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Soil-borne vascular pathogens pose serious threats to agriculture with complex invasion strategies that remain poorly characterized compared to foliar pathogens. While foliar pathogens like Magnaporthe oryzae employ specialized appressoria to penetrate plant surfaces through a combination of mechanical force and enzymatic degradation, the invasion mechanisms of vascular pathogens that lack classical appressoria have remained largely theoretical. The nanoscale processes governing root penetration and colonization by these pathogens are particularly challenging to visualize due to technical limitations of conventional microscopy. Here we show, using phase-contrast X-ray computed microtomography and advanced microscopy, that Fusarium oxysporum ( Fo ) employs distinct mitogen-activated protein kinase (MAPK) cascades to orchestrate root invasion through unprecedented morphological plasticity. We identify previously undocumented appressoria-like structures that facilitate physical penetration, while demonstrating that Fo exhibits remarkable cellular adaptability, reducing hyphal diameter by more than 20-fold (from 5 μm to 220 nm) to navigate confined plant spaces, a dramatic morphological transition previously thought impossible. By using cellulase-deficient mutants, we demonstrate that cellulolytic activity is dispensable for surface breach and submicrometric hyphal colonization, establishing that mechanical force generation rather than enzymatic degradation is the primary determinant of successful host penetration. Three-dimensional reconstruction reveals a quantitative correlation between fungal proliferation and progressive embolism formation, with distinct MAPK pathways differentially regulating penetration force generation (Fmk1), osmotic adaptation during apoplastic colonization (Hog1), and directional growth toward vascular tissues (Mpk1). These findings provide a mechanistic framework for vascular wilt pathogenesis and reveal potential targets for controlling these economically devastating plant diseases. Using X-ray microtomography, Turrà and colleagues show that Fusarium oxysporum penetrates plant roots via appressoria-like structures through mechanical force. Three MAPK pathways govern force generation, osmoadaptation, and vascular  bundle reach.

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.

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.

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.

Background ATP-binding cassette proteins have been recognized as playing a crucial role in the regulation of growth and resistance processes in all kingdoms of life. They have been deeply studied in vertebrates because of their role in drug resistance, but much less is known about ABC superfamily functions in plants. Results Recently released plant genome sequences allowed us to identify 803 ABC transporters in four vascular plants ( Oryza. sativa, Solanum lycopersicum, Solanum tuberosum and Vitis vinifera ) and 76 transporters in the green alga Volvox carteri , by comparing them with those reannotated in Arabidopsis thaliana and the yeast Saccharomyces cerevisiae . Retrieved proteins have been phylogenetically analysed to infer orthologous relationships. Most orthologous relationships in the A, D, E and F subfamilies were found, and interesting expansions within the ABCG subfamily were observed and discussed. A high level of purifying selection is acting in the five ABC subfamilies A, B, C, D and E. However, evolutionary rates of recent duplicate genes could influence vascular plant genome diversification. The transcription profiles of ABC genes within tomato organs revealed a broad functional role for some transporters and a more specific activity for others, suggesting the presence of key ABC regulators in tomato. Conclusions The findings achieved in this work could contribute to address several biological questions concerning the evolution of the relationship between genomes of different species. Plant ABC protein inventories obtained could be a valuable tool both for basic and applied studies. Indeed, interpolation of the putative role of gene functions can accelerate the discovering of new ABC superfamily members.

Background The efficacy of biocontrol agents depends critically on their environmental adaptability, with nutrient availability being a key determinant of their success. In Trichoderma species, the mechanisms linking nutrient sensing to physiological responses remain poorly understood, despite their importance for biocontrol applications. Results This study reveals how different nitrogen sources fundamentally regulate the physiology and metabolism of Trichoderma harzianum through pH modulation. Under nutrient-rich conditions, T. harzianum exhibited a biphasic pH response characterized by initial acidification followed by alkalinization, which correlated with enhanced sporulation. Examining specific nitrogen sources, we found that sodium nitrate induced environmental alkalinization, while ammonium nitrate caused sustained acidification. These pH changes were linked to distinct physiological responses: alkaline conditions promoted sporulation, while acidic conditions enhanced mycelial growth and triggered specific metabolic responses. Notably, acidic conditions specifically induced the production of harzianic acid and related bioactive compounds, suggesting pH-dependent regulation of secondary metabolism. This nitrogen-dependent pH modulation pattern was conserved across Trichoderma species, as demonstrated by parallel responses in T. asperellum . Conclusions Our findings establish nitrogen source availability as a master regulator of Trichoderma physiology through pH-dependent mechanisms, controlling both development and secondary metabolism. This understanding provides new strategies for optimizing biocontrol formulations by manipulating nitrogen sources and pH conditions to enhance both fungal fitness and beneficial metabolite production. Graphical Abstract

Beneficial interactions of rhizosphere microorganisms are widely exploited for plant biofertilization and mitigation of biotic and abiotic constraints. To provide new insights into the onset of the roots–beneficial microorganisms interplay, we characterised the transcriptomes expressed in tomato roots at 24, 48 and 72 h post inoculation with the beneficial fungus Trichoderma harzianum T22 and analysed the epigenetic and post-trascriptional regulation mechanisms. We detected 1243 tomato transcripts that were differentially expressed between Trichoderma -interacting and control roots and 83 T. harzianum transcripts that were differentially expressed between the three experimental time points. Interaction with Trichoderma triggered a transcriptional response mainly ascribable to signal recognition and transduction, stress response, transcriptional regulation and transport. In tomato roots, salicylic acid, and not jasmonate, appears to have a prominent role in orchestrating the interplay with this beneficial strain. Differential regulation of many nutrient transporter genes indicated a strong effect on plant nutrition processes, which, together with the possible modifications in root architecture triggered by ethylene/indole-3-acetic acid signalling at 72 h post inoculation may concur to the well-described growth-promotion ability of this strain. Alongside, T. harzianum -induced defence priming and stress tolerance may be mediated by the induction of reactive oxygen species, detoxification and defence genes. A deeper insight into gene expression and regulation control provided first evidences for the involvement of cytosine methylation and alternative splicing mechanisms in the plant– Trichoderma interaction. A model is proposed that integrates the plant transcriptomic responses in the roots, where interaction between the plant and beneficial rhizosphere microorganisms occurs. Symbiosis: the genetics of beneficial tomato root–fungus interaction The fungus Trichoderma harzianum induces differentiated protein production in tomato roots that benefits plant nutrition and survivability. Microorganisms advantageous to plants have long been exploited in agriculture; however, with limited studies into the interface of Trichoderma fungus and plants—the roots. Italian researchers, led by the Research Centre for Vegetable and Ornamental Crops' Nunzio D'Agostino and the National Research Council’s Marina Tucci, inoculated tomato plant roots with T. harzianum over 72 h, finding over 1200 examples of differential gene expression and post-expression modification that resulted in improved plant growth and immune system regulation. The interaction also induces a root change that likely promotes further interaction with the fungus and increased stress tolerance via promoting antioxidation and defensive activity. The team’s work provides early evidence of the molecular mechanisms behind root–fungus symbiosis and may help to inform crop breeding and fertilisation strategies.

Viticulture is vital to Italy’s agricultural sector, since it significantly contributes to the global wine industry. Microflora and microfauna are considered important factors for soil quality, improving grapevine growth, and promoting resistance to biotic and abiotic stresses. This study examined the impact of selected Trichoderma strains (T. harzianum M10 and T. afroharzianum T22) and their secondary metabolite 6-pentyl-α-pyrone (6PP) on the soil microbiome, the metabolome, and physiological changes of grapevines. Before treatment application, low levels of plant-parasitic nematodes (Rotylenchulus spp., Xiphinema pachtaicum) were found in the soil, together with pathogens (Fusarium spp., Neonectria spp.) and beneficial microbes (Clonostachys rosea, Pseudomonas spp.). Metagenomic analysis revealed significant treatment impacts in the soil microbiome, with T22 and 6PP treatments increasing Proteobacteria abundance, while slight variations of fungal communities and no significant differences in nematofauna were found. Metabolomic analysis showed that treatments induced grapevines to produce antioxidant secondary metabolites able to boost plant defense against abiotic and biotic stresses and increase nutraceutical grapes’ value. Finally, T22 treatment increased the grapes’ winemaking value, raising their Brix grade. Our results demonstrate that microbial or metabolite-based treatments could affect the soil microbiome composition, grapevine health and resilience, and grapes’ oenological and nutraceutical properties.

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 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 ( , ) and three potato ( , , ) genes in 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, and . Unexpectedly, overexpression of the Kunitz-type or of the and Potato type I inhibitor genes also lead to a dramatic reduction in the propagation and symptom development produced by the bacterial pathogen . We further found that localized expression of and in 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.

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.