Mechanisms for plant growth promotion activated by Trichoderma in natural and managed terrestrial ecosystems.

Trichoderma spp. are free-living fungi present in virtually all terrestrial ecosystems. These soil fungi can stimulate plant growth and increase plant nutrient acquisition of macro- and micronutrients and water uptake. Generally, plant growth promotion by Trichoderma is a consequence of the activity of potent fungal signaling metabolites diffused in soil with hormone-like activity, including indolic compounds as indole-3-acetic acid (IAA) produced at concentrations ranging from 14 to 234 µg l-1, and volatile organic compounds such as sesquiterpene isoprenoids (C15), 6-pentyl-2H-pyran-2-one (6-PP) and ethylene (ET) produced at levels from 10 to 120 ng over a period of six days, which in turn, might impact plant endogenous signaling mechanisms orchestrated by plant hormones. Plant growth stimulation occurs without the need of physical contact between both organisms and/or during root colonization. When associated with plants Trichoderma may cause significant biochemical changes in plant content of carbohydrates, amino acids, organic acids and lipids, as detected in Arabidopsis thaliana, maize (Zea mays), tomato (Lycopersicon esculentum) and barley (Hordeum vulgare), which may improve the plant health status during the complete life cycle. Trichoderma-induced plant beneficial effects such as mechanisms of defense and growth are likely to be inherited to the next generations. Depending on the environmental conditions perceived by the fungus during its interaction with plants, Trichoderma can reprogram and/or activate molecular mechanisms commonly modulated by IAA, ET and abscisic acid (ABA) to induce an adaptative physiological response to abiotic stress, including drought, salinity, or environmental pollution. This review, provides a state of the art overview focused on the canonical mechanisms of these beneficial fungi involved in plant growth promotion traits under different environmental scenarios and shows new insights on Trichoderma metabolites from different chemical classes that can modulate specific plant growth aspects. Also, we suggest new research directions on Trichoderma spp. and their secondary metabolites with biological activity on plant growth.

First report of Irpex rosettiformis causing white root rot in avocado trees in Michoacán, México.

In México, avocado production is an important economic source. In the last season it generated $ 3. 27 billion USD of foreign currency in the country. Irpex spp. are wood decay fungi. In the period 2019-2022, in the state of Michoacán (19°13' N; 101°55' W), México, basidiomes of Irpex sp. were observed on the base of trunks and crowns of 5-years-old and older avocado (Persea americana) trees. The trees exhibited disease symptoms that included white root rot, leaf yellowing, small leaves, branch diebacks, generalized defoliation, apical flaccidity, abundant but small sun burnt fruits due to the lack of foliage, and after 2-4 years of first disease appearance, the infected trees died. In the place where fungus was established, abundant white and cottony mycelium was formed, which caused trees decay. The incidence of the disease in the sampled orchards was estimated to be 30% per ha with 350 - 400 trees, which was determined through a simple sampling design focused on trees with signs and symptoms of the disease due to the phytopathogen. Samples of infected tissue (roots and stems) and fungal basidiomes were collected from 90 trees (5-6 per orchard). The symptomatic avocado trees studied were randomly selected from 17 orchards. For the fungal macroscopic characterization, the synoptic keys described by Gilbertson and Ryvarden (1986) and by Largent (1973) were used. The samples showed typical structures corresponding to Irpex sp., including rosettes, annual basidiomes, a system of monomitic hyphae, and subglobose basidiospores. In vitro fungal isolation from basidiomes and infected tree tissues was done according to the protocol of Agrios (2004). The fungal strains were maintained on PDA at 28 °C. At 16 days of incubation the colonies were opaque, whitish with fluffy and corky mycelium. Microscopic analysis of the fungus showed typical yellowish spores, with an ellipsoid shape of 3-4 x 4-5.5 µm (50 accounted structures per isolate [N=19]) and basidia of 20-25 x 4.5-5.5 µm (n=20 basidiomes). For molecular characterization, two molecular markers were used, the internal transcribed spacer rDNA-ITS1 5.8 rDNA-ITS2 (ITS; White et al. 1990) and the large ribosomal subunit (LSU; Vilgalys and Hester 1990). The PCR reaction was performed as described by Martínez-González et al. (2017). The consensus sequences were compared with those deposited in the NCBI-GenBank, using the BLASTN 2.2.19 tool (Zhang et al. 2000), the samples showed 99% match with the species, Irpex rosettiformis. GenBank accession numbers of the submitted isolates are summarized in supplementary Table 4. To test Koch's postulates, 3-months old avocado plants grown in greenhouse conditions were inoculated (n = 10 per each isolate [N= 19]) on the roots with 3 g of I. rosettiformis mycelium. The experiment was done twice with 20 non-inoculated plants as control. After 67 days, basidiomes (50 x 70 x 1.5 mm in average) were observed where the disease incidence was >77%, with subsequent tree decline. The pathogen was re-isolated in vitro in PDA and its identity was confirmed by morphological characteristics of mycelium. This work shows that I. rosettiformis is not only a wood decay fungus, but also a phytopathogen, the causative agent of white root rot disease in P. americana var. drymifolia, cultivar 'Hass', which establishes a precedent for monitoring and preventing its proliferation to other regions in the American continent and the world where nursery avocado seedlings are exported.

Volatiles released by Beauveria bassiana induce oviposition behavior in the fall armyworm Spodoptera frugiperda (Lepidoptera: Noctuidae).

Microbial volatile organic compounds may act as semiochemicals, inciting different behavioral responses in insects. Beauveria bassiana is an entomopathogenic fungus, and physiological and environmental factors are positively related to fungal virulence. In this study, we examined the volatile profiles produced by eight B. bassiana strains, isolated from soil plots and mycosed insect cadavers, with different speeds of kill and determined if these compounds induce oviposition behavior in Spodoptera frugiperda. Fungal volatilome analysis revealed differences between the isolates. Isolates from mycosed insects showed higher virulence, larger egg mass area and length, and a higher number of eggs by mass, than those obtained from soil. Furthermore, a dilution of the fungal odoriferous compounds increased the insect response, suggesting that S. frugiperda is highly susceptible to the fungal compound's fingerprint. Otherwise, the insect response to the natural blend of volatiles released by the fungus was different from that obtained with 3-methylbutanol, which was the most abundant compound in all isolates. The ability of an entomopathogen to produce volatiles that can induce olfactory stimulation of egg-laying behavior could represent an ecological adaptive advantage in which the entomopathogen stimulates the insect population growth.

In a belowground multitrophic interaction, Trichoderma harzianum induces maize root herbivore tolerance against Phyllophaga vetula.

BACKGROUND: Trichoderma spp. are soil fungi that interact with plant roots and associated biota such as other microorganisms and soil fauna. However, information about their interactions with root-feeding insects is limited. Here, interactions between Trichoderma harzianum and the root-feeding insect Phyllophaga vetula, a common insect pest in maize agroecosystems, were examined. RESULTS: Applications of T. harzianum and P. vetula to the root system increased and decreased maize growth, respectively. Induced tolerance against herbivore attack was provided by T. harzianum maintaining a robust and functional root system as evidenced by the increased uptake of Cu, Ca, Mg, Na and K. Herbivore tolerance also coincided with changes in the emission of root volatile terpenes known to induce indirect defense responses and attract natural enemies of the herbivore. More importantly, T. harzianum induced de novo emission of several sesquiterpenes such as ß-caryophyllene and δ-cadinene. In addition, single and combined applications of T. harzianum and P. vetula altered the sucrose content of the roots. Finally, T. harzianum produced 6-pentyl-2H-pyran-2-one (6-PP) a volatile compound that may act as an antifeedant-signaling compound mitigating root herbivory by P. vetula. CONCLUSION: Our results provide novel information about belowground multitrophic plant-microbe-arthropod interactions between T. harzianum and P. vetula in the maize rhizosphere resulting in alterations in maize phenotypic plant responses, inducing root herbivore tolerance.

The interactions of Trichoderma at multiple trophic levels: inter-kingdom communication.

Trichoderma spp. are universal saprotrophic fungi in terrestrial ecosystems, and as rhizosphere inhabitants, they mediate interactions with other soil microorganisms, plants, and arthropods at multiple trophic levels. In the rhizosphere, Trichoderma can reduce the abundance of phytopathogenic microorganisms, which involves the action of potent inhibitory molecules, such as gliovirin and siderophores, whereas endophytic associations between Trichoderma and the seeds and roots of host plants can result in enhanced plant growth and crop productivity, as well as the alleviation of abiotic stress. Such beneficial effects are mediated via the activation of endogenous mechanisms controlled by phytohormones such as auxins and abscisic acid, as well as by alterations in host plant metabolism. During either root colonization or in the absence of physical contact, Trichoderma can trigger early defense responses mediated by Ca2+ and reactive oxygen species, and subsequently stimulate plant immunity by enhancing resistance mechanisms regulated by the phytohormones salicylic acid, jasmonic acid, and ethylene. In addition, Trichoderma release volatile organic compounds and nitrogen or oxygen heterocyclic compounds that serve as signaling molecules, which have effects on plant growth, phytopathogen levels, herbivorous insects, and at the third trophic level, play roles in attracting the natural enemies (predators and parasitoids) of herbivores. In this paper, we review some of the most recent advances in our understanding of the environmental influences of Trichoderma spp., with particular emphasis on their multiple interactions at different trophic levels.

Increased maize growth and P uptake promoted by arbuscular mycorrhizal fungi coincide with higher foliar herbivory and larval biomass of the Fall Armyworm Spodoptera frugiperda.

Most plant species naturally associate with arbuscular mycorrhizal fungi (AMF), which are known to promote crop nutrition and health in agroecosystems. However, information on how mycorrhizal associations affect plant biotic interactions that occur aboveground with foliar herbivores is limited and needs to be further addressed for the development of pest management strategies. With the objective to examine the influence of maize mycorrhizas on foliar herbivory caused by larvae of Spodoptera frugiperda, a serious pest in maize agroecosystems, we performed a fully factorial greenhouse pot experiment with three factors: Maize genotype (Puma and Milpal H318), AMF (with and without AMF, and without AMF with mineral P) and Insect herbivory (with and without S. frugiperda). Main results showed that inoculation with AMF improved plant growth and foliar P concentration, which coincided with increased foliar damage from herbivory and higher biomass of S. frugiperda larvae. A significant positive correlation between shoot P concentration and larval biomass was also observed. Finally, foliar herbivory by S. frugiperda slightly increased and decreased AMF root colonization in Puma and H318, respectively. In conclusion, our results show that maize plant benefits from AMF in terms of promotion of growth and nutrition, and may also increase the damage caused from insects by improving the food quality of maize leaves for larval growth, which seems to be linked to increased P uptake by the maize mycorrhizal association.

Trichoderma atroviride promotes tomato development and alters the root exudation of carbohydrates, which stimulates fungal growth and the biocontrol of the phytopathogen Phytophthora cinnamomi in a tripartite interaction system.

Several species of Trichoderma promote plant growth and help in defense against root pathogens. The role of root-exuded carbohydrates as chemo-attractive stimuli for Trichoderma colonization is attracting considerable interest. In this project, we studied the interaction between Trichoderma atroviride and tomato (Lycopersicon esculentum L. cv. Río Grande) plants in two different stages, before and during root colonization. In addition, the biocontrol capacity of T. atroviride against the phytopathogen Phytophthora cinnamomi in a tripartite interaction system was examined. We found that the beneficial effects of T. atroviride on plant growth were fine-tuned depending on the progress of interaction. Interestingly, the composition of the carbohydrate exudate from plants interacting with T. atroviride was different from that produced by other treatments and probably provided a nutritional source for the fungus. Particularly, sucrose was found only during root colonization by the fungus. Our data show that root-derived sugars enabled a higher Trichoderma growth rate, and that, in the tripartite interaction system with P. cinnamomi, the fungus competes for space and available soil nutrients more efficiently than P. cinnamomi, thereby antagonizing the growth of the phytopathogen.

Ecological functions of Trichoderma spp. and their secondary metabolites in the rhizosphere: interactions with plants.

Trichodermaspp. are common soil and root inhabitants that have been widely studied due to their capacity to produce antibiotics, parasitize other fungi and compete with deleterious plant microorganisms. These fungi produce a number of secondary metabolites such as non-ribosomal peptides, terpenoids, pyrones and indolic-derived compounds. In the rhizosphere, the exchange and recognition of signaling molecules byTrichodermaand plants may alter physiological and biochemical aspects in both. For example, severalTrichodermastrains induce root branching and increase shoot biomass as a consequence of cell division, expansion and differentiation by the presence of fungal auxin-like compounds. Furthermore,Trichoderma, in association with plant roots, can trigger systemic resistance and improve plant nutrient uptake. The present review describes the most recent advances in understanding the ecological functions ofTrichodermaspp. in the rhizosphere at biochemical and molecular levels with special emphasis on their associations with plants. Finally, through a synthesis of the current body of work, we present potential future research directions on studies related toTrichodermaspp. and their secondary metabolites in agroecosystems.

Mitogen-Activated Protein Kinase 6 and Ethylene and Auxin Signaling Pathways Are Involved in Arabidopsis Root-System Architecture Alterations by Trichoderma atroviride.

Trichoderma atroviride is a symbiotic fungus that interacts with roots and stimulates plant growth and defense. Here, we show that Arabidopsis seedlings cocultivated with T. atroviride have an altered root architecture and greater biomass compared with axenically grown seedlings. These effects correlate with increased activity of mitogen-activated protein kinase 6 (MPK6). The primary roots of mpk6 mutants showed an enhanced growth inhibition by T. atroviride when compared with wild-type (WT) plants, while T. atroviride increases MPK6 activity in WT roots. It was also found that T. atroviride produces ethylene (ET), which increases with l-methionine supply to the fungal growth medium. Analysis of growth and development of WT seedlings and etr1, ein2, and ein3 ET-related Arabidopsis mutants indicates a role for ET in root responses to the fungus, since etr1 and ein2 mutants show defective root-hair induction and enhanced primary-root growth inhibition when cocultivated with T. atroviride. Increased MPK6 activity was evidenced in roots of ctr1 mutants, which correlated with repression of primary root growth, thus connecting MPK6 signaling with an ET response pathway. Auxin-inducible gene expression analysis using the DR5:uidA reporter construct further revealed that ET affects auxin signaling through the central regulator CTR1 and that fungal-derived compounds, such as indole-3-acetic acid and indole-3-acetaldehyde, induce MPK6 activity. Our results suggest that T. atroviride likely alters root-system architecture modulating MPK6 activity and ET and auxin action.

Trichoderma spp. Improve growth of Arabidopsis seedlings under salt stress through enhanced root development, osmolite production, and Na⁺ elimination through root exudates.

Salt stress is an important constraint to world agriculture. Here, we report on the potential of Trichoderma virens and T. atroviride to induce tolerance to salt in Arabidopsis seedlings. We first characterized the effect of several salt concentrations on shoot biomass production and root architecture of Arabidopsis seedlings. We found that salt repressed plant growth and root development in a dose-dependent manner by blocking auxin signaling. Analysis of the wild type and eir1, aux1-7, arf7arf19, and tir1abf2abf19 auxin-related mutants revealed a key role for indole-3-acetic acid (IAA) signaling in mediating salt tolerance. We also found that T. virens (Tv29.8) and T. atroviride (IMI 206040) promoted plant growth in both normal and saline conditions, which was related to the induction of lateral roots and root hairs through auxin signaling. Arabidopsis seedlings grown under saline conditions inoculated with Trichoderma spp. showed increased levels of abscissic acid, L-proline, and ascorbic acid, and enhanced elimination of Na⁺ through root exudates. Our data show the critical role of auxin signaling and root architecture to salt tolerance in Arabidopsis and suggest that these fungi may enhance the plant IAA level as well as the antioxidant and osmoprotective status of plants under salt stress.

Trichoderma-induced plant immunity likely involves both hormonal- and camalexin-dependent mechanisms in Arabidopsis thaliana and confers resistance against necrotrophic fungi Botrytis cinerea.

Filamentous fungi belonging to the genus Trichoderma have long been recognized as agents for the biocontrol of plant diseases. In this work, we investigated the mechanisms involved in the defense responses of Arabidopsis thaliana seedlings elicited by co-culture with Trichoderma virens and Trichoderma atroviride. Interaction of plant roots with fungal mycelium induced growth and defense responses, indicating that both processes are not inherently antagonist. Expression studies of the pathogenesis-related reporter markers pPr1a:uidA and pLox2:uidA in response to T. virens or T. atroviride provided evidence that the defense signaling pathway activated by these fungi involves salicylic acid (SA) and/or jasmonic acid (JA) depending on the amount of conidia inoculated. Moreover, we found that Arabidopsis seedlings colonized by Trichoderma accumulated hydrogen peroxide and camalexin in leaves. When grown under axenic conditions, T. virens produced indole-3-carboxaldehyde (ICAld) a tryptophan-derived compound with activity in plant development. In Arabidopsis seedlings whose roots are in contact with T. virens or T. atroviride, and challenged with Botrytis cinerea in leaves, disease severity was significantly reduced compared to axenically grown seedlings. Our results indicate that the defense responses elicited by Trichoderma in Arabidopsis are complex and involve the canonical defense hormones SA and JA as well as camalexin, which may be important factors in boosting plant immunity.

The role of microbial signals in plant growth and development.

Plant growth and development involves a tight coordination of the spatial and temporal organization of cell division, cell expansion and cell differentiation. Orchestration of these events requires the exchange of signaling molecules between the root and shoot, which can be affected by both biotic and abiotic factors. The interactions that occur between plants and their associated microorganisms have long been of interest, as knowledge of these processes could lead to the development of novel agricultural applications. Plants produce a wide range of organic compounds including sugars, organic acids and vitamins, which can be used as nutrients or signals by microbial populations. On the other hand, microorganisms release phytohormones, small molecules or volatile compounds, which may act directly or indirectly to activate plant immunity or regulate plant growth and morphogenesis. In this review, we focus on recent developments in the identification of signals from free-living bacteria and fungi that interact with plants in a beneficial way. Evidence has accumulated indicating that classic plant signals such as auxins and cytokinins can be produced by microorganisms to efficiently colonize the root and modulate root system architecture. Other classes of signals, including N-acyl-L-homoserine lactones, which are used by bacteria for cell-to-cell communication, can be perceived by plants to modulate gene expression, metabolism and growth. Finally, we discuss the role played by volatile organic compounds released by certain plant growth-promoting rhizobacteria in plant immunity and developmental processes. The picture that emerges is one in which plants and microbes communicate themselves through transkingdom signaling systems involving classic and novel signals.

Trichoderma virens, a plant beneficial fungus, enhances biomass production and promotes lateral root growth through an auxin-dependent mechanism in Arabidopsis.

Trichoderma species belong to a class of free-living fungi beneficial to plants that are common in the rhizosphere. We investigated the role of auxin in regulating the growth and development of Arabidopsis (Arabidopsis thaliana) seedlings in response to inoculation with Trichoderma virens and Trichoderma atroviride by developing a plant-fungus interaction system. Wild-type Arabidopsis seedlings inoculated with either T. virens or T. atroviride showed characteristic auxin-related phenotypes, including increased biomass production and stimulated lateral root development. Mutations in genes involved in auxin transport or signaling, AUX1, BIG, EIR1, and AXR1, were found to reduce the growth-promoting and root developmental effects of T. virens inoculation. When grown under axenic conditions, T. virens produced the auxin-related compounds indole-3-acetic acid, indole-3-acetaldehyde, and indole-3-ethanol. A comparative analysis of all three indolic compounds provided detailed information about the structure-activity relationship based on their efficacy at modulating root system architecture, activation of auxin-regulated gene expression, and rescue of the root hair-defective phenotype of the rhd6 auxin response Arabidopsis mutant. Our results highlight the important role of auxin signaling for plant growth promotion by T. virens.