Differential Response of Tomato Plants to the Application of Three Trichoderma Species When Evaluating the Control of Pseudomonas syringae Populations

Trichoderma species are well known biocontrol agents that are able to induce responses in the host plants against an array of abiotic and biotic stresses. Here, we investigate, when applied to tomato seeds, the potential of Trichoderma strains belonging to three different species, T. parareesei T6, T. asperellum T25, and T. harzianum T34, to control the fully pathogenic strain Pseudomonas syringae pv. tomato (Pst) DC3000, able to produce the coronatine (COR) toxin, and the COR-deficient strain Pst DC3118 in tomato plants, and the molecular mechanisms by which the plant can modulate its systemic defense. Four-week old tomato plants, seed-inoculated, or not, with a Trichoderma strain, were infected, or not, with a Pst strain, and the changes in the expression of nine marker genes representative of salicylic acid (SA) (ICS1 and PAL5) and jasmonic acid (JA) (TomLoxC) biosynthesis, SA- (PR1b1), JA- (PINII and MYC2) and JA/Ethylene (ET)-dependent (ERF-A2) defense pathways, as well as the abscisic acid (ABA)-responsive gene AREB2 and the respiratory burst oxidase gene LERBOH1, were analyzed at 72 hours post-inoculation (hpi) with the bacteria. The significant increase obtained for bacterial population sizes in the leaves, disease index, and the upregulation of tomato genes related to SA, JA, ET and ABA in plants inoculated with Pst DC3000 compared with those obtained with Pst DC3118, confirmed the COR role as a virulence factor, and showed that both Pst and COR synergistically activate the JA- and SA-signaling defense responses, at least at 72 hpi. The three Trichoderma strains tested reduced the DC3118 levels to different extents and were able to control disease symptoms at the same rate. However, a minor protection (9.4%) against DC3000 was only achieved with T. asperellum T25. The gene deregulation detected in Trichoderma-treated plus Pst-inoculated tomato plants illustrates the complex system of a phytohormone-mediated signaling network that is affected by the pathogen and Trichoderma applications but also by their interaction. The expression changes for all nine genes analyzed, excepting LERBOH1, as well as the bacterial populations in the leaves were significantly affected by the interaction. Our results show that Trichoderma spp. are not adequate to control the disease caused by fully pathogenic Pst strains in tomato plants.

Download Full-text

Pseudomonas syringae pv. tomato OxyR Is Required for Virulence in Tomato and Arabidopsis

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-09-15-0204-r ◽

2016 ◽

Vol 29 (2) ◽

pp. 119-131 ◽

Cited By ~ 17

Author(s):

Yasuhiro Ishiga ◽

Yuki Ichinose

Keyword(s):

Oxidative Stress ◽

Pseudomonas Syringae ◽

Molecular Mechanisms ◽

Expression Profiles ◽

High Sensitivity ◽

Defense Responses ◽

Bacterial Populations ◽

Disease Symptoms ◽

Expression Of Genes ◽

Genes Encoding

Reactive oxygen species (ROS) have been shown to have a crucial role in plant defense responses and signaling pathways. In addition, ROS also have direct toxicity against pathogens. However, the molecular mechanisms of plant ROS in the direct effects against pathogens is still unclear. To investigate the function of plant ROS in the interactions of plant and bacterial pathogens, we focused on oxyR, encoding an oxidative stress-regulated transcription factor in Pseudomonas syringae pv. tomato DC3000 (DC3000), and generated an ΔoxyR mutant. The DC3000 ΔoxyR mutant showed high sensitivity to oxidative stress in comparison with wild type and the complemented line. The host plants of DC3000, including tomato and Arabidopsis inoculated with the ΔoxyR mutant, clearly showed reduced disease symptoms as well as reduced bacterial populations. Expression profiles of DC3000 genes revealed that OxyR could regulate the expression of genes encoding ROS-detoxifying enzymes, including catalases (KatB and KatG), in response to ROS. We also demonstrated that the expression of katB could be regulated by OxyR during the infection of DC3000 in Arabidopsis. These results suggest that OxyR has an important role in the virulence of DC3000 by regulating the expression of genes related to oxidative stress.

Download Full-text

C-Terminal Region of Plant Ferredoxin-Like Protein Is Required to Enhance Resistance to Bacterial Disease in Arabidopsis thaliana

Phytopathology ◽

10.1094/phyto-08-10-0220 ◽

2011 ◽

Vol 101 (6) ◽

pp. 741-749 ◽

Cited By ~ 8

Author(s):

Yi-Hsien Lin ◽

Hsiang-En Huang ◽

Yen-Ru Chen ◽

Pei-Luan Liao ◽

Ching-Lian Chen ◽

...

Keyword(s):

Disease Resistance ◽

Transgenic Plants ◽

Nadph Oxidase ◽

Pseudomonas Syringae ◽

Induced Defense ◽

Defense Responses ◽

Bacterial Disease ◽

Oxidase Gene ◽

Induced Defense Responses ◽

Resistance To Ralstonia Solanacearum

Protein phosphorylation is an important biological process associated with elicitor-induced defense responses in plants. In a previous report, we described how plant ferredoxin-like protein (PFLP) in transgenic plants enhances resistance to bacterial pathogens associated with the hypersensitive response (HR). PFLP possesses a putative casein kinase II phosphorylation (CK2P) site at the C-terminal in which phosphorylation occurs rapidly during defense response. However, the contribution of this site to the enhancement of disease resistance and the intensity of HR has not been clearly demonstrated. In this study, we generated two versions of truncated PFLP, PEC (extant CK2P site) and PDC (deleted CK2P site), and assessed their ability to trigger HR through harpin (HrpZ) derived from Pseudomonas syringae as well as their resistance to Ralstonia solanacearum. In an infiltration assay of HrpZ, PEC intensified harpin-mediated HR; however, PDC negated this effect. Transgenic plants expressing these versions indicate that nonphosphorylated PFLP loses its ability to induce HR or enhance disease resistance against R. solanacearum. Interestingly, the CK2P site of PFLP is required to induce the expression of the NADPH oxidase gene, AtrbohD, which is a reactive oxygen species producing enzyme. This was further confirmed by evaluating the HR on NADPH oxidase in mutants of Arabidopsis. As a result, we have concluded that the CK2P site is required for the phosphorylation of PFLP to enhance disease resistance.

Download Full-text

The Internal Glycine-Rich Motif and Cysteine Suppress Several Effects of the HpaGXooc Protein in Plants

Phytopathology ◽

10.1094/phyto-96-1052 ◽

2006 ◽

Vol 96 (10) ◽

pp. 1052-1059 ◽

Cited By ~ 37

Author(s):

Fengquan Liu ◽

Hongxia Liu ◽

Qin Jia ◽

Xiaojing Wu ◽

Xiaojing Guo ◽

...

Keyword(s):

Plant Growth ◽

Pseudomonas Syringae ◽

Growth Promotion ◽

Marker Genes ◽

Pathogen Defense ◽

Hypersensitive Cell Death ◽

Tobacco Plants ◽

Tomato Plants ◽

Aerial Parts ◽

Stimulate Plant Growth

HpaGXooc, produced by Xanthomonas oryzae pv. oryzicola, is a member of harpin group of proteins that stimulate plant growth, hypersensitive cell death (HCD), and pathogen defense. The protein contains two copies of the glycine-rich motif (GRM), a characteristic of harpins, and a cysteine, which is absent in other harpins. Genetic modification generated the pro-tein mutants HpaGXoocMG (MG) by deleting GRMs and HpaGXoocC47T (C47T) by replacing cysteine with threonine. When applied to tobacco plants, C47T and MG were 1.2- and 1.7-fold stronger, respectively, than HpaGXooc in inducing HCD, which occurred consistently with expression of the marker genes hin1 and hsr203. The proteins markedly alleviated infection of tobacco by Tobacco mosaic virus and Arabidopsis and tomato by Pseudomonas syringae. Treating tobacco plants with HpaGXooc, C47T, and MG decreased the viral infection by 58, 81, and 92%, respectively. In Arabidopsis and tomato plants treated with HpaGXooc, C47T, or MG, P. syringae multiplication was inhibited; bacterial population multiplied in 5 days in these plants were ca. 160-, 1,260-, or 15,860-fold smaller than that in control plants. So pathogen defense was induced in both plants. Defense-related genes Chia5, NPR1, and PR-1a were expressed consistently with resistance. In response to HpaGXooc, C47T, and MG, aerial parts and roots of tomato plants increased growth by 15 and 53%, 25 and 77%, and 46 and 106%, relative to controls. The expansin gene, EXP2, involved in the cell expansion and plant growth was expressed coordinately with plant growth promotion. These results suggest that the presence of GRM and cysteine in HpaGXooc represses the effects of the protein in plants.

Download Full-text

Sm1, a Proteinaceous Elicitor Secreted by the Biocontrol Fungus Trichoderma virens Induces Plant Defense Responses and Systemic Resistance

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-19-0838 ◽

2006 ◽

Vol 19 (8) ◽

pp. 838-853 ◽

Cited By ~ 217

Author(s):

Slavica Djonović ◽

Maria J. Pozo ◽

Lawrence J. Dangott ◽

Charles R. Howell ◽

Charles M. Kenerley

Keyword(s):

Plant Defense ◽

Defense Mechanisms ◽

Molecular Mechanisms ◽

Pathogenic Fungi ◽

Defense Responses ◽

Systemic Resistance ◽

Trichoderma Virens ◽

Trichoderma Spp ◽

Toxic Activity ◽

Small Protein

The soilborne filamentous fungus Trichoderma virens is a biocontrol agent with a well-known ability to produce antibiotics, parasitize pathogenic fungi, and induce systemic resistance in plants. Even though a plant-mediated response has been confirmed as a component of bioprotection by Trichoderma spp., the molecular mechanisms involved remain largely unknown. Here, we report the identification, purification, and characterization of an elicitor secreted by T. virens, a small protein designated Sm1 (small protein 1). Sm1 lacks toxic activity against plants and microbes. Instead, native, purified Sm1 triggers production of reactive oxygen species in monocot and dicot seedlings, rice, and cotton, and induces the expression of defense-related genes both locally and systemically in cotton. Gene expression analysis revealed that SM1 is expressed throughout fungal development under different nutrient conditions and in the presence of a host plant. Using an axenic hydroponic system, we show that SM1 expression and secretion of the protein is significantly higher in the presence of the plant. Pretreatment of cotton cotyledons with Sm1 provided high levels of protection to the foliar pathogen Colletotrichum sp. These results indicate that Sm1 is involved in the induction of resistance by Trichoderma spp. through the activation of plant defense mechanisms.

Download Full-text

Molecular characterization of differences between the tomato immune receptors Fls3 and Fls2

10.1101/2020.02.04.934133 ◽

2020 ◽

Author(s):

Robyn Roberts ◽

Alexander E. Liu ◽

Lingwei Wan ◽

Annie M. Geiger ◽

Sarah R. Hind ◽

...

Keyword(s):

Pseudomonas Syringae ◽

Molecular Mechanisms ◽

Bacterial Pathogen ◽

Transcript Abundance ◽

Defense Responses ◽

Structural Features ◽

Host Responses ◽

Pathogen Invasion

AbstractPlants mount defense responses by recognizing indications of pathogen invasion, including microbe-associated molecular patterns (MAMPs). Flagellin from the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) contains two MAMPs, flg22 and flgII-28, that are recognized by tomato receptors Flagellin sensing 2 (Fls2) and Flagellin sensing 3 (Fls3), respectively. It is unknown to what degree each receptor contributes to immunity and if they promote immune responses using the same molecular mechanisms. Characterization of CRISPR/Cas9-generated Fls2 and Fls3 tomato mutants revealed that the two receptors contribute equally to disease resistance both on the leaf surface and in the apoplast. However, striking differences were observed in certain host responses mediated by the two receptors. Compared to Fls2, Fls3 mediated a more sustained production of reactive oxygen species (ROS) and an increase in transcript abundance of 44 tomato genes, with two genes serving as reporters for Fls3. Fls3 had greater in vitro kinase activity and interacted differently with the Pst effector AvrPtoB as compared to Fls2. Using chimeric Fls2/Fls3 proteins, we found that no receptor domain was solely responsible for the Fls3 sustained ROS, suggesting involvement of multiple structural features. This work reveals differences in the immunity outputs between Fls2 and Fls3, suggesting they use distinct molecular mechanisms to activate pattern-triggered immunity in response to flagellin-derived MAMPs.

Download Full-text

molecular link from PAMP perception to a MAPK cascade associated with tomato disease resistance

10.32747/2012.7597918.bard ◽

2012 ◽

Author(s):

Guido Sessa ◽

Gregory B. Martin

Keyword(s):

Cell Death ◽

Disease Resistance ◽

Immune Responses ◽

Map Kinase ◽

Pseudomonas Syringae ◽

Molecular Mechanisms ◽

Plant Immunity ◽

Defense Responses ◽

Research Activities ◽

Mapk Cascades

The research problem: The detection of pathogen-associated molecular patterns (PAMPs) by plant pattern recognition receptors (PRRs) is a key mechanism by which plants activate an effective immune response against pathogen attack. MAPK cascades are important signaling components downstream of PRRs that transduce the PAMP signal to activate various defense responses. Preliminary experiments suggested that the receptor-like cytoplasmickinase (RLCK) Mai5 plays a positive role in pattern-triggered immunity (PTI) and interacts with the MAPKKK M3Kε. We thus hypothesized that Mai5, as other RLCKs, functions as a component PRR complexes and acts as a molecular link between PAMP perception and activation of MAPK cascades. Original goals: The central goal of this research was to investigate the molecular mechanisms by which Mai5 and M3Kε regulate plant immunity. Specific objectives were to: 1. Determine the spectrum of PAMPs whose perception is transmitted by M3Kε; 2. Identify plant proteins that act downstream of M3Kε to mediate PTI; 3. Investigate how and where Mai5 interacts with M3Kε in the plant cell; 4. Examine the mechanism by which Mai5 contributes to PTI. Changes in research directions: We did not find convincing evidence for the involvement of M3Kε in PTI signaling and substituted objectives 1 and 3 with research activities aimed at the analysis of transcriptomic profiles of tomato plants during the onset of plant immunity, isolation of the novel tomato PRR FLS3, and investigation of the involvement of the RLCKBSKs in PTI. Main achievements during this research program are in the following major areas: 1. Functional characterization of Mai5. The function of Mai5 in PTI signaling was demonstrated by testing the effect of silencing the Mai5 gene by virus-induced gene silencing (VIGS) experiments and in cell death assays. Domains of Mai5 that interact with MAPKKKs and subcellular localization of Mai5 were analyzed in detail. 2. Analysis of transcriptional profiles during the tomato immune responses to Pseudomonas syringae (Pombo et al., 2014). We identified tomato genes whose expression is induced specifically in PTI or in effector-triggered immunity (ETI). Thirty ETI-specific genes were examined by VIGS for their involvement in immunity and the MAPKKK EPK1, was found to be required for ETI. 3. Dissection of MAP kinase cascades downstream of M3Kε (Oh et al., 2013; Teper et al., 2015). We identified genes that encode positive (SGT and EDS1) and negative (WRKY1 and WRKY2) regulators of the ETI-associated cell death mediated by M3Kε. In addition, the MKK2 MAPKK, which acts downstream of M3Kε, was found to interact with the MPK3 MAPK and specific MPK3 amino acids involved interaction were identified and found to be required for induction of cell death. We also identified 5 type III effectors of the bacterial pathogen Xanthomonaseuvesicatoria that inhibited cell death induced by components of ETI-associated MAP kinase cascades. 4. Isolation of the tomato PRR FLS3 (Hind et al., submitted). FLS3, a novel PRR of the LRR-RLK family that specifically recognizes the flagellinepitope flgII-28 was isolated. FLS3 was shown to bind flgII-28, to require kinase activity for function, to act in concert with BAK1, and to enhance disease resistance to Pseudomonas syringae. 5. Functional analysis of RLCKs of the brassinosteroid signaling kinase (BSK) family.Arabidopsis and tomato BSKs were found to interact with PRRs. In addition, certain ArabidospsisBSK mutants were found to be impaired in PAMP-induced resistance to Pseudomonas syringae. Scientific and agricultural significance: Our research activities discovered and characterized new molecular components of signaling pathways mediating recognition of invading pathogens and activation of immune responses against them. Increased understanding of molecular mechanisms of immunity will allow them to be manipulated by both molecular breeding and genetic engineering to produce plants with enhanced natural defense against disease.

Download Full-text

Trichoderma harzianum Strain T22 Modulates Direct Defense of Tomato Plants in Response to Nezara viridula Feeding Activity

Journal of Chemical Ecology ◽

10.1007/s10886-021-01260-3 ◽

2021 ◽

Author(s):

Tuğcan Alınç ◽

Antonino Cusumano ◽

Ezio Peri ◽

Livio Torta ◽

Stefano Colazza

Keyword(s):

Growth Rate ◽

Plant Defense ◽

Trichoderma Harzianum ◽

Defense Responses ◽

Nezara Viridula ◽

Marker Genes ◽

Stink Bug ◽

Tomato Plants ◽

Southern Green Stink Bug ◽

The Impact

AbstractPlant growth-promoting fungi belonging to genus Trichoderma are known to help plants when dealing with biotic stressors by enhancing plant defenses. While beneficial effects of Trichoderma spp. against plant pathogens have long been documented, fewer studies have investigated their effect on insect pests. Here, we studied the impact of Trichoderma root colonization on the plant defense responses against stink bug feeding attack. For this purpose, a model system consisting of tomato plant, Solanum lycopersicum cv Dwarf San Marzano, Trichoderma harzianum strain T22 and the southern green stink bug, Nezara viridula, was used. We firstly determined stink bug performance in terms of relative growth rate and survival on tomato plants inoculated by T. harzianum T22. Then, we evaluated relative expression of plant defense-related genes on inoculated plants induced by stink bug feeding. We found evidence that T. harzianum T22 affects tomato defense responses against N. viridula nymphs leading to reduction of growth rate. Our results also showed that T. harzianum T22 enhances plant direct defenses by an early increase of transcript levels of jasmonic acid marker genes. Yet this effect was time-dependent and only detected 8 h after herbivore induction. Taken together, our findings provide better understanding on the mechanisms underlying tomato induced resistance against herbivorous stink bugs.

Download Full-text

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

Frontiers in Plant Science ◽

10.3389/fpls.2021.678830 ◽

2021 ◽

Vol 12 ◽

Author(s):

Ilaria Di Lelio ◽

Mariangela Coppola ◽

Ernesto Comite ◽

Donata Molisso ◽

Matteo Lorito ◽

...

Keyword(s):

Plant Defense ◽

Environmental Conditions ◽

Insect Pests ◽

Plant Protection ◽

Growth Promotion ◽

Defense Responses ◽

Macrosiphum Euphorbiae ◽

Trichoderma Species ◽

Tomato Plants ◽

The Impact

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.

Download Full-text

The Phytotoxin Coronatine Contributes to Pathogen Fitness and Is Required for Suppression of Salicylic Acid Accumulation in Tomato Inoculated with Pseudomonas syringae pv. tomato DC3000

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-20-8-0955 ◽

2007 ◽

Vol 20 (8) ◽

pp. 955-965 ◽

Cited By ~ 149

Author(s):

Srinivasa Rao Uppalapati ◽

Yasuhiro Ishiga ◽

Tamding Wangdi ◽

Barbara N. Kunkel ◽

Ajith Anand ◽

...

Keyword(s):

Salicylic Acid ◽

Pseudomonas Syringae ◽

Defense Responses ◽

Tomato Dc3000 ◽

Defense Proteins ◽

Tomato Plants ◽

Expression Of Genes ◽

Genes Encoding ◽

Salicylic Acid Accumulation ◽

Pathogen Fitness

The roles of the phytotoxin coronatine (COR) and salicylic acid (SA)-mediated defenses in the interaction of Pseudomonas syringae pv. tomato DC3000 and tomato (Solanum lycopersicum) were investigated. Unlike findings reported for Arabidopsis thaliana, DC3000 mutants impaired for production of COR or one of its components, coronafacic acid (CFA) or coronamic acid (CMA), induced distinctly different disease lesion phenotypes in tomato. Tomato plants inoculated with the CFA- CMA- mutant DB29 showed elevated transcript levels of SlICS, which encodes isochorismate synthase, an enzyme involved in SA biosynthesis in S. lycopersicum. Furthermore, expression of genes encoding SA-mediated defense proteins were elevated in DB29-inoculated plants compared with plants inoculated with DC3000, suggesting that COR suppresses SlICS-mediated SA responses. Sequence analysis of SlICS revealed that it encodes a protein that is 55 and 59.6% identical to the A. thaliana ICS-encoded proteins AtICS1 and AtICS2, respectively. Tomato plants silenced for SlICS were hypersusceptible to DC3000 and accumulated lower levels of SA after infection with DC3000 compared with inoculated wild-type tomato plants. Unlike what has been shown for A. thaliana, the COR- mutant DB29 was impaired for persistence in SlICS-silenced tomato plants; thus, COR has additional roles in virulence that are SA independent and important in the latter stages of disease development. In summary, the infection assays, metabolic profiling, and gene expression results described in this study indicate that the intact COR molecule is required for both suppression of SA-mediated defense responses and full disease symptom development in tomato.

Download Full-text

Molecular defense responses of apple genotypes in compatible and incompatible interactions with Erwinia amylovora

Plant Protection Science ◽

10.17221/10339-pps ◽

2002 ◽

Vol 38 (SI 1 - 6th Conf EFPP 2002) ◽

pp. S139-S140

Author(s):

M.N. Brisset ◽

J.S. Venisse ◽

J.P. Paulin

Keyword(s):

Pseudomonas Syringae ◽

Type Strain ◽

Fire Blight ◽

Molecular Mechanisms ◽

Erwinia Amylovora ◽

Wild Type Strain ◽

Defense Responses ◽

Bacterial Disease ◽

Wild Type ◽

Susceptible Host

Erwinia amylovora is the causal agent of fire blight, a bacterial disease of apple and pear. Pathogenicity determinants of the bacteria are identified (hrp-dsp cluster, capsule, siderophore) but molecular mechanisms leading to susceptibility or resistance of the plant are not yet understood. To address this question, we challenged two genotypes of apple, known for their contrasting susceptibility to fire blight, with a wild-type strain of E. amylovora (Ea wt), an avirulent hrp mutant of this bacteria (Ea hrp) or a wild-type strain of the incompatible pathogen Pseudomonas syringae pv. tabaci (Pst wt). Mechanisms usually related to resistance responses were investigated i.e. oxidative stress, accumulation of PR-proteins and induction of genes encoding various enzymes of the phenylpropanoid pathway. Results showed two kinds of responses (i) some mechanisms were elicited in both susceptible and resistant genotypes by Ea wt and Pst wt with similar kinetics and not induced by Ea hrp, (ii) others were specifically repressed by Ea wt in its susceptible host, when induced by Pst wt and Ea hrp. These results suggest several hypothesis about the cross-talk between E. amylovora and its host plants.

Download Full-text