modulate the Physiological Processes Positively Affecting Fruit Production and Quality.

Many Trichoderma spp. are successful plant beneficial microbial inoculants due to their ability to act as biocontrol agents with direct antagonistic activities to phytopathogens, and as biostimulants capable of promoting plant growth. This work investigated the effects of treatments with three selected Trichoderma strains (T22, TH1, and GV41) to strawberry plants on the productivity, metabolites and proteome of the formed fruits. Trichoderma applications stimulated plant growth, increased strawberry fruit yield, and favored selective accumulation of anthocyanins and other antioxidants in red ripened fruits. Proteomic analysis of fruits harvested from the plants previously treated with Trichoderma demonstrated that the microbial inoculants highly affected the representation of proteins associated with responses to stress/external stimuli, nutrient uptake, protein metabolism, carbon/energy metabolism and secondary metabolism, also providing a possible explanation to the presence of specific metabolites in fruits. Bioinformatic analysis of these differential proteins revealed a central network of interacting molecular species, providing a rationale to the concomitant modulation of different plant physiological processes following the microbial inoculation. These findings indicated that the application of Trichoderma-based products exerts a positive impact on strawberry, integrating well with previous observations on the molecular mechanisms activated in roots and leaves of other tested plant species, demonstrating that the efficacy of using a biological approach with beneficial microbes on the maturing plant is also able to transfer advantages to the developing fruits.

Heterologous Expression of PKPI and Pin1 Proteinase Inhibitors Enhances Plant Fitness and Broad-Spectrum Resistance to Biotic Threats.

Kunitz-type (PKPI) and Potato type I (Pin1) protease inhibitors (PIs) are two families of serine proteinase inhibitors often associated to plant storage organs and with well known insecticidal and nematicidal activities. Noteworthy, their ability to limit fungal and bacterial pathogenesis in vivo or to influence plant physiology has not been investigated in detail. To this aim, we generated a set of PVX-based viral constructs to transiently and heterologously express two potato PKPI (PKI1, PKI2) and three potato Pin1 (PPI3A2, PPI3B2, PPI2C4) genes in Nicotiana benthamiana plants, a widely used model for plant-pathogen interaction studies. Interestingly, transgenic plants expressing most of the tested PIs showed to be highly resistant against two economically important necrotrophic fungal pathogens, Botrytis cinerea and Alternaria alternata. Unexpectedly, overexpression of the PKI2 Kunitz-type or of the PPI2C4 and PPI3A2 Potato type I inhibitor genes also lead to a dramatic reduction in the propagation and symptom development produced by the bacterial pathogen Pseudomonas syringae. We further found that localized expression of PPI2C4 and PKI2 in N. benthamiana leaves caused an increase in cell expansion and proliferation which lead to tissue hypertrophy and trichome accumulation. In line with this, the systemic expression of these proteins resulted in plants with enhanced shoot and root biomass. Collectively, our results indicate that PKPI and Pin1 PIs might represent valuable tools to simultaneously increase plant fitness and broad-spectrum resistance toward phytopathogens.

Effect of Trichoderma Bioactive Metabolite Treatments on the Production, Quality, and Protein Profile of Strawberry Fruits.

Fungi of the genus Trichoderma produce secondary metabolites having several biological activities that affect plant metabolism. We examined the effect of three Trichoderma bioactive metabolites (BAMs), namely, 6-pentyl-α-pyrone (6PP), harzianic acid (HA), and hydrophobin 1 (HYTLO1), on yield, fruit quality, and protein representation of strawberry plants. In particular, 6PP and HA increased the plant yield and number of fruits, when compared to control, while HYTLO1 promoted the growth of the roots and increased the total soluble solids content up to 19% and the accumulation of ascorbic acid and cyanidin 3-O-glucoside in red ripened fruits. Proteomic analysis showed that BAMs influenced the representation of proteins associated with the protein metabolism, response to stress/external stimuli, vesicle trafficking, carbon/energy, and secondary metabolism. Results suggest that the application of Trichoderma BAMs affects strawberry plant productivity and fruit quality and integrate previous observations on deregulated molecular processes in roots and leaves of Trichoderma-treated plants with original data on fruits.

New Strategies in the Cultivation of Olive Trees and Repercussions on the Nutritional Value of the Extra Virgin Olive Oil.

The health advantages of extra-virgin olive oil (EVOO) are ascribed mainly to the antioxidant ability of the phenolic compounds. Secoiridoids, hydroxytyrosol, tyrosol, phenolic acid, and flavones, are the main nutraceutical substances of EVOO. Applications of beneficial microbes and/or their metabolites impact the plant metabolome. In this study the effects of application of selected Trichoderma strains or their effectors (secondary metabolites) on the phenolic compounds content and antioxidant potential of the EVOOs have been evaluated. For this purpose, Trichoderma virens (strain GV41) and Trichoderma harzianum (strain T22), well-known biocontrol agents, and two their metabolites harzianic acid (HA) and 6-pentyl-α-pyrone (6PP) were been used to treat plants of Olea europaea var. Leccino and var. Carolea. Then the nutraceutical potential of EVOO was evaluated. Total phenolic content was estimated by Folin-Ciocalteau's assay, metabolic profile by High-Resolution Mass spectroscopy (HRMS-Orbitrap), and antioxidant activity by DPPH and ABTS assays. Our results showed that in the cultivation of the olive tree, T22 and its metabolites improve the nutraceutical value of the EVOOs modulating the phenolic profile and improving antioxidants activity.

Bivalent Metal-Chelating Properties of Harzianic Acid Produced by Trichoderma pleuroticola Associated to the Gastropod Melarhaphe neritoides.

Harzianic acid is a secondary metabolite of Trichoderma, structurally belonging to the dienyltetramic acid subgroup of the tetramic acids. Biological activities of harzianic acid are of great interest for its antimicrobial and plant growth-promoting activities, which might be related to its chelating properties. In the present work harzianic acid, isolated from cultures of a strain of Trichoderma pleuroticola associated to the gastropod Melarhaphe neritoides, was studied as a complexant agent of a number of biologically relevant transition metals (i.e., Zn2+, Fe2+, Cu2+, and Mn2+), using UV-VIS, potentiometry, MS and NMR techniques. Our findings show the coordination capacity of harzianic acid toward the above cations through the formation of neutral or charged complexes in a variable ratio depending on the metal and pH conditions.

Plant Dynamic Metabolic Response to Bacteriophage Treatment After Xanthomonas campestris pv. campestris Infection.

Periodic epidemics of black rot disease occur worldwide causing substantial yield losses. Xanthomonas campestris pv. campestris (Xcc) represents one of the most common bacteria able to cause the above disease in cruciferous plants such as broccoli, cabbage, cauliflower, and Arabidopsis thaliana. In agriculture, several strategies are being developed to contain the Xanthomonas infection. The use of bacteriophages could represent a valid and efficient approach to overcome this widespread phenomenon. Several studies have highlighted the potential usefulness of implementing phage therapy to control plant diseases as well as Xcc infection. In the present study, we characterized the effect of a lytic phage on the plant Brassica oleracea var. gongylodes infected with Xcc and, for the first time, the correlated plant metabolic response. The results highlighted the potential benefits of bacteriophages: reduction of bacterium proliferation, alteration of the biofilm structure and/or modulation of the plant metabolism and defense response.

Antibiofilm Activity of a Trichoderma Metabolite against Xanthomonas campestris pv. campestris, Alone and in Association with a Phage.

Biofilm protects bacteria against the host's immune system and adverse environmental conditions. Several studies highlight the efficacy of lytic phages in the prevention and eradication of bacterial biofilms. In this study, the lytic activity of Xccφ1 (Xanthomonas campestris pv. campestris-specific phage) was evaluated in combination with 6-pentyl-α-pyrone (a secondary metabolite produced by Trichoderma atroviride P1) and the mineral hydroxyapatite. Then, the antibiofilm activity of this interaction, called a φHA6PP complex, was investigated using confocal laser microscopy under static and dynamic conditions. Additionally, the mechanism used by the complex to modulate the genes (rpf, gumB, clp and manA) involved in the biofilm formation and stability was also studied. Our results demonstrated that Xccφ1, alone or in combination with 6PP and HA, interfered with the gene pathways involved in the formation of biofilm. This approach can be used as a model for other biofilm-producing bacteria.

Autotrophic and Heterotrophic Growth Conditions Modify Biomolecole Production in the Microalga Galdieria sulphuraria (Cyanidiophyceae, Rhodophyta).

Algae have multiple similarities with fungi, with both belonging to the Thallophyte, a polyphyletic group of non-mobile organisms grouped together on the basis of similar characteristics, but not sharing a common ancestor. The main difference between algae and fungi is noted in their metabolism. In fact, although algae have chlorophyll-bearing thalloids and are autotrophic organisms, fungi lack chlorophyll and are heterotrophic, not able to synthesize their own nutrients. However, our studies have shown that the extremophilic microalga Galderia sulphuraria (GS) can also grow very well in heterotrophic conditions like fungi. This study was carried out using several approaches such as scanning electron microscope (SEM), gas chromatography/mass spectrometry (GC/MS), and infrared spectrophotometry (ATR-FTIR). Results showed that the GS, strain ACUF 064, cultured in autotrophic (AGS) and heterotrophic (HGS) conditions, produced different biomolecules. In particular, when grown in HGS, the algae (i) was 30% larger, with an increase in carbon mass that was 20% greater than AGS; (ii) produced higher quantities of stearic acid, oleic acid, monounsaturated fatty acids (MUFAs), and ergosterol; (iii) produced lower quantities of fatty acid methyl esters (FAMEs) such as methyl palmytate, and methyl linoleate, saturated fatty acids (SFAs), and poyliunsaturated fatty acids (PUFAs). ATR-FTIR and principal component analysis (PCA) statistical analysis confirmed that the macromolecular content of HGS was significantly different from AGS. The ability to produce different macromolecules by changing the trophic conditions may represent an interesting strategy to induce microalgae to produce different biomolecules that can find applications in several fields such as food, feed, nutraceutical, or energy production.

Reinforced Olive Pâté as a Source of Antioxidants with Positive Effects on Young Smokers.

Background and objectives: Olive pâté (OP) is an olive-derived product with potentially beneficial effects on human health due to the presence of natural antioxidants. The present dietary supplementation study aimed to evaluate the effects on blood antioxidant levels of an olive pâté reinforced with natural antioxidants (ROP) recovered from olive mill waste. Materials and methods: Ninety-eight healthy volunteers (M = 54, 55%, age 18-25) were divided into two groups: A (n = 49), practicing three or more days of physical activity a week, and B (n = 49), practicing less than two. Each group was split into two subgroups, receiving dietary supplementation with OP or ROP. The status of smoker was also recorded, and a biological antioxidant potential (BAP) test was performed on each subject. Results: The BAP values increased with both OP (n = 30) and ROP (n = 68) but ROP supplementation showed higher increments (736.9 µmol/L) than OP (339.6). The increment was significantly higher for smokers (n = 15), 1122.9 vs. non-smokers (n = 53), 635.7, with values in percent of baseline, respectively, 34.6% and 16.2% (P < 0.01). Conclusions: The ROP nutritional supplementation appears useful to increase antioxidant activity, with better effect in smokers; further studies should confirm the finding and investigate its biological bases.

Transcriptome and Metabolome Reprogramming in Tomato Plants by Trichoderma harzianum strain T22 Primes and Enhances Defense Responses Against Aphids.

Beneficial fungi in the genus Trichoderma are among the most widespread biocontrol agents of plant pathogens. Their role in triggering plant defenses against pathogens has been intensely investigated, while, in contrast, very limited information is available on induced barriers active against insects. The growing experimental evidence on this latter topic looks promising, and paves the way toward the development of Trichoderma strains and/or consortia active against multiple targets. However, the predictability and reproducibility of the effects that these beneficial fungi is still somewhat limited by the lack of an in-depth understanding of the molecular mechanisms underlying the specificity of their interaction with different crop varieties, and on how the environmental factors modulate this interaction. To fill this research gap, here we studied the transcriptome changes in tomato plants (cultivar "Dwarf San Marzano") induced by Trichoderma harzianum (strain T22) colonization and subsequent infestation by the aphid Macrosiphum euphorbiae. A wide transcriptome reprogramming, related to metabolic processes, regulation of gene expression and defense responses, was induced both by separate experimental treatments, which showed a synergistic interaction when concurrently applied. The most evident expression changes of defense genes were associated with the multitrophic interaction Trichoderma-tomato-aphid. Early and late genes involved in direct defense against insects were induced (i.e., peroxidase, GST, kinases and polyphenol oxidase, miraculin, chitinase), along with indirect defense genes, such as sesquiterpene synthase and geranylgeranyl phosphate synthase. Targeted and untargeted semi-polar metabolome analysis revealed a wide metabolome alteration showing an increased accumulation of isoprenoids in Trichoderma treated plants. The wide array of transcriptomic and metabolomics changes nicely fit with the higher mortality of aphids when feeding on Trichoderma treated plants, herein reported, and with the previously observed attractiveness of these latter toward the aphid parasitoid Aphidius ervi. Moreover, Trichoderma treated plants showed the over-expression of transcripts coding for several families of defense-related transcription factors (bZIP, MYB, NAC, AP2-ERF, WRKY), suggesting that the fungus contributes to the priming of plant responses against pest insects. Collectively, our data indicate that Trichoderma treatment of tomato plants induces transcriptomic and metabolomic changes, which underpin both direct and indirect defense responses.

Trichoderma atroviride P1 Colonization of Tomato Plants Enhances Both Direct and Indirect Defense Barriers Against Insects.

Numerous microbial root symbionts are known to induce different levels of enhanced plant protection against a variety of pathogens. However, more recent studies have demonstrated that beneficial microbes are able to induce plant systemic resistance that confers some degree of protection against insects. Here, we report how treatments with the fungal biocontrol agent Trichoderma atroviride strain P1 in tomato plants induce responses that affect pest insects with different feeding habits: the noctuid moth Spodoptera littoralis (Boisduval) and the aphid Macrosiphum euphorbiae (Thomas). We observed that the tomato plant-Trichoderma P1 interaction had a negative impact on the development of moth larvae and on aphid longevity. These effects were attributed to a plant response induced by Trichoderma that was associated with transcriptional changes of a wide array of defense-related genes. While the impact on aphids could be related to the up-regulation of genes involved in the oxidative burst reaction, which occur early in the defense reaction, the negative performance of moth larvae was associated with the enhanced expression of genes encoding for protective enzymes (i.e., Proteinase inhibitor I (PI), Threonine deaminase, Leucine aminopeptidase A1, Arginase 2, and Polyphenol oxidase) that are activated downstream in the defense cascade. In addition, Trichoderma P1 produced alterations in plant metabolic pathways leading to the production and release of volatile organic compounds (VOCs) that are involved in the attraction of the aphid parasitoid Aphidius ervi, thus reinforcing the indirect plant defense barriers. Our findings, along with the evidence available in the literature, indicate that the outcome of the tripartite interaction among plant, Trichoderma, and pests is highly specific and only a comprehensive approach, integrating both insect phenotypic changes and plant transcriptomic alterations, can allow a reliable prediction of its potential for plant protection.