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.

Regulation of the expression of the two major chitinase genes,ech42 (encoding the CHIT42 endochitinase) andnag1 (encoding the CHIT73N-acetyl-β-d-glucosaminidase), of the chitinolytic system of the mycoparasitic biocontrol fungusTrichoderma atroviride (= Trichoderma harzianumP1) was investigated by using a reporter system based on theAspergillus niger glucose oxidase. Strains harboring fusions of the ech42 or nag1 5′ upstream noncoding sequences with the A. niger goxAgene displayed a glucose oxidase activity pattern that was consistent under various conditions with expression of the nativeech42 and nag1 genes, as assayed by Northern analysis. The expression product of goxA in the mutants was completely secreted into the medium, detectable on Western blots, and quantifiable by enzyme-linked immunosorbent assay. nag1gene expression was triggered during growth on fungal (Botrytis cinerea) cell walls and on the chitin degradation productN-acetylglucosamine. N-Acetylglucosamine, di-N-acetylchitobiose, or tri-N-acetylchitotriose also induced nag1 gene expression when added to mycelia pregrown on different carbon sources.ech42 expression was also observed during growth on fungal cell walls but, in contrast, was not triggered by addition of chitooligomers to pregrown mycelia. Significant ech42expression was observed after prolonged carbon starvation, independent of the use of glucose or glycerol as a carbon source, suggesting that relief of carbon catabolite repression was not involved in induction during starvation. In addition, ech42 gene transcription was triggered by physiological stress, such as low temperature, high osmotic pressure, or the addition of ethanol. Four copies of a putative stress response element (CCCCT) were found in the ech42promoter.

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.

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.

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.

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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.

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